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297 publications mentioning hsa-mir-93 (showing top 100)

Open access articles that are associated with the species Homo sapiens and mention the gene name mir-93. Click the [+] symbols to view sentences that include the gene name, or the word cloud on the right for a summary.

1
[+] score: 497
To determine the molecular mechanisms of mir-93 CSC regulation, we employed an unbiased approach assessing the effect of mir-93 expression on early changes in global gene expression profile coupled with prediction of miRNA target sequences. [score:8]
Of the 2,000 genes downregulated at least two-fold upon DOX treatment in the ALDH [+] population (Table S1), 127 overlapped with the predicted target sequences of mir-93 including twenty-four genes known to be involved in stem cell regulation (Figure 4A and Table S2) including JAK1, SOX4, STAT3, AKT, E2H1 and HMGAZ. [score:7]
Continued expression of mir-93 simultaneously downregulates a number of stem cell self-renewal pathways including JAK/STAT, AKT, EZH1 and HMGH2, promoting cellular differentiation and depleting the CSC population. [score:6]
In contrast, only 352 genes were significantly downregulated by DOX in the ALDH [−] population (Table S3) with twelve of these genes (no stem cell genes) overlapping with the predicted mir-93 targets. [score:6]
To determine the relationship between mir-93 expression and cell cycle kinetics, we assessed mir-93 expression in quiescent and cycling ALDH [+] and ALDH [−] populations. [score:5]
To determine the effect of mir-93 expression on tumor invasion, we examined the effect of mir-93 induction and/or downregulation on invasion of SUM159 cells using a matrigel invasion assay. [score:5]
To determine the relationship between mir-93 expression and tumor initiating capacity, we constructed a mir-93 sensor tagged with GFP (mir-93-sensor-GFP) containing a mir-93 target UTR coupled to GFP. [score:5]
Interestingly, expression of mir-93 significantly reduced expression of the mRNA for TGFβR2 in both ALDH [+] and ALDH [−] SUM159 cells. [score:5]
Expression of mir-93 in SUM159 cells causes them to assume a more epithelial appearance associated with a decrease in Vimentin and an increase in membrane localized E-Cadherin expression. [score:5]
miRNA93 (mir-93) is frequently overexpressed in human cancer but, paradoxically, has been found to function as a tumor suppressor in some contexts. [score:5]
The mir-93 miRNA target sequence was engineered into the 3′ untranslated region (UTR) of the cDNA encoding for the GFP fluorescent protein. [score:5]
These experiments suggested that unlike cytotoxic agents which primarily target the bulk cell population, mir-93 overexpression was able to reduce the CSC population. [score:5]
In less differentiated tumors, enforced expression of mir-93 completely blocks tumor development in mammary fat pads and development of metastases following intracardiac injection in mouse xenografts by reducing breast cancer stem cells. [score:5]
Expression of this construct in cells that express the mir-93 miRNA results in a RNAi pathway -dependent degradation of GFP mRNA and thus no green fluorescence. [score:5]
In cells transfected with this vector, mir-93 expression results in degradation of GFP mRNA (sensor -negative), whereas mir-93 -negative cells express GFP (sensor -positive) (Figure 1B). [score:5]
These studies suggest that mir-93 regulates the CSC population by simultaneously targeting a number of stem cell regulatory genes. [score:5]
mir-93 also inhibits TGFβ signaling by targeting TGFβR2, an effect seen within twelve hours of mir-93 induction. [score:5]
mir-93 downregulates stem cell regulatory genes in BCSCs. [score:5]
mir-93 overexpression inhibits the growth of established tumor xenografts. [score:5]
To confirm this, we utilized a luceriferase reporter assay to determine the effect of mir-93 on the expression of the stem cell regulatory genes AKT3, SOX4 and STAT3 selected from the expression profiling data. [score:5]
To confirm and extend these results we determined the effect of mir-93 expression on mRNA expression of a wider panel of epithelial and mesenchymal markers. [score:5]
Overlap between mir93 predicted targets from TargetScan 5.1 and profiling data from DOX -treated cells (DOX) to non-DOX -treated cells (CTRL) in the ALDH [−] population (12 genes) or in the ALDH [+] population (352 genes). [score:5]
Expression of mir-93 in these cells caused them to assume a more epithelial appearance associated with a decrease in Vimentin and an increase in E-Cadherin expression (Figure 6A). [score:5]
As shown in Figure 2G, mir-93 induction completely suppressed whereas docetaxel partially suppressed metastasis formation. [score:5]
Furthermore, overexpression of mir-93 in freshly isolated normal breast cells or in immortalized non-transformed MCF-10A cells increased the proportion of cells expressing EpCAM (Figure 7B, 7C). [score:5]
To simulate the effects of CSC targeting agents in advanced disease, tumors were inoculated into mammary fatpads and when the tumors were palpable mir-93 was induced by addition of doxycycline to the mouse drinking water. [score:5]
Expression of mir-93 in SUM159 cells resulted in a time dependent decrease in expression of mesenchymal markers, Vimentin, N-cadherin and Twist, and an increase in the epithelial markers E-Cadherin and Claudin (Figure 6B). [score:5]
In order to determine whether down-regulation of mir-93 promoted tumorigenesis, we utilized a mirZip anti-sense miRNA in SUM159 cells. [score:4]
mir-93 expression was also associated with and in turn regulates cellular proliferation. [score:4]
To determine the relationship between mir-93 expression and level of cellular differentiation, we compared the expression of mir-93 in claudin [low] (SUM159), basal (HCC1954) and luminal (MCF7) cells. [score:4]
com/RNAi) was used to generate mir-93 -expressing lentiviruses; and mirZIP-lentivector (SBI, Mountain View, CA) was used to generate mir-93-knockdown lentiviruses in UM Vector Core Facility. [score:4]
Overexpression of mir-93 significantly inhibited the ability of SUM159 cells to invade in this assay (Figure S8E). [score:4]
For construction of the mir-93 sensor, miRNA-complementary oligonucleotides were annealed and cloned into a Marx vector that directs GFP expression. [score:4]
A. Schematic representation of the experimental design to identify the direct targets of mir-93 in SUM159 cells. [score:4]
Expression of mir-93 reduced the level of these stem cell regulatory genes in SUM159 (Figure 4B) and HCC1954 cells (Figure S18) but not in luminal MCF7 and MDA-MB-453 cells (Figure S19). [score:4]
The downregulation of these genes in pTRIPZ-SUM159-mir-93, pTRIPZ-HCC1954-mir-93 cell lines and pTRIPZ-MC1-mir-93 were confirmed with customized PCR array plates (Figures S15, S16, S17). [score:4]
Concomitantly, mir-93 and its related miRNA cluster is co-synthesized which promotes further proliferation while simultaneously downregulating TGFβ signaling. [score:4]
We utilized a DOX inducible system to determine the effects of enforced mir-93 expression on the CSC populations assessed by expression of the stem cell markers ALDH and CD24 [−]CD44 [+] as well as by mouse xenograft assays [14], [22]. [score:4]
1002751.g004 Figure 4 A. Schematic representation of the experimental design to identify the direct targets of mir-93 in SUM159 cells. [score:4]
mir-93 targets stem cell regulatory genes. [score:4]
Among these three cell lines, mir93 expression level is highest in MCF7 cells and lowest in the SM159 cells. [score:3]
However, the effect of mir-93 on the cancer stem cell population is dependent on the cellular differentiation state, with mir-93 expression increasing the cancer stem cell population in more differentiated breast tumors. [score:3]
The effects of mir-93 expression on tumor initiating capacity was confirmed using two primary breast xenografts generated without in vitro culture. [score:3]
mir-93 overexpression decreases CSCs in vitro. [score:3]
The mir106b-25 cluster is composed of the highly conserved miRNA106b (mir-106b), miRNA93 (mir-93) and miRNA25 (mir-25) that have been reported to be overexpressed in a number of cancers including gastric, prostate and pancreatic neural endocrine tumors, neuroblastoma and multiple myeloma [1], [2], [3]. [score:3]
mir-93 overexpression decreases CSCs in vitro We utilized a tetracycline (TET) inducible mir-93 construct tagged with RFP (pTRIPZ-mir-93-RFP) to determine the functional role of mir-93 in CSCs. [score:3]
Figure S17 Validation of the 127 overlapped gene expression with customerized StellArray PCR array plate in pTRIPZ-MC1-mir93. [score:3]
B. A schematic of mir-93-Sensor-GFP lentiviral construct; C. SUM159 cells were transduced with the mir-93-sensor-GFP lentivirus and selected with hygromycin B, and cells were sorted based on the GFP expression. [score:3]
These studies demonstrated that in these breast cancer cell lines low mir-93 expression is associated with the CSC phenotype characterized by increased aldehyde dehydrogenase expression, tumor initiating capacity and the ability to generate heterogeneous tumors containing both stem cell and non-stem cell populations. [score:3]
Figure S11Endogenous mir93 expression levels parallel cell differentiation state. [score:3]
Induction of mir-93 reduced the quiescent cell population from 64% to 42% suggesting that this miRNA has the capacity to directly regulate the cell cycle (Figure 5B). [score:3]
Tumor initiating capacity is associated with low mir-93 expression. [score:3]
In contrast, CD24 [−]CD44 [+] in MCF7 cells showed no difference for mir93 expression level in comparison to the bulk population. [score:3]
Figure S9 mir-93 inhibits tumor metastasis in SUM159 cells. [score:3]
A. ALDH -positive cells from HCC1954 cells shows lower mir93 expression level in comparison to ALDH -negative cells by qRT-PCR. [score:3]
Interestingly, we found that the highest level of mir-93 is expressed in the EpCAM [+]CD49f [+] population (Figure 7A), which suggested mir-93 was required to maintain the cells as EpCAM [+]CD49f [+]. [score:3]
mir-93 overexpression increases the CSC population and accelerates tumor growth in luminal subtype MCF7 cells. [score:3]
Interestingly, this analysis revealed that twenty-four genes known to be involved in stem cell self-renewal including JAK1, SOX4, STAT3, AKT, EZH1, HMGA2 are targeted by mir-93. [score:3]
Figure S21 Validation of the 127 overlapped gene expression with customerized StellArray PCR array plate in pTRIPZ-MCF7-mir93. [score:3]
In contrast, overexpresson of mir-93 in the luminal MCF7 cells line resulted in an increase in CD24 [−]CD44 [+] CSC resulting in increased tumor growth. [score:3]
We determined the effects of mir-93 expression on MET of SUM159 cells by assessing markers of these states at the protein and mRNA levels. [score:3]
The relative increase in the Aldefluor -positive population seen with docetaxel treatment was abrogated by simultaneous mir-93 expression (Figure 2B). [score:3]
ALDH [+] cells from SUM159 cells shows lower mir-93 expression level in comparison to ALDH [−] cells as accessed by qRT-PCR. [score:3]
In contrast, in cells with repressed mir-93 miRNA, the GFP mRNA is not degraded and resulting in expression of the fluorescent GFP. [score:3]
The relationship between mir-93 expression and tumor initiation was determined by introducing serial dilutions of sensor -positive (mir-93 -negative) and sensor -negative (mir-93 -positive) SUM159 cells into the mammary fatpads of NOD/SCID mice. [score:3]
In order to determine the cellular targets of mir-93 in BCSCs, ALDH [+] and ALDH [−] populations of SUM159 cells were separated and cultured in suspension in the presence or absence of DOX for ten hours. [score:3]
Since TGFβ is a major inducer of the EMT [26], [27], we examined the effect of mir-93 expression on components of this pathway. [score:3]
Quiescent G0/G1 cells expressed lower levels of mir-93 than proliferating cells in S/G2/M phase. [score:3]
Furthermore, enforced expression of mir-93 increased the fraction of cycling cells. [score:3]
The levels of endogenous mir-93 expression paralleled cellular differentiation states with mir-93 levels lowest in the most primitive “claudin [low]” SUM159 cells, highest in the “luminal” MCF7 cells and intermediate in the “basal” HCC1915 cells. [score:3]
Figure S4 mir-93 expression in Sensor-GFP -positive and Sensor-GFP -negative SUM159 cells. [score:3]
Tetracycline (DOX) induces mir-93 expression in suspension-cultured SUM159 cells by 10 hours; B. 1×10 [6] SUM159 cells or pTRIPZ-SUM159 -mir-93 cells were plated in T75 flasks and, after overnight, the cells were treated with Vehicle control, with (DOX) or without (CTRL) DOX (1 ug/ml), docetaxel (10 nM) or the combination for 7 days. [score:3]
In addition to breast cancer cells, we also determined the effects of mir-93 expression on normal breast cell differentiation. [score:3]
As shown in Figure S3, mir-93 expression was lower in CSCs which were characterized by their expression of the CSC markers: ALDH [+] or CD24 [−]CD44 [+]. [score:3]
Figure S25 mir93 expression induces MET in HCC1954 cells. [score:3]
B. The effect of mir-93 expression on a panel of epithelial and mesenchymal markers at the mRNA level as accessed by qPCR. [score:3]
mir-93 expression decreased the CSC in these claudin [low] primary xenografts. [score:3]
qRT-PCR was utilized to access the effects of mir-93 on mRNA expression of mesenchymal markers (Vimentin, N-Cadherin and Twist), epithelial markers (E-Cadherin and Claudin), and TGFβR2. [score:3]
A. microRNA RT-PCR demonstrated SUM159-mirZip93-DsRed reduced mir93 expression by more than 90%. [score:3]
Furthermore, GFP was significantly reduced by overexpression of mir-93 (Figure S6), demonstrating that the sensor reports mir-93 function. [score:3]
Figure S15 Validation of the 127 overlapped gene expression with customerized StellArray PCR array plate in pTRIPZ-SUM159-mir93. [score:3]
Figure S16 Validation of the 127 overlapped gene expression with customerized StellArray PCR array plate in pTRIPZ-HCC1954-mir93. [score:3]
To determine whether the expression of mir-93 affect the growth of tumor metastasis in vivo, SUM159 (Figure 2G) and HCC1954 (1G) cells co -transfected with the inducible mir-93 vector and luciferase were introduced into NOD/SCID mice by intracardiac injection and metastasis formation monitored by bioluminescence imaging. [score:3]
Induction of mir-93 significantly inhibited the growth of SUM159 and HCC1954 xenografts (Figure 2C and Figure S1C). [score:3]
Induction of mir93 expression by DOX decreased the ALDH -positive population. [score:3]
mir-93 expression was significantly higher in GFP -negative cells than GFP -positive cells (Figure S4) and the ALDH1A1 was much lower in GFP -negative cells than GFP -positive cells as accessed by western blot or immunohistochemical staining (Figure S5). [score:3]
mir-93 expression in the adjuvant setting prevents tumor growth. [score:3]
B. Single cells were isolated from normal breast tissues and infected with mir-93 -expressing lentiviruses in suspension. [score:3]
A. ALDH [+] cells from SUM159 cells shows lower mir-93 expression level in comparison to ALDH [−] cells as accessed by qRT-PCR. [score:3]
mir-93 inhibits tumor growth and metastasis by decreasing CSCs in SUM159 cells. [score:3]
Cycling (S/G2/M) cells expressed significantly higher levels of mir-93 compared to quiescent (G0/G1) cells in both the ALDH [−] and ALDH [+] compartments (Figure 5A). [score:2]
Mir-93 is expressed equally in CD24 [−]CD44 [+] and bulk (non-CD24 [−]CD44 [+]) populations of MCF7 cells. [score:2]
mir-93 expression was significantly higher in DOX group compared to the control group at the end of treatment (Figure S2). [score:2]
Using a series of breast cancer cell lines representing different stages of differentiation and mouse xenograft mo dels, we demonstrate that mir-93 modulates the fate of breast cancer stem cells by regulating their proliferation and differentiation states. [score:2]
mir-93 regulates cell proliferation. [score:2]
ALDH [+] cells were significantly increased after mir-93 was knocked down (mirZip93-DsRed) (Figure S8B). [score:2]
Furthermore, in the claudin [low] SUM159 cells and basal HCC1954 cells, mir-93 expression is significantly lower in Aldefluor -positive as compared to Aldefluor -negative populations (Figure S11). [score:2]
As assessed by qRT-PCR, mir-93 expression was significantly increased in the ALDH [−] compared to ALDH [+] populations in SUM159 claudin [low] and HCC1954 basal subtype of human breast cancer (Figure 1A and Figure S1A). [score:2]
Furthermore, since the TET-inducible mir-93 system allows for the controlled regulation of CSC populations, it provides a valuable tool for assessing the role of CSCs in tumor growth in mouse xenograft mo dels. [score:2]
In summary, our experiments suggest that CSCs can exist in two alternative epithelial and mesenchymal states, the balance of which is regulated by miRNAs including mir-93 (Figure 8). [score:2]
As shown in Figure S8C, knockdown of mir-93 significantly promoted the growth of SUM159 cells in tumor xenografts and increased the CSC frequency. [score:2]
Furthermore, the proportion of ALDH [+] cells were significantly increased after mir-93 was knocked down (mirZip93-DsRed) (Figure S8D). [score:2]
Together these studies suggested that mir-93 regulated the CSC population and that this population mediates tumor growth following adjuvant therapy. [score:2]
In contrast, knockdown of mir-93 utilizing the mirZip93-DsRed promoted tumor invasion (Figure S8F). [score:2]
qRT-PCR was utilized to confirm the efficient knock-down of mir-93 (Figure S8A). [score:2]
mir-93 expression level was significantly higher in DOX group compared to the control group at the end of treatment (Figure S14). [score:2]
Enforced mir-93 expression in claudin [low] and basal breast cancer cell lines significantly reduced the CSC populations as assessed by the Aldefluor assay. [score:2]
The 127 genes in pTRIPZ-MCF7-mir-93 were also tested with customized PCR array plates, and interestingly, most of the stem cell genes were not knocked-down by mir-93 induction in the ALDH [+] proportion of MCF7 (Figure S21). [score:2]
In both SUM159 cells and HCC1954 cells, ALDH [+] cells have lower mir93 expression compared to ALDH [−] cells. [score:2]
We compared mir-93 expression levels in these four populations. [score:2]
In these studies, we demonstrate that mir-93 is capable of modulating breast CSC populations by regulating their proliferation and differentiation states. [score:2]
Consistent with this mo del, induction of mir-93 immediately after fatpad implantation or after development of micrometastasis by intracardiac injection completely blocked tumor recurrence. [score:2]
A hypothetic mo del illustrating regulation of normal and malignant mammary stem cell states and fates by mir-93. [score:2]
Figure S19 Luciferase assay testing mir93 targets. [score:2]
Figure S18 Luciferase assay confirming mir93 targets. [score:2]
We have utilized breast cancer cell lines representing different molecular subtypes of breast cancer as well as primary xenografts of breast cancer and normal mammary cells to examine the role of mir-93 in the regulation of normal and malignant breast stem cells. [score:2]
mir-93 regulates the cell cycle in SUM159 cells. [score:2]
In order to extend these observations to primary breast tumors, we examined the effect of mir-93 induction on three primary breast xenografts, MC1 (Figure S10A), UM2 (Figure S10B) and UM1 (Figure S10C) which were directly established from patient tumors and not passaged in vitro. [score:2]
However, development of metastasis were delayed and reduced in mice following mir-93 induction with or without docetaxel chemotherapy (Figure S1G). [score:2]
A. Mir-93 is expressed equally in CD24 [−]CD44 [+] and bulk (non-CD24 [−]CD44 [+]) populations of MCF7 cells. [score:2]
Figure S8Modulation of mir93 level in SUM159 cells altered cell invasion in vitro and knockdown increases tumor growth in vivo. [score:2]
Moreover, this did not appear to result from increased CSC apoptosis suggesting a potential role for mir-93 in promoting differentiation of CSCs. [score:1]
mir-93-sensor-GFP SUM159 cells were sorted for GFP -positive and GFP -negative cells by flow cytometry. [score:1]
To assess the functional relevance of this, we determined the effect of mir-93 induction in SUM159 and HCC1954 cells on tumor growth in NOD/SCID mouse xenografts. [score:1]
Cells isolated from primary xenografts MC1 (A) or UM2 (B) or UM1 (C) were transduced with the pTRIPZ-mir93 lentivirus in suspension. [score:1]
SUM159 cells were infected with no virus (non-infection), DsRed control lentivirus (SUM159-Neg-DsRed) or mirZip antisense mir93 lentivirus (SUM159-mirZip93-DsRed). [score:1]
Moreover, mir-93 -negative cells gave rise to tumors containing both mir-93 -negative and mir-93 -positive populations, whereas mir-93 -positive cells gave rise only to small, slow growing tumors containing exclusively mir-93 -positive populations (Figure 1D). [score:1]
This demonstrates that the effect of mir-93 on CSC populations is dependent on the cellular differentiation state. [score:1]
G. 200k pTRIPZ-HCC1954-mir93-luc cells in 100 ul of PBS were injected into the left ventricle of NOD/SCID mice. [score:1]
C. 100k pTRIPZ-HCC1954-mir93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
pTRIPZ-SUM159-mir93 cells are less invasive than the control cells in vitro accesses at 27 hours. [score:1]
10k pTRIPZ-MC1-mir93 or pTRIPZ-UM2-mir93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
To determine whether mir-93 induces or is a consequence of cellular proliferation, we utilized the DOX inducible mir-93 construct to determine the effect of mir-93 induction on cell cycle distribution. [score:1]
In contrast, cells isolated from tumors with mir-93 induction with or without docetaxel chemotherapy had markedly reduced tumor initiating capacity in secondary mice with no tumors observed from introduction of fifty cells from the mir-93 docetaxel treated group (Figure 2E, Figure S1E). [score:1]
Mir-93-sensor-GFP SUM159 cells were sorted for GFP -positive and GFP -negative cells by flow cytometry and RNA was isolated from both group of sorted cells and the expression level of mir-93 or RNU24 level was measured by qRT-PCR. [score:1]
Figure S71×10 [6] pTRIPZ-SUM149-mir93 cells were plated in T75 flasks and, after overnight, the cells were treated with Vehicle control or DOX (1 ug/ml) for 3–7 days. [score:1]
In contrast, the CSC population in MCF7 cells characterized by the phenotype CD24 [−]CD44 [+] [25] expressed the same high level of mir-93 as did the other (non-stem) cells constituting the bulk population (Figure 3A, Figure S11). [score:1]
mir-93 levels were significantly increased by ten hours following tetracycline induction in these cells (Figure 2A). [score:1]
In contrast, no metastases developed in mice following mir-93 induction with or without docetaxel chemotherapy (Figure 2G). [score:1]
Figure S23 Analysis of cell cycle for pTRIPZ-HCC1954-mir93 cells and pTRIPZ-MCF7-mir93cells. [score:1]
Figure S6 mir-93 induction reduces GFP in mir93-sensor-GFP-SUM159 cells. [score:1]
mir-93 -negative SUM159 cells have increased tumor-initiating capacity. [score:1]
Mir93 induced by DOX treatment resulted in a decreased proportion of cells in the G0/G1 phase and an increased proportion of cells in the S/G2/M phase for pTRIPZ-HCC1954-mir93 cells. [score:1]
mir93 level was analyzed by microRNA qRT-PCR. [score:1]
When tumors reached 0.2–0.3 cm in diameter, we induced mir-93 (with doxycycline treatment, hereafter DOX) or initiated cytotoxic chemotherapy with docetaxel or the combination. [score:1]
pTRIPZ-SUM159-mir-93 cells were plated in 2-well chamber slides with (DOX) or without (CTRL) Doxycycline for 7 days. [score:1]
mir-93 was shown as one of the most abundant miRNAs in ALDH [−] cells [21]. [score:1]
SUM159 cells were transduced with the pTRIPZ-mir-93 lentivirus and selected with Puromycin for 7 days. [score:1]
As shown in Figure 1C, sensor -positive (mir-93 -negative) cells had significantly higher tumor initiating capacity and CSC frequency than sensor -negative (mir-93 -positive) cells. [score:1]
Consistent with this, induction of mir-93 in MCF7 cells increased the CD24 [−]CD44 [+] population (Figure 3B). [score:1]
D. mir-93 -negative cells gave rise to tumors containing both mir-93 -negative and mir-93 -positive cell populations, but mir-93 -positive cells only gave rise to tumors containing mir-93 -positive cell populations. [score:1]
These studies demonstrated that mir-93 induction reduced the CSC population reducing growth of established tumor xenografts. [score:1]
Addition of the chemotherapeutic agent docetaxel resulted in a more significant reduction in tumor size, an effect that was accentuated by mir-93 induction. [score:1]
A. 500k pTRIPZ-T47D-mir93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
Cells from G0/G1 and S/G2/M were sorted from ALDH [+] or ALDH [−] populations and mir-93 expression was measured with qRT-PCR. [score:1]
We utilized a tetracycline (TET) inducible mir-93 construct tagged with RFP (pTRIPZ-mir-93-RFP) to determine the functional role of mir-93 in CSCs. [score:1]
The mo del depicted in Figure 8 is consistent with our observation that mir-93 level is highest in the EpCAM [+]CD49f [+] normal mammary cells and decreased with terminal differentiation. [score:1]
A. SUM159 cells were transduced with the pTRIPZ-mir-93 lentivirus and selected with Puromycin for 7 days. [score:1]
mir-93 initiates MET in SUM159 cells. [score:1]
These results were confirmed and extended by demonstrating that mir-93 induction in primary tumors increased their tumor-initiating capacity when implanted into secondary recipients (Figure 3E, Figures S12C, S13C). [score:1]
mir-93 promotes Mesenchymal-Epithelial Transition (MET) in SUM159 cells. [score:1]
Docetaxel also increased this population, as did the combination of mir-93 plus docetaxel (Figure 3B). [score:1]
C. 1000k pTRIPZ-MCF7-mir-93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
To simulate the adjuvant setting we induced mir-93 and/or administered docetaxel immediately after tumor cell implantation. [score:1]
Of interest, it has been recently reported that the mir-106b-25 cluster including mir-93 is induced in the early stages of nuclear reprogramming of fibroblasts into IPS cells [30]. [score:1]
Figure S3Induction of mir93 decreases both ALDH [+] and CD24 [−]CD44 [+] cells in the primary breast tumor xenografts. [score:1]
A serial dilution of mir-93 -negative (sensor/GFP -positive) SUM159 cells and mir-93 -positive (sensor/GFP -negative) SUM159 cells were injected into the 4 [th] fatpads of NOD/SCID mouse. [score:1]
200k pTRIPZ-SUM159-mir-93-Luc cells in 100 ul of PBS were injected into the left ventricle of NOD/SCID mice. [score:1]
In this setting, mir-93 induction had only a modest effect in reducing tumor growth. [score:1]
The 3′UTR of AKT3, SOX4, and STAT3 pMIR-REPORT firefly luciferase reporter plasmids with the wild-type 3′UTR sequences of AKT3, SOX4, or STAT3 were transiently transfected into pTRIPZ-MCF7-mir93 (A) or pTRIPZ-MDA-MB-453-mir93 (B) cells and an internal control ACTB luciferase reporter was co -transfected for normalization. [score:1]
mir-93 prevents tumor metastasis in the adjuvant setting. [score:1]
F. 10k pTRIPZ-SUM159-mir-93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
G. 200k pTRIPZ-SUM159-mir-93-Luc cells in 100 ul of PBS were injected into the left ventricle of NOD/SCID mice. [score:1]
The 3′UTR of AKT3, SOX4, and STAT3 pMIR-REPORT firefly luciferase reporter plasmids with the wild-type 3′UTR sequences of AKT3, SOX4, or STAT3 were transiently transfected into pTRIPZ-HCC1954-mir93 cells and an internal control ACTB luciferase reporter was co -transfected for normalization. [score:1]
mir93-sensor-GFP-SUM159 cells were infected with pTRIPZ-mir93 lentivirus and grow in T75 flasks. [score:1]
The pMIR-REPORT firefly luciferase reporter plasmids with the wild-type 3′UTR sequences of AKT3, SOX4, or STAT3 were transiently transfected into pTRIPZ-mir-93-SUM159 cells and an internal control ACTB luciferase reporter was co -transfected for normalization. [score:1]
pTRIPZ-HCC1954-mir93 cells were plated in 2-well chamber slides with (DOX) or without (CTRL) Doxycycline for 7 days. [score:1]
Recently, Ibarra, et al, showed that Let7, as well as mir-93 are highly depleted in mouse mammary stem/progenitor cells isolated with the stem cell marker ALDH [21]. [score:1]
F. 10k pTRIPZ-HCC1954-mir93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
Although tumors grew after four to five weeks in control animals, there was no observed tumor growth following mir-93 induction and/or docetaxel treatments for eight weeks (Figure 2F, Figure S1F). [score:1]
mir93 promotes MET in normal breast epithelial cells. [score:1]
A. pTRIPZ-SUM159-mir-93 cells were plated in 2-well chamber slides with (DOX) or without (CTRL) Doxycycline for 7 days. [score:1]
Furthermore, when treatment was discontinued at eight weeks, animals that received chemotherapy alone developed local tumor growth and metastasis while those with mir-93 induction with or without chemotherapy showed no recurrence when animals were sacrificed after four months. [score:1]
In contrast, DOX treatment has no effects on the cell cycle for pTRIPZ-MCF7-mir93 cells. [score:1]
100k pTRIPZ-SUM159-mir-93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
mir-93 maintains normal breast stem cells in an epithelial state. [score:1]
RNA was isolated from each group of sorted cells and the expression level of mir-93 or RNU24 level was measured by qRT-PCR. [score:1]
Analysis of treated MCF7 tumors confirmed that mir-93 induction increased the proportion of CD24 [−]CD44 [+] cells and ALDH [+] cells in tumors as did docetaxel or DOX plus docetaxel (Figure 3D). [score:1]
C. 100 k pTRIPZ-SUM159-mir-93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
Furthermore, induction of mir-93 increased the proliferation of SUM159 by 29% (Figure S22). [score:1]
Although mir-93 induction had similar effects on the basal HCC1954 cell lines it had no significant effect on the cell cycle of the luminal MCF7 cells (Figure S22, Figure S23). [score:1]
B. of pTRIPZ-SUM159-mir-93 cells in the presence or absence of DOX for 7 days. [score:1]
These studies suggested that mir-93 played a role in maintaining normal breast cells in an epithelial (EpCAM [+]) state. [score:1]
We demonstrate that induction of mir-93 in mesenchymal-like SUM159 cells induces an MET in the ALDH [+] CSC population characterized by increased expression of E-Cadherin and Claudin, with concomitant downreguation of mesenchymal genes, such as Vimentin, N-Cadherin and Twist. [score:1]
B. 1×10 [6] pTRIPZ-HCC1954-mir93 cells were plated in T75 flasks and, after overnight, the cells were treated with Vehicle control, DOX (1 ug/ml), docetaxel (10 nM) or the combination for 3–7days. [score:1]
In contrast, no tumors developed in mice following mir-93 induction with or without docetaxel (Figure 2F, Figure S1F). [score:1]
As shown in Figure S11, mir-93 levels correlate with postulated differentiation state of these cell lines. [score:1]
Together, these experiments suggested that the effects of mir-93 on the CSC population differed in different molecular subtypes of breast cancer, an observation consistent with the hypothesis that miRNA effects might be differentiation state dependent. [score:1]
mir-93 induction with DOX resulted in a decreased proportion of cells in G0/G1 and an increased proportion of cells in S/G2/M. [score:1]
The mesenchymal state associated with an invasive phenotype characterized by quiescence and low mir-93 expression is maintained by growth factors such as TGFβ. [score:1]
B. 1×10 [6] pTRIPZ-MCF7-mir-93 cells were plated in T75 flasks and, after overnight, the cells were treated with Vehicle control, DOX (1 ug/ml), docetaxel (10 nM) or the combination for 7 days. [score:1]
After sorting, total RNA were isolated from different sorted groups (EpCAM [+]CD49f [−], EpCAM [+]CD49f [+], EpCAM [−]CD49f [+], EpCAM [−]CD49f [−]) and mir-93 expression were measured by qRT-PCR. [score:1]
1000k pTRIPZ-MCF7-mir-93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
A. 200k pTRIPZ-MDA-MB-453-mir93 cells were injected into the 4 [th] fatpads of NOD/SCID mice. [score:1]
of pTRIPZ-HCC1954-mir93 cells and pTRIPZ-MCF7-mir93 cells in the presence or absence of DOX. [score:1]
Furthermore, induction of mir-93 further reduced tumor growth when added to the docetaxel chemotherapy (Figure 2C and Figure S1). [score:1]
Induction of mir-93 upon cell implantation completely prevented tumor growth in this mo del. [score:1]
mir-93 promotes tumor growth by increasing CSCs in MCF7 cells. [score:1]
At the end of treatment, cells from Control and DOX groups were isolated from the tumors and mir-93 expression level was measured by qRT-PCR. [score:1]
Figure S22The effects of mir93 on cell proliferation. [score:1]
mir-93 induction increased the proportion of ALDH [+] cells from 1.01% to 9.5% in MDA-MB-453 tumors (Figure S12B) and from 1.26% to 3.84% in T47D tumors (Figure S13B). [score:1]
This suggests that mir-93 may play a different role in more differentiated luminal breast cancer than in the more undifferentiated claudin [low] and basal subtype. [score:1]
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[+] score: 360
As indicated in Fig.  3A and B, inhibition of miR-93 significantly suppressed U87 cell invasion, while upregulation of miR-93 enhanced SF126 cell invasion. [score:8]
We observed that inhibition of miR-93 expression caused a significantly reduction in U87 cell proliferation, while overexpression of miR-93 markedly promoted SF126 cell proliferation (Fig.  2C,D). [score:7]
We further set three groups using U87 cells: miR-93 inhibitor, miR-93 inhibitor+siRNA NC, and miR-93 inhibitor+P21 siRNA. [score:7]
Fig. 3. Downregulation of miR-93 inhibits cell invasion in U87 and SF126 cells. [score:6]
In addition, we also examined several other cell cycle-related proteins, and found that knockdown of miR-93 increased the protein levels of P27 and P53, but decreased the Cyclin D1 protein levels in U87 cells, while overexpression of miR-93 reduced the protein levels of P27 and P53, but upregulated the Cyclin D1 protein levels in SF126 cells, when compared to the control groups, respectively (Fig.  4D). [score:6]
Based on these above data, our study demonstrates that miR-93, upregulated in glioma, promotes the proliferation, cell cycle progression, migration and invasion of human glioma cells and suppresses their chemosensitivity to TMZ. [score:6]
Therefore, we suggest that miR-93 may not only directly inhibit the expression of P21 at the post-transcriptional level, but also through mediation of P53. [score:6]
Ohta et al. reported miR-93 could enhance the proliferation, migration and invasion, while inhibited the apoptosis in hepatocellular carcinoma cells through directly targeting PTEN and CDKN1A and activating the c-Met/PI3K/Akt pathway (Ohta et al., 2015). [score:6]
These data suggest that knockdown of P21 reversed the suppressive effects of miR-93 inhibition on the cell cycle progression in U87 cells. [score:6]
Jiang et al. also reported that the expression of miR-93 was markedly upregulated in glioma tissues, and that the miR-93 levels were significantly correlated with clinicopathologic grade and overall survival in glioma, consistent with our findings (Jiang et al., 2015). [score:6]
Moreover, knockdown of P21 reversed the suppressive effects of miR-93 inhibition on the cell cycle progression in U87 cells. [score:6]
Fig. 5. Knockdown of p21 reverses miR-93 inhibitor mediated inhibition of cell proliferation. [score:6]
Fig. 2. Downregulation of miR-93 inhibits cell proliferation and arrests cell cycle in U87 and SF126 cells. [score:6]
Knockdown of P21 attenuated the suppressive effects of miR-93 inhibition on cell cycle progression and colony formation. [score:6]
As miRNAs play a role via regulating their targets expression (Areeb et al., 2015), we further investigated the putative targets of miR-93 by using several common miRNA analysis software, including Pictar, MicroRNA. [score:6]
In the present study, we used in situ hybridization and real-time RT-PCR to examined the expression of miR-93 in glioma, and found that it was significantly upregulated in glioma tissues compared to normal brain tissues. [score:5]
org, and Targetscan were used to analyze the putative target genes of miR-93. [score:5]
On the contrary, knockdown of miR-93 induced a cell cycle arrest at G0/G1 stage, accompanied with an upregulation of P21 in U87 cells. [score:5]
We also found that upregulation of miR-93 promoted the proliferation, migration and invasion of glioma cells, and that miR-93 was involved in the regulation of cell cycle progression by mediating the protein levels of P21, P27, P53 and Cyclin D1. [score:5]
Our data showed that knockdown of miR-93 enhanced the protein expression of P21 in U87 cells, while overexpression of miR-93 led to a significant decrease in the protein levels of P21 in SF127 cells, when compared to the control groups, respectively (Fig.  4C). [score:5]
To study the role of miR-93 in glioma cells, miR-93 inhibitor was used to decrease the miR-93 levels in U87 cells, while miR-93 mimic was used to increase its expression in SF126 cells. [score:5]
MiR -negative control (miR-NC, GenePharma, Shanghai, China), miR-93 mimic (GenePharma), miR-93 inhibitor (GenePharma), inhibitor-NC (GenePharma), p21 siRNA, p21-NC, or Lipofectamine 2000 was diluted with OPTI-MEM (Life Technologies), respectively. [score:5]
In conclusion, our study reveals an oncogenic role of miR-93 in the regulation of proliferation, cell cycle progression, colony formation, migration, invasion, and chemoresistance in glioma cells, possibly via directly targeting P21. [score:5]
org, and Targetscan, and P21 (encoded by CDKN1A) was predicated to be a target gene of miR-93 (Fig.  4A). [score:5]
Further investigation showed that upregulation of miR-93 enhanced the proliferation, cell cycle progression, migration and invasion of SF126 cells, while knockdown of miR-93 suppressed these malignant phenotypes of U87 cells, suggesting that miR-93 may promote the growth and metastasis of glioma. [score:5]
Recently, some specific miRNAs, e. g. miR-93, have been found to be involved in pathological processes by targeting some oncogenes or tumor suppressors in glioma. [score:5]
In situ hybridization data showed that miR-93 was positively expressed in 38 cases of glioma tissues, and the positive expression rate was 88.4% (38/43). [score:5]
To upregulate the miR-93 levels in SF126 cells, miR-93 mimic was used. [score:4]
Moreover, the upregulated miR-93 level was significantly associated with the advanced malignancy. [score:4]
P21, a direct target of miR-93, is involved in miR-93 -mediated glioma cell proliferation. [score:4]
Therefore, our study aimed to explore the expression and function of miR-93 in the regulation of the malignant phenotypes of glioma cells, as well as the underlying mechanism. [score:4]
Accordingly, we suggest that miR-93 can negatively mediate the protein expression of P21 by directly binding to its 3′UTR in glioma cell lines. [score:4]
In the present study, we observed that miR-93 was significantly upregulated in glioma, and increased miR-93 levels were significantly associated with the advanced malignancy. [score:4]
Knockdown of miR-93 in U87 cells significantly induced a cell cycle arrest at G0/G1 stage (Fig.  2E), while overexpression of miR-93 promoted the cell cycle progression in SF126 cells (Fig.  2F). [score:4]
In addition, we found that knockdown of miR-93 promoted the suppressive effects of on the protein levels of P21, P27 and P53, as well as the promoting effects on Cyclin D1. [score:4]
In the present study, we for the firstly time reported that knockdown of miR-93 enhanced the chemosensitivity of glioma cells to TMZ, by inhibiting cell proliferation and colony formation and inducing cell apoptosis. [score:4]
Moreover, we showed that its upregulation was significantly associated with the malignant progression as well as poor prognosis of glioma patients, suggesting that miR-93 may play an oncogenic role in glioma. [score:4]
P21 was further identified as a direct target of miR-93. [score:4]
To knockdown the miR-93 levels in U87 cells, they were transfected with inhibitor. [score:4]
In addition, in situ hybridization data also indicated that miR-93 was gradually upregulated as the malignant progression of glioma (Fig.  1A,B). [score:4]
In our study, we found that P21 was a direct target of miR-93, and its protein levels were negatively mediated by miR-93 in glioma cells. [score:4]
Based on these data, we suggest that the promoting effect of miR-93 on cell cycle progression as well as proliferation in glioma cells may be directly via inhibition of P21. [score:4]
In vitro study showed that miR-93 could directly target P21, and promote the malignant phenotypes of glioma cells, as well as their chemoresistance to TMZ. [score:4]
As indicated in Fig.  5B, cells in the G0/G1 stage were markedly decreased in the miR-93 inhibitor+P21 siRNA, while cells in the S stage were significantly increased in the miR-93 inhibitor+P21 siRNA, when compared to the other two groups, respectively. [score:4]
These data indicate that P21 is a direct target gene of miR-93. [score:4]
These data are consistent with the result of cell cycle analysis in U87 and SF126 cells after knockdown or overexpression of miR-93. [score:4]
We speculated that P21, as a direct target of miR-93, might be involved in the miR-93 -mediated chemoresistance of glioma cells to TMZ, which should be verified in the future studies. [score:4]
The expression levels of miR-93 were positively correlated to the glioma grade (P<0.01). [score:3]
Therefore, miR-93 may become a promising diagnostic marker and therapeutic target for glioma. [score:3]
MicroRNA miR-93 promotes tumor growth and angiogenesis by targeting integrin-beta8. [score:3]
In addition, they identified integrin-β8 as a target of miR-93, and found that higher levels of integrin-β8 were associated with cell death in tumor mass and in human glioblastoma (Fang et al., 2011). [score:3]
Moreover, the cell proliferation was further decreased in the TMZ+miR-93 inhibitor group. [score:3]
Similarly, knockdown of miR-93 decreased U87 cell migration, while overexpression of miR-93 promoted SF126 cell migration, compared to the control group, respectively. [score:3]
MicroRNA-93 activates c-Met/PI3K/Akt pathway activity in hepatocellular carcinoma by directly inhibiting PTEN and CDKN1A. [score:3]
MiR-93 was also found to promote the proliferation, invasion and metastasis of nasopharyngeal carcinoma cells in vitro and in vivo by directly targeting TGFbetaR2 (Lyu et al., 2014). [score:3]
In the present study, overexpression of miR-93 enhanced cell cycle progression, accompanied with decreased P21 levels in SF126 cells. [score:3]
In the present study, we found that miR-93 also had a suppressive effect on the protein levels of P53 in U87 and SF126 cells. [score:3]
U87 and SF126 cells were transfected with miR-93 inhibitor or mimic, respectively. [score:3]
These results suggest that P21, as a downstream effecter of miR-93, had suppressive effects on the colony formation capacities of U87 cells. [score:3]
To reveal the role of miR-93 in glioma, we firstly examined the expression levels of miR-93 in 43 cases of glioma tissues as well as eight cases of normal brain tissues by conducting in-site hybridization and real-time RT-PCR. [score:3]
Fig. 1. The expression of miR-93 in glioma. [score:3]
However, the positive expression rate of miR-93 in normal brain tissues was only 25% (2/8), significantly lower than that in glioma tissues (P<0.01). [score:3]
TGFbetaR2 is a major target of miR-93 in nasopharyngeal carcinoma aggressiveness. [score:3]
We further studied the effects of miR-93 overexpression or inhibition on the invasion and migration of glioma cells by conducting transwell assay and wound healing assay. [score:3]
In addition, Kaplan–Meier survival time analysis showed that higher miR-93 levels were significantly associated with shorter survival time of patients with astrocytoma, indicating that its expression level is associated with the worse prognosis (Fig.  1D). [score:3]
Recently, miR-93 was found to promote tumor growth and angiogenesis by targeting integrin-β8 and activation of PI3K/Akt signaling pathway (Fang et al., 2011; Jiang et al., 2015). [score:3]
They showed that miR-93-oveexpressing glioma cells induced the formation of blood vessels in the tumor tissue, which in return facilitated cell survival, resulting in enhanced tumor growth (Fang et al., 2011). [score:3]
These above data indicate that inhibition of miR-93 could enhance the chemosensitivity of U87 cells to TMZ. [score:3]
TMZ (100 μmol/l) was used to treat U87 cells with or without transfection with miR-93 inhibitor for 24 h. MTT was used to examine the cell proliferation. [score:3]
U87 cells were treated with miR-93 inhibitor or TMZ alone, or co -treated with both. [score:3]
Temozolomide was used to treat U87 cells with or without transfection with miR-93 inhibitor. [score:3]
In the present study, in situ hybridization and real-time RT-PCR data indicated that miR-93 was significantly upregulated in glioma patients (n=43) compared with normal brain tissues (n=8). [score:3]
Real-time RT-PCR also showed similar findings that miR-93 was significantly upregulated in glioma tissues compared to normal brain tissues (Fig.  1C). [score:3]
In addition, inhibition of miR-93 enhanced the chemosensitization of glioma cells to temozolomide (TMZ). [score:3]
We further examined the effects of miR-93 on the expression of P21 in U87 cells. [score:3]
Fig. 4. MiR-93 directly targets p21. [score:3]
However, the regulatory mechanism of miR-93 in the biological behaviors and chemoresistance of glioma cells remains unclear. [score:2]
The detailed role and underlying mechanism of miR-93 in the regulation of glioma growth and chemoresistance remain largely unclear. [score:2]
However, the detailed regulatory mechanism of miR-93 in glioma remains still largely unclear. [score:2]
MiR-93 is upregulated in glioma tissues compared to normal brain tissues. [score:2]
Knockdown of miR-93 enhances the chemosensitivity of U87 cells to TMZ. [score:2]
Our data showed that treatment with TMZ significantly induced cell apoptosis compared to the control group, and inhibition of miR-93 further enhanced the cell apoptosis in U87 cells treated with TMZ (Fig.  6B). [score:2]
As indicated in Fig.  5C, the colony formation rate was significantly higher in the miR-93 inhibitor+P21 siRNA group, when compared with that in the other two groups, respectively. [score:2]
We observed that knockdown of miR-93 promoted the effects of on the protein levels of these cell cycle-related genes (Fig.  6D), suggesting that these proteins are involved in the miR-93 -mediated chemosensitivity of U87 cells to TMZ. [score:2]
As demonstrated in Fig.  2B, transfection with miR-93 inhibitor led to a significant decrease in the mir-93 levels in U87 cells, when compared to the non -transfected U87 cells. [score:2]
Briefly, the miR-93 staining intensity was scored as 0 (negative), 1(+), 2(++), and 3 (+++). [score:1]
We further studied the effects of miR-93 on the drug resistance in glioma cells. [score:1]
miR-93 promotes cell proliferation in gliomas through activation of PI3K/Akt signaling pathway. [score:1]
We further studied whether P21 acted as a downstream effecter of miR-93 in glioma cells. [score:1]
Therefore, miR-93 may be a cause of chemoresistance of glioma to TMZ. [score:1]
These findings suggest that miR-93 plays an oncogenic role in the growth of glioma probably via promoting the cell cycle progression. [score:1]
As indicated in Fig.  2A, U87 cells showed the highest miR-93 levels, while SF126 cells showed the lowest miR-93 levels. [score:1]
High miR-93 level is associated with the advanced malignancy and poor prognosis of glioma patients. [score:1]
Besides, miR-93 was also suggested to be involved in glioma immune escape (Codo et al., 2014). [score:1]
Besides, several other cell cycle-related proteins including P27, P53 and Cyclin D1 were also mediated by miR-93 in glioma cells. [score:1]
HEK293 cells were co -transfected with WT P21-3′UTR or MUT P21-3′UTR plasmid (400 ng), and miR-NC or miR-93 mimic (50 nM), using Lipofectamine 2000. [score:1]
We further analyze the relationship between the miR-93 levels and the clinicopathological features of glioma. [score:1]
HEK293 cells were co -transfected with WT P21-3′UTR or MUT P21-3′UTR plasmid, and miR-NC or miR-93 mimic, respectively. [score:1]
To verify this prediction, we constructed the wild type (WT) P21 3′UTR containing the putative binding sequences (GCACUUU) of miR-93 and the mutant type (MUT) P21 3′UTR, within which the binding sequences were changed into ‘AAAAAAA’. [score:1]
Therefore, we investigated whether inhibition of miR-93 could enhance the chemosensitivity of U87 cells to TMZ. [score:1]
We further examined the miR-93 levels in several common glioma cell lines, and found that U87 cells showed the highest miR-93 levels, while SF126 cells showed the lowest miR-93 levels. [score:1]
Accordingly, we suggest that miR-93 may play a promoting role in glioma metastasis. [score:1]
Following 3 h incubation with pre-hybridization solution at 37°C, slides were incubated with miR-93 probe (2 μg/ml, Exiqon, Denmark) overnight at 55°C. [score:1]
MiR-93 has been demonstrated to play a key role in the development, progression and chemotherapy resistance in several common human cancers. [score:1]
Indeed, Fang et al. found that miR-93 could enhance the tumor growth of glioma cells in vivo (Fang et al., 2011). [score:1]
The expression of miR-93 in ISH was evaluated as others described (Luo et al., 2015) by two independent pathologists. [score:1]
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[+] score: 308
Other miRNAs from this paper: hsa-mir-30a, hsa-mir-181a-2, hsa-mir-181a-1, hsa-mir-125a
miR-93 did not directly inhibit the expression of AFP through direct binding on the 3′UTR as assessed by the DIANA microT v3.0 algorithm database, Target Scan and RNA22. [score:9]
We demonstrated that miR-93 promoted HCC cell proliferation, migration and invasion through activation of the oncogenic c-Met/PI3K/Akt pathway, and also inhibited apoptosis and drug-sensitivity by directly inhibiting PTEN and CDKN1A expression in HCC cells. [score:8]
Figure 7Schematic representation of miR-93 with the oncogenic c-Met/PI3K/Akt pathway miR-93 directly binds to the PTEN and CDKN1A 3′UTRs Growth inhibition curves for high (HepG2) and low (SNU449) miR-93 expressing HCC cell lines in the presence of tyrosine kinase inhibitors for 24-72 hrs. [score:8]
We also found that the inhibition of miR-93 improves the anti-HCC action of tyrosine kinase inhibitors by targeting the c-Met/PI3K/Akt pathway. [score:7]
Figure 6 Growth inhibition curves for high (HepG2) and low (SNU449) miR-93 expressing HCC cell lines in the presence of tyrosine kinase inhibitors for 24-72 hrs. [score:7]
Furthermore, we checked the expression of Akt and found that phospho-Akt levels were decreased by inhibiting miR-93 expression (Figure 4E). [score:7]
In our in vitro studies, we demonstrated that overexpression of miR-93 promoted cell proliferation, migration, invasion, and 3D culture sphere formation, while knockdown of miR-93 inhibited these respective functions. [score:6]
This study indicated that miR-93 is a novel regulator of the oncogenic c-Met/PI3K/Akt pathway in HCC, and inhibition of miR-93 hinders tumorigenesis and enhances sensitivity to conventional chemotherapy and tyrosine kinase inhibitor treatment. [score:6]
The expression of miR-93 was also suppressed by siMet in HepG2 and PLC/RPF/5 cells (Figure 5F-G). [score:5]
Anti-miR-93 suppressed miR-93 expression 6.3-fold in anti-miR-93 transfected HepG2 cells compared to control cells, and mimic-miR-93 induced miR-93 expression 3.3-fold in mimic-miR-93 transfected SNU449 cells compared to control-miR transfected cells (Supplemental Figure 3A-B). [score:5]
To test the sensitivity of HCC cells to specific drugs used in current treatments, we examined the response of high (HepG2) and low miR-93 (SNU449) expressing cells in the presence of two known HCC tyrosine kinase inhibitors, tivantinib and sorafenib. [score:5]
Therefore, transfection with anti-miR-93 increases cell sensitivity to sorafenib, since inhibition of miR-93 eliminates the inhibition of PTEN and CDKN1A, thereby activates these respective genes. [score:5]
This suggestes that PTEN and CDKN1A expression was suppressed by miR-93 binding to the 3′UTRs of those genes (Figure 4B-C). [score:5]
These results suggest that the expression of miR-93 is not only significantly higher in HCC than normal hepatocytes, but also correlates with worse disease-free survival in advanced HCC patients. [score:5]
miR-93 expression is correlated with c-Met expression. [score:5]
Phospho-Akt level was reduced by inhibiting miR-93 expression. [score:5]
HepG2 cells, which expressed high levels of miR-93, and SNU449 cells, which expressed low levels of miR-93, were selected for the studies. [score:5]
We demonstrated the mechanisms through which miR-93 inhibits PTEN and CDKN1A, thereby activating proliferation through the c-Met/PI3K/Akt pathway and inhibiting apoptosis in HCC. [score:5]
We examined the relationship between miR-93 expression and cell sensitivity to two chemotherapeutic drugs for advanced HCC (5-FU and cisplatin) and two tyrosine kinase inhibitors. [score:5]
Because HCC tumors with stronger c-Met staining showed higher miR-93 expression, we hypothesized that the efficacy of tivantinib could be associated with miR-93 expression. [score:5]
Previous studies have indicated that miR-93 may promote cell growth in ovarian cancer by directly targeting PTEN [27]. [score:4]
Figure 7Schematic representation of miR-93 with the oncogenic c-Met/PI3K/Akt pathway miR-93 directly binds to the PTEN and CDKN1A 3′UTRs miR q-PCR array screening of six HCC cell lines identified 29 miRs whose expression levels changed more than 2-fold up or down (Figure 1A). [score:4]
miR-93 expression correlates with HGF and c-Met IHC intensity. [score:3]
miR-93 inhibits drug-sensitivity. [score:3]
The expression of miR-93 was significantly higher in the specimens with vascular invasion (p=0.022), while miR-125a-5p was not significantly associated with the vascular-invasive status (p=0.073) (Figure 2B and Supplemental Figure 1C). [score:3]
miR-93 targets PTEN and CDKN1A in HCC. [score:3]
Our results indicate that miR-93 inhibits both PTEN and CDKN1A. [score:3]
Figure 5We performed c-Met IHC staining in PEAT and examined the relationship with miR-93 expression. [score:3]
The relative expression of miR-93 was linearly correlated with the c-Met IHC staining score (Fisher exact test; Supplemental Figure 8). [score:3]
These findings were corroborated by the stimulated expression of the PTEN and CDKN1A proteins after anti-miR-93 transfection (Figure 4D). [score:3]
miR-93 inhibits PTEN and CDKN1A. [score:3]
The combination of anti-miR-93 transfection and treatment with individual tyrosine kinase inhibitors for 24 hrs and 72 hrs significantly increased the sensitivity of HCC cells to both sorafenib (Figure 6A-a, b, c) and tivantinib (Figure 6A-d, e, f). [score:3]
The binding was specific because the Luc-reporter vector with mutated PTEN and CDKN1A 3′UTR was not affected by the changes of miR-93 expression (Figure 4B-C). [score:3]
Anti-miR-93 could improve the effectiveness of current chemotherapeutic agents and possibly decrease the dosage of those drugs, when accompanied with anti-miR-93, even for those with Childs-Pugh B or C liver disease. [score:3]
Anti-miR-93 suppressed proliferation in HepG2 (Figure 3A). [score:3]
Since α-fetoprotein (AFP) is an important HCC-specific diagnostic biomarker, we examined the association between miR-93 and AFP expression [24]. [score:3]
Using qRT-PCR analysis, AFP mRNA expression was significantly lower in anti-miR-93 transfected cells than in control-miR transfected cells (Supplemental Figure 6). [score:3]
2×10 [5] HCC cells were transfected with 36 nM of mirVana human miRNA inhibitor (anti-miR-93) or with negative-control (control-miR) oligonucleotides (Ambion). [score:3]
miR-93 expression is enhanced in HCC tumors. [score:3]
Figure 4Pathway analysis to confirm the target of miR-93. [score:3]
Because classical statistical tests can yield many false -positive and false -negative results when only three replicates for each case are used, fold-change was used to screen for differentially expressed genes between the anti-miR-93 and control transfected cells, and p-values were generated for the genes of interest. [score:3]
We performed c-Met IHC staining in PEAT and examined the relationship with miR-93 expression. [score:3]
Analysis identified PTEN and CDKN1A as candidate targets of miR-93 (Figure 4A). [score:3]
Two candidate miRs (miR-93, miR-125a-5p) exhibited greater than a 10-fold increase in expression. [score:3]
Treatment of cell lines with 50 ng/mL HGF for 24 hrs induced the expression of miR-93; HepG2 cells were the most influenced by HGF treatment (Figure 5D). [score:3]
The database showed that the potential targets of miR-93 are the PTEN and CDKN1A 3′UTR. [score:3]
We demonstrated that elevated miR-93 promoted HCC tumorigenesis and resistance to targeted therapy and chemotherapy (Figure 7). [score:3]
The combination of anti-miR-93 transfection and treatment with a tyrosine kinase inhibitor increased the sensitivity of HCC cells to tivantinib by 22.1-fold (IC50; control 1.44 ± 0.07 μM anti-miR 0.07± 0.02 μM) and sorafenib by 24.5-fold (IC50; control 8.31 ± 0.41 μM anti-miR 0.34 ± 0.02 μM) after 24 hrs, respectively. [score:3]
To determine the relative expression of miR-93 and related pathways, we performed on HCC cells transfected with anti-miR-93 versus control-miR transfected HCC cells. [score:3]
Transfection of anti-miR-93 inhibited cell growth activity, while mimic-miR-93 stimulates cell growth in HepG2 cell spheres (Figure 3J). [score:3]
miR-93 high group was significantly correlated with worse disease-free survival (p=0.035) but not overall survival (p=0.179) (Figure 2D and Supplemental Figure 2). [score:3]
There was no significant difference in their expression between anti-miR-93 and control-miR. [score:3]
In order to understand the role of miR-93 on the c-Met/PI3K/PTEN/Akt pathway, we then analyzed the protein expressions of PI3K class 1A and 1B (p85, phospho-p85, p110-alpha, -beta, and -gamma) by western blotting of HCC cells transfected with anti-miR-93. [score:3]
Overall, we also demonstrated that miR-93 inhibits the PTEN and CDKN1A genes, thereby controlling HCC cell apoptosis and c-Met/PI3K/Akt pathway activity. [score:3]
Furthermore, we demonstrated that high levels of miR-93 in tumors are significantly related to worse disease-free survival in HCC patients. [score:3]
The transfection of anti-miR-93 in SNU449 cells, a low miR-93 expressing cell line, had no effect on the sensitivity to both sorafenib (Figure 6B-a, b, c) or tivantinib after 24 hr and 72 hr treatment (Figure 6B-d, e, f). [score:3]
A significant correlation between c-Met IHC staining levels and miR-93 expression levels was observed (p=0.002, Wilcoxon test, Figure 5C). [score:3]
These results indicated that miR-93 functions as an oncogenic miR in HCC cells, however the efficacy of mimic or anti-miR-93 relies on the base miR-93 expression levels in the cells. [score:3]
Sorafenib targets MAPK (C-Raf and B-Raf) and VEGFR-2, but not the PI3K/Akt pathway, which allowed us to observe the effects of miR-93 on the c-Met pathway. [score:3]
The expression of AFP decreased significantly after the transfection of HCC cells with anti-miR-93, as shown by data. [score:3]
We discovered miR-93 is an oncogenic miR that stimulates tumorigenesis and is a potential therapeutic target in HCC. [score:3]
When we analyzed other cut-off points (20-80%), the miR-93 expression did not have a significant correlation with the cell number percentage of positive staining (Table 1). [score:3]
We then verified that miR-93 expression was significantly higher in HCC tumors (n=47) than normal liver tissues obtained from cancer-free patients (n=16), liver tissues from non-cancer patients with liver cirrhosis (n=8), and specimens of histopathologically tumor-free liver tissue removed during liver resection of melanoma hepatic metastases (n=10) (Figure 2C). [score:3]
DEF6, ASB16, CRLF1, ABCD1, HIC2, DHRS2 Drug-Sensitivity PBFKB4, MAD1L1, NHEJ1, RNASET2, MRPL3-18-30-40, TRIB3, TXNRD2, FOXA2, HNRNPC Pathway analysis to confirm the target of miR-93. [score:3]
miR-93 inhibited the PTEN and CDKN1A genes, thereby controlling HCC cell apoptosis and c-Met/PI3K/Akt pathway activity. [score:3]
Anti-miR-93 enhanced drug-sensitivity to tyrosine kinase inhibitors. [score:3]
Furthermore, the percentage of positive cells (>50% or <50%) had no significant correlation with miR-93 expression. [score:3]
To explore the relation between miR-93 and c-Met, we assessed the association of miR-93 and c-Met expression in HCC and non-HCC specimens. [score:3]
Figure 3Effect of miR-93 knockdown on HCC cell function in vitroClinical HCC specimens showed the relevance of miR-93, therefore we confirmed the significance in vitro. [score:2]
HCC tumor specimens (n=47) were separated into AFP mRNA positive (n=25) and negative groups (n=22); miR-93 expression was significantly higher in the AFP positive group compared to the negative group (p=0.048, student t-test, Figure 4F). [score:2]
Effect of miR-93 knockdown on HCC cell function in vitro. [score:2]
The expression of AFP was reduced by 1.6-fold in anti-miR-93 transfected cells compared to control-miR transfected cells. [score:2]
The expression levels of miR-93 and miR-125a-5p were stimulated 4.5-fold and 9-fold, respectively, in a cohort of 47 HCC tumors compared to 40 normal liver and liver cirrhosis tissues (Figure 2A and Supplemental Figure 1B). [score:2]
Schematic representation of miR-93 with the oncogenic c-Met/PI3K/Akt pathway miR-93 directly binds to the PTEN and CDKN1A 3′UTRs. [score:2]
These results suggested that miR-93 levels indirectly affect AFP activity. [score:2]
To demonstrate direct binding of miR-93 to these genes, we performed the co-transfection of mimic-, anti-miR-93, or control-miR with the vectors of luc-3′UTR PTEN or CDKN1A in HCC cells followed by luciferase activity analysis. [score:2]
We also examined the response of anti-miR-93 transfected HCC cells and control-miR transfected cells to chemotherapeutic agents used to treat advanced HCC patients, such as hepatic arterial infusion chemotherapy (HAIC) and transarterial chemoembolization (TACE) [26]. [score:1]
Therefore, we proceeded to focus on the mechanistic action of miR-93 in HCC. [score:1]
The mechanistic effects of miR-93 on oncogenic pathways were further examined by in-silico analysis using the DIANA microT v3.0 algorithm database. [score:1]
Clinical HCC specimens showed the relevance of miR-93, therefore we confirmed the significance in vitro. [score:1]
We propose that anti-miR-93 could be used in combination with sorafenib and tivantinib to increase their respective drug-sensitivity in HCC. [score:1]
Interestingly, tivantinib plus anti-miR-93 was significantly more effective than sorafenib plus anti-miR-93. [score:1]
Statistical analysis showed stimulation of miR-93 in the AFP positive group. [score:1]
In this study, we evaluated the miR expression profiles of HCC and non-HCC cell lines using miR q-PCR array analysis and identified miR-93 as a significant miR associated with HCC progression. [score:1]
The 47 HCC specimens were then categorized into a miR-93 high group (–ΔCq>1.4, n=25) and a miR-93 low group (–ΔCq<1.4, n=22) (Table 1). [score:1]
HCC clinical specimens and non-cancer liver specimens were assessed to confirm the significance of miR-93. [score:1]
Relative genes in miR-93 pathway. [score:1]
We also demonstrated that miR-93 plays a crucial role in modulating c-Met/PI3K/Akt pathway activity. [score:1]
miR-93 is enhanced in HCC tumor tissues assessed in PEAT. [score:1]
We have demonstrated that miR-93 is associated with drug-sensitivity to 5FU, CDDP, sorafenib, and tivantinib. [score:1]
Three HCC cell lines (HepG2, Hep3B, and PLC/RFP/5) were treated with either anti-miR-93 or control-miR and were divided into 96-well micro plates. [score:1]
In contrast, mimic-miR-93 increased proliferation of HepG2 and SNU449 cells (Figure 3A and Supplemental Figure 4A). [score:1]
Activation of miR-93 blocked PTEN, therefore Akt phosphorylation occurred. [score:1]
To show the specificity of binding, we also performed the co-transfection of mimic- or anti-miR-93 with the vectors of luc-mut/3′UTR PTEN or CDKN1A that contained mutated sequences at the miR-93 binding site (Supplemental Figure 5). [score:1]
Transfected HepG2 cells with control-, mimic-, or anti-miR-93 were grown for 10 days in regular medium condition on 3D cultured spheres (Figure 3I). [score:1]
We propose that miR-93 has an important function as a modulator of PTEN activity in HCC. [score:1]
The sensitivity was highest with the combination of tivantinib and anti-miR-93 in HepG2 cells (Figure 6A-f). [score:1]
The dead cells were visualized with propidium iodide (PI); anti-miR-93 transfected HepG2 cells were strongly PI stained relative to controls, demonstrating that apoptosis is enhanced with a loss of miR-93 function (Figure 3G). [score:1]
miR-93 increases proliferation, migration and invasion of HCC cells. [score:1]
showed significant increasing of miR-93 going from the c-Met negative group (0) to the c-Met staining group (1+, 2+ and 3+). [score:1]
Total RNA was isolated and mRNA was sequenced from cell lines transfected with anti-miR-93 or control miR. [score:1]
This was consistent with our hypothesis that miR-93 induces phosphorylation of Akt through PTEN degradation. [score:1]
Figure 2 HCC clinical specimens and non-cancer liver specimens were assessed to confirm the significance of miR-93. [score:1]
This study identified miR-93 as a novel oncogenic miR that is stimulated in both in vitro and in vivo studies in HCC. [score:1]
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An miRNA, miR-93, was significantly up-regulated, whereas phosphatase and tensin homologue (PTEN) expression was significantly down-regulated in all tested OS cells, when compared with hMSCs. [score:8]
Our cDNA array analysis demonstrated that PTEN was the only miR-93 target gene whose expression was uniformly up-regulated in all five OS cell lines. [score:8]
As our results indicated that miR-93 expression was consistently up-regulated in OS cell lines, we employed genome-wide mRNA profiling by cDNA array to identify possible targets of miR-93 in OS cells. [score:8]
miR-93 overexpression plays an important role in promoting lung cancer cell growth, angiogenesis, and metastasis, while its inhibition suppresses cell proliferation, migration, and colony formation [17, 18]. [score:7]
Effect of anti-miR-93 and PTEN expression vector on PTEN protein expression and its downstream factors in Saos cells after 48 hours of transfection Since the introduction of anti-miR-93 significantly inhibited cell proliferation of OS Saos cell lines, we hypothesized that anti-miR-93 might reduce the cell cycle progression and⁄or induce-apoptosis of the cells. [score:7]
It is possible that miR-93 may down-regulate PTEN expression via an indirect pathway. [score:7]
In addition, the down-regulation of miR-93 in these cells significantly suppressed tumor growth in vivo. [score:6]
The expression of phosphorylated Akt protein, which is known to be inversely correlated with that of PTEN, was significantly down-regulated in anti-miR-93 -transfected cells. [score:6]
e Up-regulation of PTEN using anti- miR-93 and PTEN -expression vector in ES cells. [score:6]
Thus, we can assume that up-regulation of miR-93 might affect OS cell cycle progression via control of PTEN expression. [score:6]
In the present study, our miRNA array results demonstrated that the expression of miR-93 was up-regulated in all five OS cell lines. [score:6]
It is noteworthy that up-regulation of PTEN by transfection with vectors encoding anti-miR-93 or PTEN itself induced apoptosis of OS cells, indicating that the repression of OS cell growth following restoration of PTEN expression came about by apoptosis, as well as by cell cycle retardation. [score:6]
The number of PTEN (d) and p-Akt (f) positive cells per unit area in the tumor lesionThe expression of PTEN (Fig.   6c) and p-AKT (Fig.   6e) within the primary tumor lesion was reduced in the anti-miR-93 and PTEN expression vector transfected tumor tissues. [score:5]
Taking these observations together, miR-93 can be seen to play a critical role in carcinogenesis through suppression of PTEN, and may serve as a therapeutic target for the treatment of OS. [score:5]
In addition, we have provided evidence that the expression of miR-93 in OS cells is significantly increased, and that this miRNA plays important roles in OS cell proliferation and tumorigenesis by targeting PTEN both in vitro and in vivo. [score:5]
The number of PTEN (d) and p-Akt (f) positive cells per unit area in the tumor lesion The expression of PTEN (Fig.   6c) and p-AKT (Fig.   6e) within the primary tumor lesion was reduced in the anti-miR-93 and PTEN expression vector transfected tumor tissues. [score:5]
Fig. 2miR-93 inhibits PTEN mRNA expression in Saos cells and silencing of PTEN with anti-miR-93 in OS cells. [score:5]
When anti-miR-93 was transfected into OS cell lines, PTEN expression was greatly increased, suggesting that PTEN might be a target of miR-93 in ES cells. [score:5]
An analysis using several algorithms, such as BLAST and TargetScan, further recommended PTEN as a putative target of miR-93. [score:5]
Saos transfected miR-93 inhibitor and PTEN vector showed the increase in expression of PTEN, phospho (p)-p21 and p-Bad. [score:5]
Although miR-93 may influence the expression of many genes, we focused on PTEN as its target in OS cells. [score:5]
Among the predicted target genes of miR-93 in the TargetScan (http://www. [score:5]
To test whether miR-93 expression affected endogenous PTEN expression, we transfected the miR-93 and miR-93 mutant oligonucleotides, as well as the negative control-miR, into Saos cells (Fig.   2b). [score:5]
Several studies have demonstrated that PTEN, a cell cycle and proliferation regulator, is the direct target of miR-93 in breast cancer [5, 20, 29]. [score:5]
The over -expression of miR-93 resulted in the suppression of PTEN protein levels, indicating that miR-93 might function as an oncogene in OS cells. [score:5]
miR-93 was down-regulated in all five OS cell lines. [score:4]
Given that PTEN acts as a major negative regulator of the PI3K/Akt signaling pathway, its inhibition by miR-93 might play an important role in the proliferation of other cell types [20]. [score:4]
Up regulation of miR-93 expression in OS cell lines. [score:4]
PTEN as a direct miR-93 target in OS cell lines. [score:4]
The miR-93 mutant did not result in any changes, whereas the miR-93 introduction group demonstrated a reduction in PTEN mRNA; therefore, miR-93 sequence can indeed be anticipated to directly inhibit PTEN mRNA function. [score:4]
The results suggested that miR-93 might affect the expression of PTEN genes by binding to 3′UTR of PTEN. [score:3]
The number of p-Akt positive cells per unit area was lower (p < 0.01) in the mice that ati-miR-93 (117.6 ± 31.1 cells/mm2) and PTEN expression vector (68.8 ± 25.1 cells/mm2) than in those that received untreated (234 ± 42.2 cells/mm2) and control-miR (206.2 ± 57 cells/mm2). [score:3]
Briefly, 1 ×  10 [6] Saos cells were seeded and incubated for 24 h, then anti-miR-93 or PTEN expression plasmid was added to the cells followed by incubation for 48 h. The cells were washed with PBS, suspended in annexin V binding buffer, then added to an annexin V-FITC solution and propidium iodide (PI) for 20 min at room temperature. [score:3]
The cell growth of Saos (Fig.   3a) and MG63 (Fig.   3b) was inhibited by the transfection of anti-miR-93 as determined by cell counting in comparison with untreated and negative control-miRNA transfected cells at 48 h after the transfection. [score:3]
Furthermore, miR-93 increased proliferation and decreased apoptosis in OS cells, whereas its silencing in these cells inhibited such carcinogenic processes. [score:3]
Several genes have been reported to be targets of miR-93, including RhoC, CCNG2, RBL2, NRF2, and DAB2 [17, 19, 26– 28]. [score:3]
Furthermore, transfection of anti-miR-93 inhibited the proliferation and cell cycle progression of ES cells. [score:3]
Four groups were; (1) untreated (n = 7); (2) transfected with of negative control-miRs (n = 7); (3) transfected with anti-miR-93 (n = 7); and (4) transfected with PTEN expression vector (n = 7). [score:3]
This observation is consistent with a previous report demonstrating that overexpression of miR-93 promotes glioma cell proliferation and cell cycle progression [30, 31]. [score:3]
a Predicted binding sites of miR-93 at the 3′UTR site of PTEN, as aligned by Target Scan and BLAST. [score:3]
The expression of miR-93 has been identified in various types of cancer, suggesting an oncogenic role [6, 30, 31]. [score:3]
The Saos and MG63 cells were plated in 6-well plates (1 × 10 [5] cells per well), and were transfected with or without anti-miR-93 inhibitor, negative control miRNA, PTEN plasmid and Mock plasmid vector using Lipofectamine 2000. [score:3]
The purpose of our study was to assess whether the expression of PTEN is repressed by miR-93 and to establish whether this pathway could play a role in tumorigenesis in OS cells. [score:3]
In order to examine whether miR-93 really inhibits PTEN mRNA function, we stopped new transcription with actinomycin D and then introduced miR-93 and a mutant with a changed seed region. [score:3]
Since the transfection of anti-miR-93 resulted in the reduction of PTEN expression, we next examine the effects of anti-miR-93 on the proliferation of ES cells. [score:3]
Based on this inverse correlation, we hypothesized that the effect of PTEN in OS cells may be directly or indirectly mediated, at least in part, via miR-93. [score:3]
), indicating that anti-miR-93 also inhibited the growth of OS cells in vivo. [score:3]
To determine the effect of anti-miR-93 and PTEN expression vector on the protein level of PTEN, we also performed immunofluorescence staining with PTEN (CST) or p-Akt antibodies (CST). [score:3]
The protein expression level of PTEN in the anti-miR-93 (20nM) transfected cells was increased to 3.45 folds of that in the control cells (p < 0.01) (Fig.   2f). [score:3]
Our results indicated that the expression of miR-93 was elevated while that of PTEN was repressed in all five OS cell lines, in comparison to hMSCs. [score:3]
Fig. 5Effects of anti-miR-93 and PTEN expression vectors on the cell cycle distribution in Saos. [score:3]
The transfection of hsa-miR-93 inhibitor (5, 10, 20, 40, 80nM) or negative control miRNAs (Control-miR) (5, 10, 20, 40, 80nM) (Invitrogen) was performed using Lipofectamine 2000 reagent (Invitrogen) in antibiotics-free OptiMEM (Invitrogen) according to the manufacturer’s instructions. [score:3]
Furthermore, the repression of miR-93 resulted in inhibition of OS tumor growth in vivo. [score:3]
The number of PTEN positive cells per unit area was higher (p < 0.01) in the mice that anti-miR-93 (208.3 ± 30.8 cells/mm2) and PTEN expression vector 233.8 ± 57.9 cells/mm2) than in those that received untreated (56.7 ± 16.7 cells/mm2) and control-miR (101.5 ± 28.1 cells/mm2) (Fig.   6d). [score:3]
Four groups were generated: (1) untreated control (n = 7); (2) transfected with negative control-miRNA (n = 7); (3) transfected with anti-miR-93 (n = 7); and (4) transfected with PTEN expression vector (n = 7). [score:3]
Ectopic expression of miR-93 decreased PTEN protein levels. [score:3]
The function of miR-93 has been reported to be important in the ecology of other types of cancer via targeting of PTEN [5, 20, 21]. [score:3]
A considerable complementarity between sequences within the seed regions of miR-93 and sequences in the 3′ untransrated region (UTR) of PTEN was predicted, using the algorithms in BLAST and TargetScan. [score:3]
Fig. 6Anti-miR-93 and PTEN vector suppressed in vivo tumor growth. [score:3]
PTEN plasmid vector transfected Saos (Fig.   3c) and MG63 (Fig.   3d) cells, as same as anti-miR-93 transfected cells, showed significant inhibition of the cell proliferation compared with the untreated and mock plasmid vector transfected cells. [score:2]
We observed an increased miR-93 expression by 4.41 ± 0.96 fold compared with miR-93 mutant (0.99 ± 0.71) or control-miR (1.00 ± 0.53) (p < 0.01) (Fig.   2c). [score:2]
analysis showed that the expression levels of p-Akt dramatically decreased in anti-miR-93 and PTEN vector transfected cells compared with untreated, negative control oligo and Mock vector transfected cells (Fig.   4). [score:2]
Furthermore, additional in vitro and in vivo experiments have demonstrated the critical role of miR-93 in regulating cancer cell growth and promotion of tumor progression [19, 20]. [score:2]
Therefore, miR-93 may have affected PTEN mRNA through direct interaction, at least in part. [score:2]
analysis showed that the expression levels of PTEN dramatically increased in anti-miR-93 transfected cells compared with untreated or negative control-miR transfected cells (Fig.   2e). [score:2]
In OS cell lines, the increased expression of miR-93 by 2.22 ~ 3.57 folds compared with hMSCs was observed. [score:2]
And decreased PTEN expression at the mRNA level following transfection with the miR-93-5p oligonucleotide 0.4 ± 0.22 compared with miR-93 mutant (0.75 ± 0.14) or control-miR (0.81 ± 0.13) (p < 0.01) (Fig.   2d). [score:2]
We next examined the function of miR-93 in regulating its potential target gene, PTEN, and in modifying the biological characteristics of OS cells. [score:2]
MiR-93 expression has been implicated in various cancer types, implying an oncogenic role [5– 7]. [score:2]
Therefore, we considered the possibility that miR-93 might affect anticarcinogenic mechanisms by targeting PTEN in OS cells. [score:2]
The transfection of miR-93-5p mimic (5′- CAAAGUGCUGUUCGUGCAGGUAG-3′) (20nM), miR-93-5p mutant (5′- CUUUCACGUGUUCGUGCAGGUAG-3′) (20nM) and negative control (NC) mRNAs (20nM) (Invitrogen) was performed using Lipofectamine 2000 reagent (Invitrogen) in antibiotics-free OptiMEM (Invitrogen) according to the manufacturer’s instructions. [score:1]
However, the role of miR-93 in the proliferation of OS cells remains unclear. [score:1]
In summary, the present study demonstrates for the first time an inverse correlation between miR-93 and PTEN in OS cells. [score:1]
Furthermore, sequence analysis suggested a possible association of miR-93 with the 3′-UTR of PTEN. [score:1]
Although future confirmation of the data presented in the current study using clinical OS samples is necessary, the novel information provided here regarding the link between miR-93 and PTEN in OS cells will be beneficial in better understanding ES oncogenesis, and may suggest certain therapeutic strategies for clinical application. [score:1]
The region complementary to the miR-93 seed region was found in the 3′-UTR of human PTEN (Fig.   2a). [score:1]
Furthermore, in flow cytometry analysis using Annexin V-FITC/PI double staining, there were no significantly differences in the distribution patterns between untreated, negative control miRNA, anti-miR-93 and PTEN plasmid transfected cells (Fig.   5h). [score:1]
f Densitometry quantification of PTEN protein after transfection of anti-miR-93. [score:1]
The experimental tumor bearing mo del was established by injection of 2 x 10 [6] cells transfected with anti-miR-93 suspended in 100 μl of normal saline into the gluteal region of nude mice (BALB/c nu/nu, Kudo, Tosu, Japan). [score:1]
The cleavage of PARP protein, a marker of caspase‑mediated apoptosis, was not observed in both anti-miR-93 and PTEN transfectants as well as untreated cells and negative control or mock plasmid vector transfectants. [score:1]
d After actinomycinD treatment, the mRNA expression level of PTEN after transfection of negative control-miR, miR-93 and miR-93 mutant was measured by qRT-PCR. [score:1]
The programmed cell death was induced by low dose ADM treatment in anti-miR-93 or PTEN plasmid vector in Saos cells (Fig.   5i). [score:1]
The introduction of anti-miR-93 into Saos cells resulted in the decreased growth of subcutaneous xenografted tumors in nude mice (Fig.   6a). [score:1]
Immunofluorescence of PTEN (c) and p-Akt (f) following transfection of anti-miR-93 and PTEN vector into tumor cells. [score:1]
The data suggested that the repression of miR-93 and restoration of PTEN might have resulted in G0⁄G1 retardation in OS cells. [score:1]
To monitor the cell cycle distributions, FACS analyses were carried out using anti-miR-93 and PTEN plasmid transfected cells (Fig.   5a-d). [score:1]
However, the biological role of miR-93 including the relationship to PTEN in OS cells has not yet been clarified. [score:1]
Induction of cell cycle arrest at G0/G1 phase by anti-miR-93. [score:1]
Our data show that miR-93 promoted the proliferation of OS cells via induction of cell cycle retardation at the G1/G0 phase. [score:1]
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Other miRNAs from this paper: hsa-mir-20a, hsa-mir-106b, hsa-mir-761
The results showed that the expression levels of N-cadherin and Vimentin were significantly upregulated in miR-93 high expression group, while E-cadherin expression was downregulated (Fig 6D). [score:13]
In this study, we found that critical molecular markers of EMT were affected by miR-93, and the expression of mesenchyme cell marker N-cadherin and Vimentin were upregulated, while the epithelia marker E-cadherin was downregulated in gastric cancer cells. [score:9]
Moreover, we found that the expression of miR-93 was downregulated (Fig 5C), while both mRNA and protein levels of TIMP2 was upregulated in the anti-miR-93 group compared to scramble group (Fig 5D and 5F). [score:8]
On the other hand, transfection of miR-93 inhibitor obviously suppressed expression of miR-93 in SGC-7901 cells (Fig 3A). [score:7]
Western blot assay showed that the expression of N-cadherin and Vimentin were significantly increased, and the expression of E-cadherin was decreased in the miR-93 mimics BGC-823 cells, whereas opposite results were shown in miR-93 inhibitor group (Fig 6A). [score:6]
0189490.g006 Fig 6 (A) The western blot assay showed that the EMT markers including N-cadherin and Vimentin expression were up-regulated after transfection of BGC-823 cells with miR-93 mimics. [score:5]
By comparing the expression profiles of miRNAs in gastric cancer, we found that miR-93 was obviously upregulated in gastric cancer tissues compared with that in matched normal mucosal tissues. [score:5]
We further performed luciferase report assay and found that overexpression of miR-93 significantly inhibited the luciferase activity of wild-type 3’-UTR of TIMP2, while knockdown of miR-93 significantly increased the luciferase activity of wild-type TIMP2 3’-UTR. [score:5]
The aberrant expression of miR-93 have been also reported and which may act as oncogene or tumor suppressor in numerous types of cancer, including glioma, hepatocellular cancer, endometrial carcinoma and breast cancer. [score:5]
In contrast, N-cadherin and vimentin protein were suppressed after transfection of SGC-7901 cells with miR-93 inhibitors. [score:5]
Furthermore, we also found the relative expression of TIMP2 was significantly decreased in 70 clinical gastric cancer tissues in which miR-93 was increased (Fig 7E), and the expression of miR-93 was negatively correlated with that of TIMP2 in gastric cancer tissues (Fig 7F, R [2] = 0.33, P< 0.01). [score:5]
In the present study, we found that miR-93 was significantly up-regulated in gastric cancer tissues compared with that in matched normal mucosal tissues, and analyzed the correlations between miR-93 expression and the clinicopathologic features. [score:5]
0189490.g003 Fig 3 (A) miR-93 inhibitors significantly decreased the expression of miR-93 in SGC-7901 cells. [score:5]
In conclusion, our study demonstrated that miR-93 was upregulated in gastric cancer and was associated with poorer clinical parameters in patients with gastric cancer. [score:4]
These results suggested that up-regulation of miR-93 may be involved in progression and metastasis of gastric cancer. [score:4]
We further clarified that miR-93 functions as a promoter in cell proliferation and metastasis in vitro and tumor formation in vivo by direct target of TIMP2. [score:4]
Inhibition of miR-93 could suppress cell viability (Fig 3B), undergo cell cycle arrest in G1-phase (Fig 3C) and induce apoptosis (Fig 3D) of SGC-7901 cells compared with their negative control. [score:4]
TIMP2 is a direct target of miR-93. [score:4]
These results demonstrated that up-regulation of miR-93 could accelerate cells proliferation. [score:4]
TIMP2 is a direct target of miR-93 in gastric cancer cells. [score:4]
Effect of miR-93 upregulation on cell viability, cell cycle and apoptosis rate of BGC-823 cells. [score:4]
In this study, we demonstrated that both mRNA and protein levels of TIMP2 were decreased after up-regulation of miR-93 in gastric cancer cells, but increased via anti-miR-93. [score:4]
Altogether, we validated that TIMP2 was a direct target of miR-93. [score:4]
Taken together, our results provided evidence that TIMP2 is a direct target of miR-93 and mediates miR-93 promotion of gastric cancer progression. [score:4]
Furthermore, silencing of TIMP2 enhanced migration and invasive ability of gastric cancer cells, which was consistent with the result obtained from the up-regulation of miR-93. [score:4]
In conclusion, our findings highlighted the key roles of miR-93 in gastric carcinogenesis and progression, and provided a promising therapeutic target in gastric cancer. [score:3]
By using bioinformatics and luciferase reporter assay, it was revealed that TIMP2 was a direct target of miR-93 and could reverse oncogenic functions of miR-93 in gastric cancer. [score:3]
0189490.g001 Fig 1 (A) qRT-PCR result for the expression of miR-93 in gastric cancer tissues and matched adjacent normal tissues. [score:3]
These results indicated that the expression of miR-93 was elevated in gastric cancer tissues and cells and may serves as a promoter in gastric cancer. [score:3]
Meanwhile, elevated expression of miR-93 was related to clinicpathological parameters of gastric cancer patients. [score:3]
In the present study, we observed high expression of miR-93 in both gastric cancer cells and tissues. [score:3]
In contrast, TIMP2 mRNA and protein levels were markedly increased in miR-93 inhibitor group. [score:3]
The results indicated that expression of miR-93 was significantly higher in gastric cancer tissues than in the adjacent normal tissues (Fig 1A). [score:3]
Effect of inhibition of miR-93 on cell proliferation, cell cycle distribution and apoptosis rate of SGC-7901 cells. [score:3]
The expression level of miR-93 in gastric cancer clinical specimens and cell lines. [score:3]
Moreover, the expression of miR-93 was negatively correlated with that of TIMP2 in gastric cancer tissues. [score:3]
Briefly, cells transfected with miR-93 mimics/inhibitor were seeded into 96-well plates at a cell density of 2×10 [3] cells per well. [score:3]
MiR-93 is up-regulated in gastric cancer cells and clinical samples. [score:3]
Because the expression of miR-93 was elevated in gastric cancer tissues, we next examined whether miR-93 expression level was associated with clinical characteristics. [score:3]
In contrast, miR-93 was also reported to suppress tumorigenesis in ovarian cancer cells [20]. [score:3]
Because miR-93 could alter the expression of EMT markers in gastric cancer cells, we further detected whether this phenotypes also occur in gastric cancer tissues. [score:3]
To validate the expression condition of miR-93 in human gastric cancer tissues, the miR-93 level in tumor tissues and paired adjacent normal tissues isolated from 70 patients was detected by using qRT-PCR. [score:3]
The correlation between miR-93 and TIMP2 expression levels was determined with Pearson’s correlation analysis. [score:3]
Several bioinformatics softwares including TargetScan, miRDB, PicTar and miRanda were used to validate complementarity between miR-93 and 3’-UTR region of TIMP2 mRNA. [score:3]
Cells were seeded in six-well plates for 24 h, and then transfected with miR-93 mimic or its inhibitor, TIMP2 siRNA, or their scrambled negative control by using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s protocol. [score:3]
of miR-93 mimics or inhibitor had no effect on the luciferase activity of mutant TIMP2 3’-UTR (Fig 7B). [score:3]
Moreover, the higher miR-93 expression was associated with advanced clinical stage (Fig 1C). [score:3]
Abnormal expression of miR-93 have been reported in several type of cancers, such as breast cancer [17], endometrial Carcinoma [18] and hepatocellular carcinoma [19]. [score:3]
However, the expression level of miR-93 was not obviously associated with gender, age at surgery, tumor size, and histological type. [score:3]
The vector was co -transfected with miR-93 mimics/inhibitor or negative control using Lipofectamine 2000. [score:3]
As shown in Table 2, the gastric cancer tissues and their paired non-tumor tissues from 70 cases were detected, and it was shown that higher expression of miR-93 was significantly associated with lymph node metastasis and advanced TNM stage (P<0.05). [score:3]
In addition, we confirmed that ectopic expression of miR-93 potentially promoted cell proliferation, migration and invasion in gastric cancer cells, while repressed cells undergo apoptosis and cell cycle arrest at G1 phase. [score:3]
48 h post-transfection of cells with miR-93 mimics, inhibitor or control (100nM), SGC-7901 and BGC-823 cells were collected and resuspended in phosphate buffered saline (PBS). [score:3]
0189490.g002 Fig 2 (A) miR-93 mimics significantly enhanced the expression of miR-93 in BGC-823 cells. [score:3]
Since transwell assays already indicated that migration and invasion were significantly increased after overexpression of miR-93, thus our results suggested that miR-93 promoted the EMT phenotypes, and then increased migration and invasion ability in vitro. [score:2]
We further established the miR-93 knock down nude mice xenografts mo del and found that miR-93 could promote tumor formation in nude mice. [score:2]
TIMP2 was identified as the top candidate target of miR-93, and was further confirmed by using luciferase activity assay. [score:2]
Knockdown of miR-93 in SGC-7901 cells increased TIMP2 mRNA and protein level. [score:2]
RT-PCR assay showed that transfection of miR-93 mimics significantly increased miR-93 expression in BGC-823 cell (Fig 2A). [score:2]
In addition, we examined whether TIMP2 was negatively regulated by miR-93 using IHC staining, and found that the protein level of TIMP2 in anti-miR-93 group was higher than that in scramble group (Fig 5G). [score:2]
Gain and loss function assays were performed by stable transfection of miR-93 mimics and miR-93 inhibitor. [score:2]
In glioma, miR-93 was found to facilitate cancer development by interacting with PI3K/AKT pathway [8]. [score:2]
MiR-93 inhibits the invasive potential of triple -negative breast cancer cells in vitro via protein kinase WNK1. [score:2]
MiR-93 expression is associated with clinicopathological characters of gastric cancer patients. [score:2]
showed that both the mRNA and protein levels of TIMP2 in miR-93 overexpression group were significantly decreased compared with negative control (Fig 7C and 7D). [score:2]
To ascertain whether TIMP2 could regulate the biological role of miR-93 in gastric cancer cells. [score:2]
MiR-93 Promotes Tumorigenesis and Metastasis of Non-Small Cell Lung Cancer Cells by Activating the PI3K/Akt Pathway via Inhibition of LKB1/PTEN/CDKN1A. [score:2]
To further confirm the interrelation between miR-93 and target gene, qRT-PCR and western blot assay were performed to evaluate the effect of miR-93 on TIMP2 expression in gastric cancer cells. [score:2]
It was found that the expression level of miR-93 was significantly increased in MKN-28, BGC-823, MKN-45, MGC-803, SGC-7901 compared to the normal gastric epithelial GES-1 cells (Fig 1D). [score:2]
In addition, it was shown that miR-93 was also involved in the occurrence or progression of gastric cancer [21]. [score:1]
Our in vitro and in vivo data together indicated that miR-93 serves as a promoter for tumor progression and acted as a predictor in gastric cancer patients. [score:1]
The opposite conclusion may be attributed to the fact that miR-93 play specific functions in differentiation degree and types of carcinoma. [score:1]
To understand the effect of miR-93 in EMT, we transfected miR-93 mimics or inhibitor into cells to investigate the variations of EMT markers. [score:1]
Additionally, because the expression of miR-93 was highest in SGC-7901 cell line, while lowest in BGC-823 cell line, these two cell lines were used to in our study for investigating the molecular mechanisms. [score:1]
MiR-93 regulates cell proliferation, apoptosis and cell cycle distribution of gastric cancer cells. [score:1]
We transduced miR-93 mimics, negative control, siRNA-TIMP2 and si-Scramble into SGC-7901 cells. [score:1]
TIMP2 mediates the biological functions of miR-93 in gastric cancer cells. [score:1]
The study in breast cancer showed that miR-93 was a new therapeutic and prognostic biomarker, which could restrain cell migratory capability and invasive potential of cancer cells [9]. [score:1]
For tumor growth research, 0.2 mL PBS containing 1×10 [6] cells of SGC-7901 transfected with miR-93 antagomir or scramble control (Ribobio, Guangzhou, China) were injected into right armpit of each nude mice after using ethanol and iodine to sterilize the insertion area. [score:1]
The miR-93 expression was measured by real-time PCR using TaqMan miRNAs Quantitation Kit (Applied Biosystems, Foster City, CA, USA). [score:1]
Multiple active results indicated that TIMP2 has candidate binding sites with miR-93 (Fig 7A). [score:1]
Hsa-miR-93 antagomir or the scramble control was transfected into SGC-7901 cells and then injected into the athymic nude mice. [score:1]
It was speculated that miR-93 enhanced cells viability through other pathways instead of TIMP2. [score:1]
Moreover, miR-93 was a metastatic promoter and may accelerate the process of epithelial-mesenchymal transition (EMT) in endometrial carcinoma cells [11]. [score:1]
The correlation between miR-93 and epithelial-mesenchymal markers in gastric cancer. [score:1]
Subsequently, we demonstrated that miR-93 played a role in promoting the cell proliferation, migration and invasion and induce EMT in gastric cancer cells. [score:1]
Correlation between clinicopathological characteristics and miR-93 expression. [score:1]
showed that compared with the control group the number of migrated and invaded BGC-823 cells was significantly increased after transfection of miR-93 mimics, while, knockdown of miR-93 significantly decreased the migration and invasion ability of SGC-7901 cells (Fig 4A and 4B). [score:1]
Effect of miR-93 on Xenograft tumor growth in vivo. [score:1]
To further validate the correlation, we monitored miR-93 levels in six gastric cancer cell lines as well. [score:1]
MTT assay showed that overexpression of miR-93 dramatically promoted cell proliferation of BGC-823 cells (Fig 2B) compared to the negative control group. [score:1]
Deciphering the molecular mechanism of miR-93’s role in human gastric cancer may advance our understanding of pathological mechanism underlying human gastric carcinogenesis, and provide a theoretical basis for further research in early diagnosis, clinical classification and biological therapy. [score:1]
To further characterize the potential action mechanism of miR-93 in gastric cancer, we employed bioinformatics software to search the target genes of miR-93 [26– 27]. [score:1]
Previous studies have demonstrated that miR-93 acts as an oncogene in cancers that can accelerate tumor growth, invasion and angiogenesis. [score:1]
Therefore, we concluded that miR-93 served as a promoter on migration and invasion in gastric cancer cells. [score:1]
Tumor volume of mice in scramble and anti-miR-93 groups. [score:1]
However, the biological functions of miR-93 and molecular mechanisms remain unclear in gastric cancer. [score:1]
was employed to assess the function of the miR-93 on the progress of gastric cancer. [score:1]
Therefore, we demonstrated that miR-93 enhances cell migration, invasion, and potentiate EMT in gastric cancer cells. [score:1]
0189490.g007 Fig 7(A) The predicted complementary sequence interaction between human miR-93 3'-UTR and TIMP2. [score:1]
In this study, we aimed to clarify the precise role of miR-93 in various biological processes and underlying mechanisms involved in gastric cancer. [score:1]
Tumor weight of mice in scramble and anti-miR-93 groups. [score:1]
TIMP2 mediates the effects of miR-93 in gastric cancer cells. [score:1]
These results demonstrated that miR-93 may acted as tumor oncogene and promote tumorigenicity in vivo. [score:1]
These results suggested that TIMP2 could mediates the biological functions of miR-93 in progression and metastasis of gastric cancer cells. [score:1]
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AGS cells were infected with a control lentivirus or a miR-93 overexpression lentivirus, or transfected with a PDCD4 overexpression plasmid, or co -transfected with the miR-93 overexpression lentivirus plus PDCD4 overexpression plasmid. [score:9]
In addition, after overexpressing or knocking down miR-93 in gastric cancer cells, we experimentally validated the direct inhibition of PDCD4 translation through miR-93. [score:9]
Subsequently, AGS cells (2 × 10 [6] cells per 0.1 mL) were infected with the miR-93 overexpression lentivirus, or transfected with the PDCD4 overexpression plasmid, or co -transfected with the miR-93 overexpression lentivirus plus PDCD4 overexpression plasmid. [score:9]
AGS cells were infected with a control lentivirus or a miR-93 overexpression lentivirus, or transfected with a PDCD4 overexpression plasmid, or co -transfected with a miR-93 overexpression lentivirus and a PDCD4 overexpression plasmid. [score:9]
Tumors with both miR-93 and PDCD4 overexpression exhibited significantly higher levels of PDCD4 compared to tumors with miR-93 overexpression (Fig. 6E,F), suggesting that PDCD4 overexpression could rescue the PDCD4 suppression caused by miR-93. [score:8]
As anticipated, overexpressing miR-93 significantly suppressed the PDCD4 protein levels in AGS cells, whereas miR-93 knockdown had the opposite effect on PDCD4 expression in these cells (Fig. 4B,C). [score:8]
Moreover, we showed that miR-93 inhibited PDCD4 expression and consequently inhibited apoptosis in cultured gastric cancer cells. [score:7]
To overexpress PDCD4, an expression plasmid designed to specifically express the full-length open reading frame (ORF) of PDCD4 without the miR-93-responsive 3′-UTR was constructed and transfected into AGS cells. [score:7]
The efficient overexpression of miR-93 and inhibition of PDCD4 protein in AGS cells by lentiviral transfection is shown in Supplementary Fig. 4. AGS cells were also transfected with a PDCD4 overexpression plasmid. [score:7]
In the present study, we observed that overexpressing miR-93 inhibits apoptosis in gastric cancer cells, and reducing PDCD4 expression mimics miR-93 induction. [score:7]
In summary, these results demonstrated that miR-93 directly binds to the 3′-UTR of the PDCD4 mRNA transcript and inhibits PDCD4 translation in gastric cancer cells. [score:6]
To determine the regulatory level at which miR-93 influenced PDCD4 expression, we repeated the above experiments and examined the expression of PDCD4 mRNA after transfection. [score:6]
Xenografts with both miR-93 and PDCD4 overexpression exhibited reduced cell mitosis compared to xenografts with miR-93 overexpression (Fig. 6G), suggesting that PDCD4 overexpression could attenuate the anti-apoptosis effect of miR-93. [score:6]
A 300-bp fragment containing the genomic miR-93 sequence was cloned into a lentiviral expression plasmid, and AGS cells were infected with the lentiviral plasmid to express miR-93. [score:5]
Among the 10 most upregulated miRNAs (Supplementary Table 2), miR-16-5p, miR-23b-3p, let-7a-5p, miR-15a-5p, miR-17-5p and miR-93 were identified as the candidate regulators of PDCD4. [score:5]
Immunohistochemical staining also revealed the presence of lower levels of PDCD4 in the tumors from mice implanted with miR-93 -overexpressing cells, whereas the tumors from the PDCD4 -overexpressing mice showed increased PDCD4 protein levels (Fig. 6G,H). [score:5]
Likewise, PDCD4 overexpression attenuated the anti-apoptosis effect caused by miR-93 overexpression (Fig. 6G,I). [score:5]
Direct post-transcriptional regulation of PDCD4 expression through miR-93. [score:5]
Interestingly, we observed that the restoration of PDCD4 expression successfully attenuates the anti-apoptotic effects of miR-93 on gastric cancer cells, although miR-93 has many other targets. [score:5]
This mutated luciferase reporter was unaffected through either the overexpression or knockdown of miR-93 (Fig. 4E). [score:4]
Knockdown of miR-93 was achieved after transfecting cells with anti-miR-93, a chemically modified antisense oligonucleotide designed to specifically target mature miR-93. [score:4]
After determining the levels of miR-93 in the same 6 pairs of gastric cancer tissues and noncancerous tissues, we observed that miR-93 levels were indeed upregulated in gastric cancer tissues (Fig. 3B). [score:4]
Furthermore, because miRNA sponge technology has been developed as an efficient tool for miRNA silencing, we employed this technology instead of transfection with miRNA antisense to knock down miR-93 and prove the role of miR-93 in suppression of PDCD4 protein in AGS cells (Supplementary Fig. 2A). [score:4]
Overexpression and knockdown of miR-93. [score:4]
Moreover, compared with cells transfected with pre-miR-93 alone, those transfected with both pre-miR-93 and the PDCD4-overexpression plasmid exhibited significantly higher apoptosis rates (Fig. 5), suggesting that miR-93-resistant PDCD4 is sufficient to rescue the suppression of PDCD4 through miR-93 and attenuate the anti-apoptosis effect of miR-93 on gastric cancer cells. [score:4]
Because animal miRNAs generally block translational processes without affecting transcript levels, these results implied the involvement of a miR-93 -mediated post-transcriptional regulatory mechanism in PDCD4 repression. [score:4]
How to cite this article: Liang, H. et al. miR-93 functions as an oncomiR for the downregulation of PDCD4 in gastric carcinoma. [score:4]
Tumors from the miR-93 -overexpressing group displayed reduced PDCD4 protein levels compared to tumors from the control group, whereas the tumors from the PDCD4 -overexpressing group showed elevated PDCD4 protein levels (Fig. 6E,F). [score:4]
These results indicate the importance of miR-93 targeting PDCD4 as a novel regulatory pathway in gastric cancer progression. [score:4]
A significant increase in the sizes and weights of the tumors was observed in the miR-93 -overexpressing group compared to the control group, whereas the sizes and weights of the tumors in the group implanted with the PDCD4-overexpression plasmid were dramatically decreased (Fig. 6A,B). [score:4]
The efficient overexpression and knockdown of miR-93 in AGS cells is shown in Fig. 4A. [score:4]
Validation of PDCD4 as a direct target of miR-93. [score:4]
To determine whether the negative regulatory effects of miR-93 on PDCD4 expression were mediated through the binding of miR-93 to the presumed sites in the 3′-UTR of the PDCD4 mRNA, the full-length 3′-UTR of PDCD4, containing the two presumed miR-93 binding sites, was placed downstream of the firefly luciferase gene in a reporter plasmid. [score:4]
The role of PDCD4 targeting through miR-93 in the regulation of apoptosis in gastric cancer cells. [score:4]
The correlation between miR-93 and PDCD4 was further examined after evaluating PDCD4 expression in AGS cells after the overexpression or knockdown of miR-93. [score:4]
After 60 days of xenograft growth in vivo, tumors from the miR-93-overexpression group showed a significant increase in the expression of miR-93 compared to tumors from the control group (Fig. 6C). [score:4]
The miR-93 sponge efficiently knocked down miR-93 expression in AGS cells (Supplementary Fig. 2B). [score:4]
Neither the overexpression nor the knockdown of miR-93 affected PDCD4 mRNA levels in AGS cells (Fig. 4D). [score:4]
We next analyzed the biological consequences of the miR-93 -driven repression of PDCD4 expression in gastric cancer cells. [score:3]
AGS cells were cultured in 12-well plates and transfected with pre-miR-93, anti-miR-93, PDCD4 siRNA, or the PDCD4 overexpression plasmid to induce apoptosis. [score:3]
The PDCD4 expression plasmid, miR-93 sponge plasmid and PDCD4 siRNA were transfected into AGS cells using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions. [score:3]
The cell proliferation rate measured by the percentage of Ki-67 -positive tumor cells was decreased in the group implanted with the PDCD4 overexpression plasmid and increased in the group implanted with the miR-93 overexpression lentivirus (Fig. 6G,I). [score:3]
Additionally, PDCD4 overexpression attenuated the promotive effect of miR-93 on tumor growth (Fig. 6A,B), suggesting that miR-93 might promote tumor growth by silencing PDCD4. [score:3]
The predicted interaction between miR-93 and the target sites in the PDCD4 3′-UTR are illustrated in Fig. 3A. [score:3]
To examine the direct binding of miR-93 to the target gene PDCD4, a luciferase reporter assay was performed as previously described 19. [score:3]
Identification of conserved miR-93 target sites within the 3′-UTR of PDCD4. [score:3]
Indeed, miR-93, derived from a paralog (miR-106b-25) of the oncogenic miR-17–92 cluster, is overexpressed in several types of cancers, including gastric cancer 10, lung cancer 11, breast cancer 17 and hepatocellular carcinoma 25. [score:3]
These results suggest that the targeting of PDCD4 is a major mechanism through which miR-93 exerts a tumor-promotive function. [score:3]
Taken together, these results indicate that miR-93 might inhibit cell apoptosis through silencing PDCD4. [score:3]
Therefore, we searched for miRNAs that target PDCD4 and identified miR-93 as a candidate. [score:3]
Taken as a whole, this study delineates a novel regulatory network employing miR-93 and PDCD4 to fine-tune apoptosis in gastric cancer cells. [score:2]
Furthermore, Hematoxylin and eosin (H & E) staining of xenograft tissues showed more cell mitosis in the group implanted with the miR-93 lentivirus compared with the control group, whereas confluent necrotic areas were observed in xenografts from the PDCD4 -overexpressing group (Fig. 6G). [score:2]
Moreover, knocking down miR-93 with the miR-93 sponge also increased apoptosis in AGS cells (Supplementary Fig. 2E–G). [score:2]
The role of miR-93 in regulating PDCD4 in gastric cancer cells. [score:2]
Furthermore, we introduced point mutations into the corresponding complementary sites in the 3′-UTR of PDCD4 to eliminate the predicted miR-93 binding sites (all two binding positions were mutated). [score:2]
As expected, AGS cells transfected with pre-miR-93 showed decreased apoptosis; in contrast, knocking down miR-93 with anti-miR-93 had the opposite effect on cell apoptosis (Fig. 5). [score:2]
These results were consistent with the findings of the in vitro assays, which firmly validated the role of miR-93 in promoting tumorigenesis through the targeting of PDCD4. [score:2]
For luciferase reporter assays, AGS cells were cultured in 24-well plates, and each well was transfected with 1 μg of firefly luciferase reporter plasmid, 1 μg of a β-galactosidase (β-gal) expression plasmid (Ambion), and equal amounts (100 pmol) of pre-miR-93, anti-miR-93 or the scrambled negative control RNA using Lipofectamine 2000 (Invitrogen). [score:2]
Cellular miR-93 levels were increased approximately 130-fold when AGS cells were transfected with pre-miR-93, and these levels decreased to approximately 30% of the normal level when AGS cells were treated with anti-miR-93. [score:1]
The resulting plasmid was transfected into AGS cells along with pre-miR-93, anti-miR-93 or scrambled negative control RNAs. [score:1]
Effects of miR-93 and PDCD4 on the growth of gastric cancer cell xenografts in mice. [score:1]
This finding suggested that the binding sites strongly contribute to the interaction between miR-93 and PDCD4 mRNA. [score:1]
Detection of an inverse correlation between miR-93 and PDCD4 levels in gastric cancer tissue samples. [score:1]
As expected, luciferase activity was markedly reduced in cells transfected with pre-miR-93 and increased in the cells transfected with anti-miR-93 (Fig. 4E). [score:1]
The influence of miR-93 and PDCD4 on the growth of gastric cancer cells in vivo. [score:1]
AGS cells were transfected with equal doses of pre-miR-control, pre-miR-93, anti-miR-control, anti-miR-93, control siRNA, PDCD4 siRNA, control plasmid, PDCD4 plasmid, pre-miR-control plus control plasmid, pre-miR-93 plus control plasmid, pre-miR-control plus PDCD4 plasmid, or pre-miR-93 plus PDCD4 plasmid. [score:1]
In addition, a miR-93 sponge was constructed by inserting six miR-93 complementary sites (in tandem) into the 3′ end of a non-coding RNA driven by the CMV promoter. [score:1]
These results suggest that miR-93 might be involved in the pathogenesis of gastric cancer as an oncomiR. [score:1]
Two predicted hybridizations were identified between miR-93 and the 3′-UTR of PDCD4. [score:1]
As a result, luciferase activity was markedly reduced in cells transfected with pre-miR-23b-3p, pre-miR-17-5p or pre-miR-93, while pre-miR-16-5p, pre-let-7a-5p and pre-miR-15a-5p have no influence on the luciferase activity (Supplementary Fig. 1B). [score:1]
These results suggested that miR-23b-3p, miR-17-5p and miR-93 could potentially bind to the 3′-UTR of PDCD4 mRNA transcript. [score:1]
The resulting plasmid was transfected into the human gastric carcinoma cell line AGS along with pre-miRNAs of miR-16-5p, miR-23b-3p, let-7a-5p, miR-15a-5p, miR-17-5p and miR-93. [score:1]
Because the luciferase activity was reduced to maximal 80% of the original activity by miR-93, we next focused on miR-93. [score:1]
The inverse correlation between miR-93 and PDCD4 protein levels (Fig. 3C) and the disparity between the miR-93 and PDCD4 mRNA levels (Fig. 3D) was further illustrated using Pearson’s correlation scatter plots. [score:1]
In each well, equal amounts of pre-miR-93, anti-miR-93 or scrambled negative control RNA were used. [score:1]
Synthetic pre-miR-93, anti-miR-93 and scrambled negative control RNAs (pre-miR-control and anti-miR-control) were purchased from Ambion (Austin, TX, USA). [score:1]
In the present study, we observed that the levels of miR-93 were higher in gastric cancer tissues than in noncancerous tissues. [score:1]
Furthermore, the miR-93 binding sequences in the PDCD4 3′-UTR are highly conserved across species. [score:1]
Detection of an inverse correlation between miR-93 and PDCD4 levels in gastric cancer tissues. [score:1]
To examine the binding specificity, the sequences that interacted with the miR-93 seed sequence were mutated (all two binding positions were mutated), and the mutant PDCD4 3′-UTR was inserted into an equivalent luciferase reporter. [score:1]
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Herein, we found that miR-93 was significantly upregulated in gliomas, and overexpressing miR-93 activated PI3K/Akt signaling through downregulating PTEN, PHLPP2 and FOXO3 expression via targeting their 3′UTRs, subsequently resulting in glioma cell proliferation and progression (Figure 7). [score:13]
Furthermore, we demonstrated that miR-93 activated PI3K/Akt signaling through downregulating PTEN, PHLPP2 and FOXO3 expression via targeting their 3′UTRs. [score:8]
Through analyzed by TCGA database, we found that the genomic copy number of miR-93 was substantially upregulated in 82.19% of GBM patients (n = 529), suggesting that the upregulated miR-93 in gliomas might be caused by genomic amplification. [score:7]
Moreover, the percentage of BrdUrd incorporating-cells and Cyclin D1 expression significantly decreased, but the expression of p27 [kip1] increased, in the miR-93-transduced cells treated with Akt inhibitor (Figure 6F and 6G). [score:7]
In the same context, such a multi-target effect of miR-93 on PI3K/Akt signaling also makes the miRNA molecule a potential target for delivery of effective and efficient inhibitory approaches against the pathway. [score:7]
These results suggested that silencing PTEN, PHLPP2 or FOXO3 expression in miR-93-repressed cells could reverse the inhibitory effect of the miR-93 inhibitor on glioma cell proliferation and tumorigenesis. [score:7]
MiR-93 overexpression has an important role in promoting lung cancer cell growth, angiogenesis and metastasis and inhibition of miR-93 significantly suppressed HepG2 cell proliferation, migration and colony formation [31– 34]. [score:7]
As shown Figure 5B, silencing PTEN, PHLPP2 or FOXO3 in miR-93 inhibitor transfected cells increased the expression level of Cyclin D1 and decreased the expression of p27 [Kip]. [score:7]
In this context, our finding that a simultaneous suppression of three potent inhibitors of the PI3K/Akt pathway by miR-93 strongly demonstrates a unique significance of miRNA in modulating essential signaling pathways as it targets multiple genes. [score:7]
miR-93 inhibitor (miR-93 inhibitor is a LNA/OMe modified antisense oligonucleotide designed specifically to bind to and inhibit endogenous miR-93 molecule) and negative control were purchased from RiboBio. [score:7]
High miR-93 expression was closely associated with shorter overall survival time, which suggests a possible link between high-level miR-93 expression and progression of human gliomas and highlights the potential value of the molecule as a predictive biomarker for disease outcome. [score:7]
showed that ectopic expression of miR-93 dramatically decreased, whereas inhibition of miR-93 increased, the protein expression of PTEN, PHLPP2 and FOXO3 in both LN18 and Hs683 glioma cells (Figure 4B). [score:7]
To assess whether miR-93 upregulation is correlated with glioma progression, we further examined miR-93 expression in 110 archived clinical glioma specimens. [score:6]
As shown in Figure 4A, PTEN and PHLPP2, which are the inhibitors of PI3K/Akt signaling pathway, and FOXO3, which critical regulator of cell-cycle, were found to be potential targets of miR-93. [score:6]
Whereas transfection with miR-93 inhibitor upregulated the luciferase activity, respectively (Figure 4C). [score:6]
As shown in Figure 6A and 6B, overexpressing miR-93 significantly increased, but silencing miR-93 decreased, the Akt activity and the expression of phosphorylated Akt (Ser 473) in glioma cells. [score:5]
Overexpressing miR-93 promoted, but inhibition of miR-93 reduced, glioma cell proliferation and cell-cycle progression. [score:5]
The expression of the miRNA was defined based on Ct, and relative expression levels were calculated as 2−[(Ct of miR-93) – (Ct of U6)] after normalization with reference to the expression of small nuclear RNA U6. [score:5]
In an attempt to identify the mRNA targets of miR-93, we performed a bioinformatics analysis using the publicly available algorithm (TargetScan 6.2). [score:5]
Besides, the PTEN, PHLPP2 or FOXO3 3′-UTR-luciferase reporter with a mutant miR-93 binding site seed sequence was not inhibited by ectopic expression of miR-93 (Supplementary Figure 2A and 2B). [score:5]
Furthermore, the level of DNA synthesis was significantly suppressed in miR-93 -inhibitor transfected LN18 and Hs683 cells, whereas the control cells displayed relatively higher BrdUrd incorporation rates (Figure 3C). [score:5]
Figure 1 (A) miR-93 is upregulated in paired glioma tissues and peripheral non-tumor tissues, each pair prepared from a same patient (P < 0.0001; NCBI/GEO/GSE44726). [score:4]
Interestingly, we found that there are several putative binding sites of NF-κB p50 and p65 on the promoter of MCM7, which suggesting that NF-κB might be involved in the upregulation of miR-93 in glioma. [score:4]
Therefore, our results suggested that miR-93 might play important roles in the development and progression of gliomas and represent as a potential therapeutic target for glioma therapy. [score:4]
These results suggest that downregulation of miR-93 could reduce the proliferation, tumorigenicity and cell cycle progression of glioma cells. [score:4]
miR-93 is upregulated in gliomas. [score:4]
PTEN, PHLPP2, and FOXO3 are direct targets of miR-93 in glioma cells. [score:4]
Taken together, the results confirm that PTEN, PHLPP2 and FOXO3 are direct targets of miR-93. [score:4]
In the present study, we found that miR-93 was significantly upregulated in glioma cell lines and glioma tissues. [score:4]
miR-93 was found to be markedly upregulated nasopharyngeal carcinoma and breast cancer and significantly correlated with poor prognosis [29, 30]. [score:4]
miR-93 directly suppresses PTEN, PHLPP2 and FOXO3 in glioma cells. [score:4]
Consistently, the transcriptional and translational levels of Cyclin D1 and p27 [kip1], two downstream effectors of PI3K/Akt signaling, were also significantly alliterated in the miR-93-deregulated glioma cells (Figure 6B and 6C). [score:4]
Taken together, our results suggest that miR-93 is upregulated in glioma and might represent a novel biomarker for the progression and prognosis of patients with glioma. [score:4]
The mo del of miR-93 -mediated PI3K/Akt signaling activation via down-regulation of PTEN, PHLPP2 and FOXO3 that results in the promotion of cell proliferation in gliomas. [score:4]
As shown in Figure 3A, suppression of miR-93 by miR-93 inhibitor significantly decreased the growth rate of LN18 and Hs683 cells compared with that of NC transfected cells. [score:4]
It has been demonstrated that PTEN and PHLPP2 are critical negative regulators of PI3K/Akt signaling, which prompted us to further examine whether dysregulation miR-93 alters the activity of PI3K/Akt signaling in glioma cells. [score:3]
Inhibition of miR-93 attenuates proliferation and cell cycle progression of glioma cells. [score:3]
Furthermore, univariate and multivariate Cox regression analyses revealed that the expression of miR-93 and glioma grade was identified as an independent prognostic factor (Supplementary Table 3). [score:3]
Loss-of-function studies using a miR-93 inhibitor were further performed to confirm the biological function of miR-93 in glioma progression. [score:3]
By analyzing a published micro-array -based high-throughput assessment (NCBI/GEO/GSE44726), we found that miR-93 was upregulated significantly in the glioma tissues compared with the peripheral noncancerous tissues (Figure 1A). [score:3]
Furthermore, when co -transfected with PTEN-, PHLPP2-, or FOXO3-3′UTR dual luciferase reporter plasmid, together with miR-93, miR-93 inhibitor or negative control, into the glioma cells, as shown in Figure 3C, miR-93 led to a consistent reduction of luciferase activity of PTEN, PHLPP2, FOXO3 reporters. [score:3]
PTEN, PHLPP2 or FOXO3 suppression is critical for miR-93 -induced cell proliferation and tumorigenesis in glioma. [score:3]
Figure 4 (A) Schematic putative target sites of miR-93 in 3′UTRs of PTEN, PHLPP2 and FOXO3, and the sequence of miR-93 mutant (performed as miR-93-mut). [score:3]
Suppression of PHLPP2, FOXO3, and PTEN by miR-93 is essential for glioma cell proliferation. [score:3]
Inhibition of miR-93 reduces cell proliferation and cell-cycle progression in glioma cells. [score:3]
Overexpression of miR-93 promotes proliferation and cell cycle progression of glioma cells. [score:3]
To evaluate the effects of PTEN, PHLPP2 and FOXO3 on miR-93 -induced glioma progression, we suppressed endogenous PTEN, PHLPP2 or FOXO3 expression with specific siRNAs (Figure 5A). [score:3]
Therefore, our results suggested that miR-93 contributes to progression of glioma and might represent as a potential therapeutic target for glioma therapy. [score:3]
Therefore, miR-93 acts as an onco-miR and might be served as a potential anti-cancer target. [score:3]
As an alternative mechanism to genetic alteration, overexpression of miR-93 might explain the observed over-activation of PI3K/Akt in glioma cells lack of PI3K amplification and genetic losses of PTEN, PHLPP2 and FOXO3. [score:3]
In addition, flow cytometry showed a significant increase in the percentage of cells in G1/G0 phase and a decrease in the percentage of cells in S phase in cells transfected with the miR-93 inhibitor compared with NC transfected cells (Figure 3D). [score:2]
The colony formation assay indicated that silencing PTEN, PHLPP2 or FOXO3 in miR-93 inhibitor transfected cells increased proliferation of glioma cells (Figure 5C). [score:2]
Consistently, an anchorage-independent growth assay revealed that miR-93 -overexpressing LN18 and Hs683 cells formed more and larger-sized colonies than control cells (Figure 2B). [score:2]
As shown in Figure 6D and 6E, inactivation of PI3K/Akt signaling using Akt inhibitor significantly decreased the growth rates of miR-93-transduced glioma cells, analyzed by colony formation and anchorage-independent growth assays. [score:2]
The result of colony formation assay revealed that ectopically expressing miR-93 in both LN18 and Hs683 cells markedly enhanced their growth ability, as indicated by the increase in colony numbers and sizes (Figure 2A). [score:2]
To investigate the biological function of miR-93 in the development and progression of glioma, glioma cells LN18 and Hs683 stably expressing miR-93 were established for the further study (Supplementary Figure 1). [score:2]
miR-93 promotes cell proliferation and cell-cycle progression in glioma cells. [score:1]
Meanwhile, the miR-93 sequence is located in the intron of the MCM7 (minichromosome maintenance 7) gene at chromosome 7q22.1. [score:1]
Therefore, it would be interesting to further examine the potential functions of miR-93 on other aspects of glioma progression. [score:1]
miR-93 activates PI3K/Akt signaling. [score:1]
Taken together, our results demonstrate that PI3K/Akt signaling plays essential function during miR-93 -induced glioma cells proliferation. [score:1]
pMSCV-miR-93 was then cotransfected with the pIK packaging plasmid into 293FT cells, using the standard calcium phosphate transfection method [35]. [score:1]
Activated PI3K/Akt signaling is essential for miR-93-promoted glioma proliferation. [score:1]
revealed that miR-93 levels strongly correlated with glioma WHO grade (P < 0.01) (Figure 1D, Supplementary Table 2). [score:1]
As shown in Figure 1D, miR-93 levels remained low in tumors of grades I but became markedly higher in those at grade III and was further elevated in grade IV gliomas. [score:1]
Kaplan-Meier analysis and log-rank test were employed and showed that the miR-93 levels significantly correlated with patient survival (P < 0.001; Figure 1E, Supplementary Table 2). [score:1]
Furthermore, we examined whether activation of PI3K/Akt signaling contributed to miR-93 -mediated gliomas cell proliferation. [score:1]
Of note, the luciferase activity was insusceptible by miR-93-mut (miR-93 mutant) transfection instead of miR-93 (Figure 4C). [score:1]
Consistently, real-time PCR analysis revealed that miR-93 was significantly overexpressed in 10 freshly collected gliomas samples as compared with paired adjacent non-tumor tissues obtained from the same patient (Figure 1B), and 9 glioma cell lines compared with the normal human astrocytes (NHA) control (Figure 1C). [score:1]
Currently, the molecular mechanisms in which miR-93 is overexpressed in gliomas are under investigation in our laboratory. [score:1]
Collectively, these results demonstrate that miR-93 functions to enhance proliferation, tumorigenicity and cell cycle progression of glioma cells. [score:1]
These findings suggest that miR-93 functions as an onco-miR in glioma progression at the level of enhancing cell proliferation. [score:1]
[1 to 20 of 72 sentences]
8
[+] score: 233
Other miRNAs from this paper: hsa-mir-106b
As a candidate target of miR-93-5p, ELPIN was down-regulated by miR-93-5p which bonded to its target site within the 3’UTR of ELPIN (Figure 4). [score:8]
The results revealed that miR-93-5p overexpression increased the number and length of branches (Figure 3A), while suppression of miR-93-5p inhibited the HUVECs lumen formation in terms of both number and length on the matrigel matrix (Figure 3B). [score:7]
The results of qRT-PCR revealed that transfection of miR-93-5p mimics reduced the mRNA level of EPLIN, while inhibition of miR-93-5p by transfection with ASO up-regulated EPLIN mRNA levels (Figure 4B). [score:6]
However, in these studies, miR-93-5p expression level was either up- or down-regulated in cancer cell lines, which were then co-cultured with endothelial cells (ECs) to examine the effect(s) of miR-93-5p on angiogenic capacities of ECs. [score:6]
As shown in Figure 1B, miR-93 expression level had positive correlation with the CD31 and Endomucin (EMCN) expression, which were considered to be strictly expressed in endothelial cells. [score:6]
The reporter assay further confirmed that miR-93-5p repressed EPLIN expression by directly combining with the 3’-UTR of EPLIN mRNA (Figure 4E), and this inhibitory effect was eliminated when the recognition site was disrupted by mutations (Figure 4E). [score:6]
We found that depletion of EPLIN by transfection of specific siRNA could attenuate the inhibitory effect of miR-93-5p ASO on the HUVECs migration (Figure 5C) and lumen formation capacities (Figure 5D), further confirming that miR-93-5p could promote angiogenesis by directly targeting EPLIN. [score:6]
In addition, the mechanistic studies have revealed that miR-93-5p promoted the angiogenesis process through inhibiting EPLIN expression in HUVECs. [score:5]
These results suggested that miR-93-5p might enhance angiogenesis and reduce vascular capillary stabilization by targeting ELPIN and that both miR-93-5p and EPLIN represented promising targets for developing novel anti-angiogenic therapeutics. [score:5]
However, miR-93-5p inhibitory oligos suppressed the angiogenic sprouting by decreasing both the number and the length of sprouts (Figure 3D). [score:5]
We also found that depletion of EPLIN by transfection of specific siRNA could attenuate the effects of miR-93-5p ASO on the ECs migration (Figure 5C) and lumen formation (Figure 5D), further confirming that miR-93-5p could regulate angiogenesis by directly targeting the EPLIN transcripts. [score:5]
EPLIN has been predicted as a target of miR-93-5p by multiple bioinformatics tools, including TargetScan, microRNA. [score:5]
In addition, the 3’-UTR of EPLIN mRNA has the targeting sequence of the seed region of miR-93-5p and EPLIN has been experimentally validated as a target of miR-93-5p by several cross-linking immunoprecipitation (CLIP) -based high throughput experiments. [score:5]
The results showed that miR-93-5p overexpression increased the number and the length of branches (Figure 3A), while blockade of miR-93-5p inhibited the HUVECs lumen formation in terms of both number and length on the matrigel matrix (Figure 3B). [score:5]
Fang et al. reported that miR-93-5p promoted tumor angiogenesis and metastasis by suppressing LATS2 expression in astrocytoma [13]. [score:5]
Furthermore, we analyzed The Cancer Genome Atlas (TCGA) database for breast invasive carcinoma (BRCA) to find the correlation of miR-93 expression with the expression of blood vessel markers. [score:5]
Overexpression of miR-93-5p accelerated human umbilical vein endothelial cells (HUVECs) proliferation and migration and promoted their lumen formation and sprouting in vitro, while blockade of miR-93-5p led to suppression in migration and angiogenic response of HUVECs. [score:5]
To reveal the mechanisms contributing to the increased cell viability induced by up-regulation of miR-93-5p, we examined the alterations in cell cycle. [score:4]
As shown in Figure 2B, up-regulated miR-93-5p levels significantly accelerated the growth of HUVECs. [score:4]
Further studies are guaranteed to fully understand the roles of miR-93-5p in regulating tumor angiogenesis, which include the expression levels of miR-93-59 in endothelial cells of tumor blood vessels and animal mo del -based in vivo studies. [score:4]
EPLIN was a direct target of miR-93-5p. [score:4]
We found that up- and down-regulated miR-93-5p levels significantly accelerated and decreased the growth of HUVECs, respectively (Figure 2B and 2D). [score:4]
Recent publications demonstrated that miR-93-5p is frequently up-regulated in multiple types of cancers, including breast cancer [10], osteosarcoma [11] and gastric cancer [12] and functions as an onco-miRNA in tumor growth [10– 12]. [score:4]
In this study, we have demonstrated that TNBC tissues with up-regulated miR-93-5p level also had a higher vessel density. [score:4]
In sum, our results have indicated that miR-93-5p promoted angiogenesis through down -regulating EPLIN and represented a promising target for developing novel anti-angiogenesis therapeutics. [score:4]
Fabbri et al. reported that miR-93 regulated the secretion of a panel of cytokines, chemokines and growth factors that were involved in angiogenesis in gliomas through suppressing IL-8 and VEGF [15]. [score:4]
We believe that this study contributed to the understanding of miR-93-5p’s biological functions, shed new light on the regulation of angiogenesis and provided a potential novel target for developing anti-angiogenic reagents. [score:4]
EPLIN was identified as a direct target of miR-93-5p. [score:4]
MiR-93-5p, a member of the miR-106b∼25 cluster, has been significantly up-regulated in multiple types of cancers, including breast cancer [10], osteosarcoma [11] and gastric cancer [12], and functions as an onco-miRNA in tumor growth [10– 12]. [score:4]
In this study, we demonstrate that the TNBC samples with high miR-93-5p expression had higher levels of blood vessels marker CD31 staining (Figure 1A), suggesting miR-93-5p might be involved in angiogenesis regulation. [score:4]
miR-93-5p expression level was positively correlated with blood vessel density in breast cancer. [score:3]
Aiming at the role(s) of miR-93-5p in TNBC angiogenesis, we conducted the CD31 staining of the TNBC tissues, and found that the samples with high miR-93-5p expression had more vessels (Figure 1A). [score:3]
qRT-PCR for miR-93 expression. [score:3]
miR-93-5p Promoted Cell Migration and Angiogenesis through Targeting EPLIN. [score:3]
A mutant reporter (pGL3-EPLIN-mt) with a mutation in the 3’-UTR complementary to the seed sequence of miR-93-5p was generated by using a Site Directed Mutagenesis Kit (Clontech, USA) according to manufacture’s instructions. [score:3]
In sum, our findings first demonstrated that miR-93-5p enhanced angiogenesis and reduced vascular capillary stabilization by targeting ELPIN in HUVECs. [score:3]
To deplete miR-93-5p, the anti-sense oligo (ASO) of miR-93-5p or control oligo were transfected into HUVECs and inhibition of miR-93-5p was confirmed by qRT-PCR (Figure 2C). [score:3]
miR-93-5p expression level was positively correlated with vessel density in breast cancer. [score:3]
In addition, EPLIN has been experimentally validated as a target of miR-93-5p by several CLIP -based high throughput experiments. [score:3]
Further studies revealed that over -expression of miR-93-5p enhanced cell cycle progression into the S/G2/M phases (Figure 2E) and promoted HUVECs proliferations (Figure 2F). [score:3]
The results indicated that overexpression of miR-93-5p enhanced the entrance of HUVECs into the S/G2/M phases of cell cycle, suggesting miR-93-5p could accelerate the cell cycle progression (Figure 2E). [score:3]
No literature has reported the role(s) of miR-93-5p in regulating the angiogenic capacity of endothelial cells directly. [score:3]
miR-93-5p promoted cell migration and angiogenesis through targeting EPLIN. [score:3]
revealed that inhibition of miR-93-5p led to decreased cell viability in HUVECs (Figure 2D). [score:3]
The expression level of miR-93 was normalized to U6 snRNA. [score:3]
HUVECs were transfected with miR-93-5p mimics or control oligos and the overexpression of miR-93-5p was validated by qRT-PCR (Figure 2A). [score:3]
The 3’ UTR of human EPLIN mRNA with mutated miR-93-5p targeting sequence was also shown. [score:3]
In another study, Fang et al. showed that miR-93 promoted astrocytoma growth and angiogenesis by down -regulating integrin-β8 [14]. [score:2]
We have reported that miR-93-5p played an oncogenic role in triple negative breast cancer (TNBC) development [17]. [score:2]
However, the role(s) of miR-93-5p in regulating angiogenesis remain largely underexplored. [score:2]
miRNA-93-5p enhanced ECs proliferation and migration in vitroWe then employed in vitro assays to examine the effect(s) of miR-93 overexpression on ECs. [score:2]
We then employed in vitro assays to examine the effect(s) of miR-93 overexpression on ECs. [score:2]
To establish the regulatory association between miR-93-5p and EPLIN, we initially measured the expression levels of EPLIN in response to changes in miR-93-5p levels. [score:2]
miR-93-5p has also been reported to participate in regulating angiogenesis in cancers. [score:2]
No reference regarding the direct effect(s) of miR-93-5p on angiogenesis of ECs has been published. [score:2]
However, these assays were performed based on the co-culture of tumor cells ectopically over -expressing miR-93-5p with endothelial cells. [score:2]
We therefore detected the effect of miR-93-5p on the migration capacity of ECs. [score:1]
Figure 4 (A) the 3’ UTR of human and mouse EPLIN mRNA were aligned with miR-93-5p. [score:1]
These results indicated that miR-93-5p might play a vital role in angiogenesis. [score:1]
miRNA-93-5p promoted ECs lumen formation and angiogenic sprouting in vitro. [score:1]
HUVECs were transfected with 100 nM of miR-93-5p mimic oligos (RiboBio, China) or miR-93-5p ASO (RiboBio, China) by using Lipofectamin 2000 reagent (Invitrogen, USA) according to manufacture’s instructions. [score:1]
Figure 2miR-93-5p enhanced ECs proliferation and migration in vitro (A) and (B) HUVECs were transfected with 100 nM miR-93-5p mimics or control oligos. [score:1]
These functions were in consistent with the oncogenic functions of miR-93-5p in cancer cells, including promoting tumor growth and cell cycle progression. [score:1]
miR-93-5p promoted ECs lumen formation and sprouting in vitro. [score:1]
In together, these results indicated that miR-93-5p could promote angiogenesis in vitro. [score:1]
Level 3 miRNA-Seq data for miR-93 and mRNA-seq data for CD31 and EMCN were retrieved from the TCGA Data Portal. [score:1]
In together, these results suggest that miR-93-5p could promote ECs proliferation and migration in vitro. [score:1]
miR-93-5p enhanced ECs proliferation and migration in vitro. [score:1]
Figure 3miR-93-5p promoted ECs lumen formation and sprouting in vitro (A and B) HUVECs were transfected with miR-93-5p mimics (A), miR-93-5p ASO (B), or corresponding control oligos; 48h after transfection the cells were digested and seeded on the matrigel. [score:1]
The correlations between the expression of miR-93 and CD31 or EMCN were calculated using the Pearson’s correlation analysis. [score:1]
miRNA-93-5p enhanced ECs proliferation and migration in vitro. [score:1]
The sequence of 3’-UTR of EPLIN mRNA is recognizable by the seed region of miR-93-5p (Figure 4A). [score:1]
Therefore, we investigated the direct role(s) of miR-93-5p in regulating the angiogenesis capacity of HUVECs. [score:1]
[1 to 20 of 73 sentences]
9
[+] score: 224
Other miRNAs from this paper: mmu-mir-93
This study showed for the first time that miR-93-3p inhibitor induced ccRCC cell apoptosis, inhibited metastasis, and upregulated PEDF expression. [score:10]
Shi, et al. found that TGF-β induces RBL2 expression and arrests renal cancer cell growth by downregulating miR-93-5p, and indirectly showed that miR-93-5p suppression induces G1 phase arrest. [score:9]
We searched the supplementary databases, TargetScan, miRanda, and PicTar, for potential miR-93-3p targets, and found that miR-93-3p was predicted to regulate PEDF. [score:6]
miR-93-3p expression was correlated with patient survival, and miR-93-3p upregulation was associated with poor prognosis (Figure 1B). [score:6]
In conclusion, we presented the first evidence that miR-93-3p inhibition suppresses ccRCC cell proliferation, metastasis, and invasion, and promotes apoptosis by regulating PEDF. [score:6]
Additionally, miR-93-3p inhibition suppressed ccRCC cell motility and invasion, and increased apoptosis. [score:5]
We also showed that miR-93-3p inhibits PEDF expression by binding the PEDF 3’-UTR, and miR-93-3p and PEDF levels are negatively correlated in ccRCC patients. [score:5]
Additionally, ccRCC tumor imaging in live mice demonstrated that miR-93-3p inhibition suppressed tumor growth and metastasis in vivo. [score:5]
miR-93-3p inhibition suppresses ccRCC tumorigenesis in vivo. [score:5]
Similar to the effects of anti-miR-93-3p, PEDF overexpression induced cell apoptosis (Figure 5A) and inhibited migration and invasion (Figure 5B–5C). [score:5]
PEDF overexpression reversed the effects of miR-93-3p inhibition in ccRCC cells. [score:5]
Wulfken, et al. demonstrated that miR-93-5p is upregulated in renal cancer patient serum and tissues, but the miR-93-5p function was not well understood [15, 16]. [score:4]
miR-93-3p directly targets PEDF. [score:4]
While miR-93 is upregulated in several solid tumors and is positively correlated with malignancy [19– 21], its oncogenic mechanisms in ccRCC have not been addressed. [score:4]
miR-93-3p upregulation was correlated with poor prognosis and shorter overall survival time in a large ccRCC patient cohort. [score:4]
miR-93-3p expression correlates with ccRCC patient outcome. [score:3]
Mutations were introduced by site-directed mutagenesis into putative binding sites in the 3′-UTR of PEDF gene for miR-93-3p using the TaKaRa MutanBEST Kit (Takara, Japan). [score:3]
This study presents the first evidence that miR-93-3p is overexpressed in ccRCC. [score:3]
miR-93-3p suppresses ccRCC cell apoptosis and promotes migration. [score:3]
Anti-miR-93-3p promotes cell apoptosis and suppresses invasion and metastasis in vitro. [score:3]
Anti-miR-93-3p promotes cell apoptosis and suppresses invasion and metastasis in vitroTo determine the effect of anti-miR-93-3p on apoptosis, we stained ACHN and 786-O cells with Annexin V and propidium iodide (PI). [score:3]
ccRCC cell transfection with an miR-93-3p expression vector did not affect cell viability or apoptosis, possibly due to the already -high viability and low apoptosis levels of these cells, but did decrease damage induced by H [2]O [2] treatment. [score:3]
miR-93-3p transfection increased treated cell viability 1.5-2-fold over controls (H [2]O [2] + NC), while anti-miR-93-3p again inhibited viability (Figure 2C). [score:3]
n=6, [**] P<0.001.786-O cells transduced to express luciferase were injected into mouse tail veins, followed by control or anti-miR-93-3p lentivirus. [score:3]
We also found that miR-93-3p overexpression was associated with poor prognosis in a large ccRCC patient cohort from a single institution. [score:3]
We found that miR-93-3p inhibition induced ccRCC cell apoptosis and decreased invasion and metastasis in vivo and in vitro. [score:3]
Average patient age (57.9 years old ≈ 60), 4cm tumor size (the Urological Diseases Guide indicates that tumor size at T1a stage is ≤4cm) and the average levels of miR-93-3p were the clinical variables in our ccRCC patients. [score:3]
“NC”, “miR” and “anti-miR” represented negative control, miR-93-3p and miR-93-3p inhibitor respectively. [score:3]
anti-miR-93-3p suppressed 786-O cell metastasis and proliferation (Figure 6D). [score:3]
miR-93-3p expression in 138 FFPE normal and ccRCC specimens (A). [score:3]
anti-miR-93-3p inhibited cell migration into the wounded area (Figure 3C). [score:3]
miR-93-3p mimic, negative control miRNA (NC), miR-93-3p antisense inhibitor (anti-miR-93-3p) and PEDF siRNA were synthesized by Shanghai GenePharma Company. [score:3]
Anti-miR-93-3p also affected expression of several apoptosis-related proteins and matrix metalloproteinases (MMPs). [score:3]
Anti-miR-93-3p suppressed ccRCC cell tumorigenesis and metastasis in vivo786-O cells were implanted subcutaneously in the flanks of nude mice, which were then injected with either control or anti-miR-93-3p lentivirus. [score:3]
Correlation of miR-93-3p expression with ccRCC patient overall survival (B). [score:3]
We detected miR-93-3p expression (normalized to matched normal kidney tissues) in 138 formalin-fixed, paraffin-embedded (FFPE) ccRCC specimens via qRT-PCR. [score:3]
Anti-miR-93-3p suppressed ccRCC cell tumorigenesis and metastasis in vivo. [score:3]
We confirmed that miR-93-3p overexpression is common in ccRCC. [score:3]
miR-93-3p inhibited relative luciferase activity in the reporter plasmid containing the wild type, but not mutants, PEDF 3’-UTR, demonstrating that (Figure 4E). [score:3]
To determine whether PEDF mediated miR-93-3p or anti-miR-93-3p activities, ACHN and 786-O cells were transfected with a control or PEDF expression plasmid. [score:3]
miR-93-3p was upregulated 38-fold on average in ccRCC specimens compared to normal kidney tissues (Figure 1A). [score:3]
786-O cells transduced to express luciferase were injected into mouse tail veins, followed by control or anti-miR-93-3p lentivirus. [score:3]
Our results suggest that miR-93-3p may predict ccRCC patient clinical outcome and serve as a novel anti-ccRCC therapeutic target. [score:3]
Thus far, only two published studies directly reported on miR-93 in ccRCC, and both focused on miR-93-5p. [score:2]
miR-93-3p expression was quantified using qRT-PCR (A) and cell viability was determined via MTT assay (B and C) in ACHN and 786-O cells 48 h after transfection with miR-93-3p mimics, anti-miR-93-3p, or NC oligo. [score:2]
Our results demonstrate that miR-93-3p is an oncogene in ccRCC, and may act by regulating PEDF. [score:2]
To determine whether there was a direct interaction between miR-93-3p and PEDF, the wild type or mutant PEDF 3’-UTR was inserted into the dual-luciferase reporter plasmid. [score:2]
Figure 2Effect of miR-93-3p on cell viability in vitromiR-93-3p expression was quantified using qRT-PCR (A) and cell viability was determined via MTT assay (B and C) in ACHN and 786-O cells 48 h after transfection with miR-93-3p mimics, anti-miR-93-3p, or NC oligo. [score:2]
miR-93-3p regulates ccRCC cell proliferation. [score:2]
Compared to the NC, miR-93-3p overexpression did not impact cell viability, whereas anti-miR-93-3p transfection decreased ACHN and 786-O cell proliferation (Figure 2B). [score:2]
These data indicated that miR-93-3p knockdown may serve as a potential anti-ccRCC therapeutic strategy. [score:2]
For experimental metastasis mo dels, control and stable knockdown miR-93-3p cells (stable luciferase -transfected 786-O cells) were injected into the tail vein of nude mice. [score:2]
The effects of miR-93-3p knockdown in ccRCC cells were abrogated by PEDF siRNA. [score:2]
anti-miR-93-3p injection inhibited tumor growth and reduced tumor weights and sizes compared to the control (Figure 6A–6C). [score:2]
Furthermore, cells were transfected with NC, miR-93-3p and anti-miR-93-3p respectively. [score:1]
While 69.5±4.0 ACHN cells and 122.8±19.3 786-O cells traversed the matrigel-coated membrane under control conditions, anti-miR-93-3p transfection decreased these numbers to 42.5±2.5 and 58.0±6.4 cells, respectively (Figure 3E). [score:1]
PEDF expression was measured using qRT-PCR (A) and western blotting (B) after NC, miR-93-3p, or anti-miR-93-3p transfection for 48 h in ACHN and 786-O cells. [score:1]
miR-93-3p relationships with clinical variables (sex, age, tumor size, location, and Fuhrman grade and pT status) were assessed using the Cox proportional hazard regression mo del. [score:1]
HEK293 cells were co -transfected on 24-well plates by Lipofectamine 2000 reagent (Invitrogen, USA) with 0.5 μg of constructed reporter plasmid and miR-93-3p or control miRNA at a final concentration of 50 nM. [score:1]
Additionally, co-transfection with PEDF siRNA reversed the effects of anti-miR-93-3p transfection in ACHN and 786-O cells (Figure 5D–5F). [score:1]
Cell apoptosis was analyzed via flow cytometry after PEDF siRNA was co -transfected with anti-miR-93-3p into ACHN and 786-O cells (D). [score:1]
Effect of miR-93-3p on cell viability in vitro. [score:1]
To determine the effect of anti-miR-93-3p on apoptosis, we stained ACHN and 786-O cells with Annexin V and propidium iodide (PI). [score:1]
To explore the role of miR-93-3p in tumorigenesis, we transfected ACHN or 786-O cells with negative control (NC), miR-93-3p, or anti-miR-93-3p. [score:1]
Neither miR-93-3p nor anti-miR-93-3p transfection affected PEDF mRNA levels in ACHN and 786-O cells (Figure 4A). [score:1]
The present study associated miR-93-3p with ccRCC patient survival via univariate and multivariate analyses, and assessed the biological functions of miR-93-3p in vitro. [score:1]
These results demonstrated that anti-miR-93-3p increased ACHN and 786-O cell apoptosis and decreased migration and invasion. [score:1]
786-O cells were implanted subcutaneously in the flanks of nude mice, which were then injected with either control or anti-miR-93-3p lentivirus. [score:1]
However, PEDF proteins were affected by miR-93-3p and anti-miR-93-3p transfections in both ACHN and 786-O cells (Figure 4B). [score:1]
miR-93-3p expression was calculated using the comparative Ct method. [score:1]
Clinical significance of miR-93-3p in ccRCC patients. [score:1]
Control refers to normal kidney tissues, and miR-93-3p levels were normalized to controls. [score:1]
Thus, the oncogenic activities of miR-93-3p in ccRCC are likely mediated by PEDF. [score:1]
These proteins were also detected after PEDF siRNA co-transfection with anti-miR-93-3p into ACHN and 786-O cells. [score:1]
ccRCC cells were transfected with miR-93-3p, NC, anti-miR-93-3p, siRNA or plasmid using Lipofectamine 2000 reagent (Invitrogen, USA) following the manufacturer’s instructions. [score:1]
Then, control lentivirus or anti-miR-93-3p lentivirus delivering approximately 2×10 [7] transforming units of recombinant lentivirus were injected into mice once through the tail vein. [score:1]
The potential interaction between miR-93-3p and putative binding sites in the PEDF 3’-UTR. [score:1]
anti-miR-93-3p transfection reduced these numbers to 54.3±11.2 and 89.5±41.3 in ACHN and 786-O cells, respectively (Figure 3D). [score:1]
Flow cytometry results showed that 13.1±2.3% and 10.6±1.4% of anti-miR-93-3p -transfected ACHN and 786-O cells, respectively, were apoptotic (Figure 3A). [score:1]
anti-miR-93-3p -transfected cells exhibited more Annexin V positivity (apoptosis) than NC -transfected cells. [score:1]
NC, miR-93-3p, or anti-miR-93-3p were co -transfected with the wild type PEDF 3’-UTR, Mut1, or Mut2 into HEK293 cells for 48 h. Luciferase activities were analyzed relative to the NC -transfected group (E). [score:1]
After transfection with 50 nM miR-93-3p, NC, anti-miR-93-3p, the viability of the cancer cells was detected with 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT; Roche; Switzerland). [score:1]
Univariate and multivariate analyses showed that miR-93-3p level was an independent predictor of ccRCC patient overall survival (Table 1). [score:1]
miR-93-3p, U6 levels and endogenous mRNA levels of PEDF and Actin were detected using SYBR Green PCR Master Mix kit in accordance with the manufacturer’s instructions (Applied Biosystems, USA). [score:1]
We then assessed the miR-93-3p protective function in ACHN and 786-O cells in an injury environment. [score:1]
The expression of miR-93-3p and PEDF mRNA were calculated using the comparative Ct method. [score:1]
Figure 4PEDF expression was measured using qRT-PCR (A) and western blotting (B) after NC, miR-93-3p, or anti-miR-93-3p transfection for 48 h in ACHN and 786-O cells. [score:1]
NC, miR-93-3p mimics, or anti-miR-93-3p were then co -transfected with the constructed plasmid into HEK293 cells. [score:1]
ccRCC cells transfected with NC, miR-93-3p, or anti-miR-93-3p were treated with H [2]O [2]. [score:1]
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10
[+] score: 199
Other miRNAs from this paper: hsa-mir-155, hsa-mir-517a
Importantly, overexpression of miR-93, miR-155, or an AID -targeting siRNA (shAID) in MCF-7 cells each reduced AID protein levels between 70 and 90%, whereas overexpression of the pAL-1 control did not significantly alter AID expression (Figure 3A). [score:9]
HSV-TKp, herpes simplex virus thymidine kinase promoter; SV40p, simian virus 40 promoter; FF Luc, firefly luciferase; Ren Luc, Renilla luciferase; pA, poly(A); 155TS, miR-155 target site; 93TS, miR-93 target site; 155SC, miR-155 target site scrambled; 93SC, miR-93 target site scrambled. [score:9]
Overexpression of miR-93 or miR-155 reduced AID protein levels in MCF-7 breast cancer cells (Figure 3A), whereas depletion of either endogenous miR-93 or miR-155 resulted in increased AID translation (Figure 4) directly connecting miR expression with changes in AID protein concentration. [score:8]
While a direct correlation between miR-93 activity and AID -induced oncogenesis remains to be described, miR-93 perturbations have been found to enhance cell survival, possess oncogenic activities, and augment tumor growth through regulating integrin-β8, the tumor suppressor gene FUS1, the Cdk inhibitor p21, and tumor protein 53 -induced nuclear protein 1 (TP53INP1) [39- 42]. [score:7]
Over -expression of miR-93 and miR-155 repressed the expression of endogenous AID, supporting the mo del that both miRs contribute to post-transcriptional regulation of AID. [score:6]
The human AID 3'UTR contains sequences complementary to miR-93 and miR-155In light of the evidence supporting AID regulation by miR-155 in mouse showing miR-155 regulates AID translation and c-myc translocation [29, 30], we asked if human AID is regulated by similar microRNA interactions. [score:6]
We therefore asked if miR-93 and miR-155 are present in two cell lines known to express AID, Ramos and MCF-7. Ramos is a B cell line that undergoes constitutive AID -induced somatic hypermutation [33], and MCF-7 is a breast carcinoma that misexpresses AID [24]. [score:6]
MiR-93 and miR-155 interact with the AID 3'UTR to inhibit its expressionThe identification of miR-93 and miR-155 binding sites in the 3'UTR of human AID strongly suggests post-transcriptional regulatory interactions. [score:6]
MCF-7 cells expressing either miR-93 or miR-155 sponges showed clear dose-responsive increases in AID protein levels, whereas control transfections did not significantly alter AID protein expression (Figure 4B). [score:5]
The AID 3'UTR luciferase reporter (AID 3'LR) and control 3'UTR luciferase reporter (Ctl 3'LR) were generated respectively by placing an AID 3'UTR sequence containing both the miR-155 and miR-93 target sites and a specific region of LacZ previously shown to be targeted by the pAL-1 shLacZ hairpin in a multiple cloning site in the Renilla luciferase 3'UTR [32]. [score:5]
In order to determine if endogenous AID can be repressed by increased miR-93 and/or miR-155 expression, we introduced pAL miR over -expression plasmids (Figure 2A) into MCF-7 cells [32]. [score:5]
If so, we anticipated that if miR-93 and/or miR-155 were present in AID expressing cells they could suppress endogenous AID protein levels. [score:5]
Our results suggest that in addition to these known oncogene regulations, loss of miR-93 (or miR-155) may permit AID upregulation and mutator phenotypes in non-B cell oncologies (e. g. breast, colon, stomach and lung [21- 24]). [score:5]
We conclude that one function of endogenous miR-93 and miR-155 expression is to restrict the translation of AID mRNA. [score:5]
In the present study, two human microRNAs, miR-155 and miR-93, were each shown to repress the translation of human AID through interactions with the AID 3'UTR in the MCF-7 breast carcinoma cell line which aberrantly expresses AID. [score:5]
This false polyA is located ~1,000 nt 3' of the AID stop codon and does not correspond to the full length cDNA polyadenylation as: (1) it is not preceded by the canonical 5' AAUAAA poly adenylation signal or other recognizable alternative poly adenylation signal, and (2) the miR-93 target site is expressed in both Ramos and MCF-7 cells (Figure 1B). [score:5]
Together, our findings demonstrate that miR-93 and miR-155 constitutively suppress AID translation in MCF-7 cells, suggesting widespread roles for these miRs in preventing genome cytidine deaminations, mutagenesis, and oncogenic transformation. [score:5]
In normal cells, top, genome stability is maintained by inhibition of AID translation by miR-93 and/or miR-155 when AID mRNA levels are low. [score:5]
MiR-93 and miR-155 interact with the AID 3'UTR to inhibit its expression. [score:5]
Together, our data suggest that low-level errant AID expression and subsequent genome damage may be prevented through protective miR-93 and/or miR-155 regulation. [score:4]
The full length AID 3'UTR contains a previously unidentified regulatory sequence, an unusually pronounced miR-93 complementarity, (Figure 1) conferring marked repression to targeted transcripts (Figure 2 and Additional File 2, Figure S1). [score:4]
Based on the involvement of AID in generating oncogenic genome mutations, our results suggest that miR-93 and miR-155 act as dual genome sentries to prevent errant translation of AID mRNA. [score:4]
In contrast, miR-93 and miR-155 can greatly reduce the potential for errant AID expression which protects the genome from unwanted cytidine deamination and mutation by restricting AID protein production (Figure 5). [score:4]
In addition, our identification of a miR-93 target site for AID located downstream of an internal adenosine repeat highlights the possibility that other miR regulations may have been overlooked because of truncated 3'UTR annotations. [score:4]
Mo del for AID repressionWe have shown that miR-93 and miR-155 have the capacity to repress AID translation and do so in the cellular context. [score:3]
Further analysis of the human AID 3'UTR revealed an internal 20 nt stretch of adenosines (sufficient for cDNA poly-T priming) separating the miR-155 and miR-93 target sites (Figure 1A). [score:3]
Figure 4 Endogenous miR-93 and miR-155 restrain AID protein expression. [score:3]
To be consistent with that hypothesis, the loss of cellular miR-93 or miR-155 should result in higher AID expression. [score:3]
Therefore, miR-93 and miR-155 facilitate molecular interactions with the AID 3'UTR sufficient to repress translation. [score:3]
Endogenous miR-93 and miR-155 restrain AID protein translation. [score:3]
Conspicuously, miR-93 is intronically encoded within the minichromosome maintenance 7 (MCM7) gene, an essential replication licensing factor, and precursor miR-93 abundance tracks with MCM7 expression levels [37, 38]. [score:3]
To achieve this, we constructed miR sponge transgenes for miR-93 and miR-155 that function by competitively inhibiting miR activity, as previously described [35, 36]. [score:3]
MiR-93 and miR-155 can repress endogenous AIDInteractions between miR-93, miR-155 and the AID 3'UTR within the context of our reporter assays (Figure 2C-D) strongly support the involvement of both miRs in AID translational regulation. [score:3]
Three expression constructs incorporating an endogenous human microRNA Alu promoter were created by cloning the endogenous miR-517a genomic locus and then replacing the miR-517a hairpin with the miR-93, miR-155 and shLacZ hairpins (pAL-93, pAL-155, and pAL-1 respectively)(Figure 2A), as described [32]. [score:3]
We have discovered moderate miR-155 and pronounced miR-93 complementary target sites encoded within the human AID mRNA. [score:3]
MicroRNAs miR-93 and miR-155 are wi dely expressed. [score:3]
Interactions between miR-93, miR-155 and the AID 3'UTR within the context of our reporter assays (Figure 2C-D) strongly support the involvement of both miRs in AID translational regulation. [score:3]
We have shown that miR-93 and miR-155 have the capacity to repress AID translation and do so in the cellular context. [score:3]
Strikingly, we find the predicted binding affinity between miR-93 and the AID 3'UTR (-30.3 kcal/mole) to be nearly double that of miR-155 (-15.3 kcal/mole) (Figure 1A) suggesting a strong post-transcriptional regulatory relationship. [score:2]
In addition, our characterization of an obscured miR-93 target site located within the AID 3'UTR supports the recent suggestion that many miR regulations have been overlooked due to the prevalence of truncated 3'UTR annotations. [score:2]
Figure 5 Mo del of miR-93 and miR-155 regulation of AID mediated genomic instability. [score:2]
To ensure results were the consequence of miR regulation, we confirmed the miR-93 repression of AID was dose -dependent (Additional File 4, Figure S3) and isolated total RNA from each of our transfections to monitor AID mRNA levels by quantitative PCR. [score:2]
The identification of miR-93 and miR-155 binding sites in the 3'UTR of human AID strongly suggests post-transcriptional regulatory interactions. [score:2]
Luciferase reporter assays indicate that both miR-93 and miR-155 can interact with the 3'UTR of AID to block expression. [score:2]
It is therefore tempting to speculate that domestication of the miR-93 microRNA along with the corresponding AID 3'UTR site occurred as a response to the selective pressure for maintaining genome fi delity during replication. [score:1]
In these situations, endogenous miR-155 and miR-93 have negligible influence on AID protein levels. [score:1]
Our miR sponge transgenes produce RNAs containing ~10 copies of the miR-93 or miR-155 AID 3'UTR complementary binding sites (Figure 4A). [score:1]
The human AID 3'UTR contains sequences complementary to miR-93 and miR-155. [score:1]
m, miR-93; β, Beta actin positive control; - Ctl, no template. [score:1]
MiR-93 and miR-155 can repress endogenous AID. [score:1]
Therefore, we next asked if miR-93 and miR-155 were capable of regulating the AID 3'UTR using standard luciferase reporter assays. [score:1]
The miR-93 sponge produces a polyadenylated concatamer of 11 individual 93 TSs. [score:1]
To our surprise, we also found pronounced complementarity between the AID 3'UTR and miR-93 (Figure 1A). [score:1]
We conclude that the human AID 3'UTR contains miR-155 and miR-93 recognition sites, which are separated by a 20 nucleotide adenosine repeat upstream from the true polyA sequence. [score:1]
Disruptions of miR-155 and miR-93 are associated with AID -induced oncogenesis. [score:1]
Therefore, we depleted miR-93 and miR-155 individually and then monitored changes to AID protein levels by western blotting. [score:1]
This miR-93 target site is markedly more extensive than the majority of characterized miR::mRNA interactions containing only a single, centrally located 4 bp mismatch. [score:1]
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[+] score: 179
Our study provides the first evidence that TIONFH patients have high miR-93-5p expression in the peripheral blood and that TIONFH inhibits osteoblast differentiation by reducing BMP-2 expression. [score:7]
BMP-2 is an important protein in osteogenic differentiation, and miR-93-5p is a negative regulator of BMP-2. miR-93-5p and the BMP-2 RNA 3'UTR region binding were not completely paired, and thus, the function of hBMSCs expressing BMP-2 was inhibited. [score:6]
miR-93-5p, as a member of the microRNA gene cluster miR-106b~25 (miR-106b, miR-93, and miR-25), is expressed in primary stem cells [21] and normal tissue [22] as well as in pathological contexts, such as tumour development [23], ageing [24], bone formation [25], and cardiovascular disease [26]. [score:6]
Therefore, miR-93-5p likely inhibits osteoblast differentiation via BMP-2. These observations contribute to our understanding of BMP-2 -associated miRNAs and also elucidate the mechanism of TIONFH -mediated inhibition of osteogenic differentiation. [score:5]
As with all miRNAs, miR-93-5p has many targets, and BMP-2 is just one target. [score:5]
By simulating and/or inhibiting miR-93-5p expression in human bone marrow mesenchymal stem cells, we confirmed that osteogenic differentiation-related indictors, including BMP-2, Osterix, Runt-related transcription factor, ALP and OPN, were decreased by miR-93-5p. [score:5]
Other evidence, such as the significantly reduced ALP and the formation of calcium nodules as well as the expression of BMP-2, Runx-2 and Osterix, demonstrated the inhibition of bone cell differentiation in response to miR-93-5p. [score:5]
These results demonstrate that miR-93-5p inhibits the expression of BMP-2 by binding to a specific region of the BMP-2 miRNA 3'UTR. [score:5]
It is unclear whether the effects of miR-93-5p on osteogenic differentiation depend solely/primarily on targeting BMP-2 or whether other targets are involved as well. [score:5]
Currently, although the function of miR-93-5p in promoting hBMSC proliferation and suppressing osteogenic differentiation is unclear, specific suppression of miR-93-5p in hBMSC osteogenic differentiation has been demonstrated. [score:5]
Based on comprehensive experimental results in vitro and in vivo, miR-93-5p was shown to be significantly increased in TIONFH patients, and its osteogenic suppression function was related to target genes, such as BMP-2, which may be one of the mechanisms of bone necrosis collapse. [score:5]
The above results confirmed that low expression of the osteogenic protein BMP-2 was associated with high miR-93-5p expression in TIONFH patients and decreased osteoblast differentiation. [score:5]
Is this process directly related to abnormal expression of miR-93-5p? [score:4]
Clinical significance of microRNA-93 downregulation in human colon cancer. [score:4]
miR-93-5p regulation of BMP-2 expression. [score:4]
These differences included 35 up-regulated miRNAs, such as miR-93-5p, miR-7i-5p, miR-320a, miR-25-3p, and miR-16-2-3p (fold > 2.0, P < 0.05). [score:4]
In vivo, we further validated the gene and protein levels of BMP-2. BMP-2 expression was significantly reduced in the Inducer + miR-93-5p mimic group (Fig 6B and 6C), which is consistent with the luciferase reporter gene results. [score:3]
In vitro and in vivo experiments indicated that osteogenic differentiation inhibition was associated with miR-93-5p. [score:3]
0182678.g005 Fig 5 (A) miR-93-5p targets the 3 'UTR of BMP-2. BMP-2-3'UTR WT or MUT was cloned into psiCHECK2.0 and co -transfected with the miR-93-5p mimic. [score:3]
Q-PCR results showed that the miR-93-5p expression was decreased in the NC group when they were cultured in differentiation medium after 3 and 7 d, but the miR-93-5p mimic group showed higher rates of expression compared to the other groups, indicating successful plasmid transfection (Fig 4B) The results of MTT assays indicated that the miR-93-5p mimic significantly promoted hBMSC cell proliferation in basic medium after 48 h, which was more pronounced after 72 h (p<0.05) (Fig 4C). [score:3]
Thus, based on multiple lines of evidence, miR-93-5p inhibits osteogenic differentiation by BMP-2. Interestingly, we found that miR-93-5p promoted cell proliferation of hBMSCs during in vitro experiments. [score:3]
The next day, the cells were transfected with 50 nM of miR-93-5p mimics or NC mimics and inhibitor using Lipo3000 (Thermo Scientific) according to the manufacturer’s protocol. [score:3]
These data indicated that miR-93-5p can suppress hBMSC osteogenic differentiation in vitro and suggest the important role of miR-93-5p in osteogenic differentiation. [score:3]
To further explore the mechanism of miR-93-5p in TIONFH, we analysed its target gene, BMP-2, after prediction by bioinformatics methods and luciferase reporter gene analysis. [score:3]
Identification of BMP-2 as a target of miR-93-5p. [score:3]
The cells were divided into three groups: miR-93-5p NC group, miR-93-5p mimic group and miR-93-5p inhibitor group. [score:3]
Q-PCR results showed that the miR-93-5p expression was decreased in the NC group when they were cultured in differentiation medium after 3 and 7 d, but the miR-93-5p mimic group showed higher rates of expression compared to the other groups, indicating successful plasmid transfection (Fig 4B)The results of MTT assays indicated that the miR-93-5p mimic significantly promoted hBMSC cell proliferation in basic medium after 48 h, which was more pronounced after 72 h (p<0.05) (Fig 4C). [score:3]
Further bioinformatics analysis indicated that miR-93-5p can target bone morphogenetic protein 2 (BMP-2). [score:3]
miRNA array and miR-93-5p expression. [score:3]
A partially differentially expressed miRNA cluster diagram is shown (Fig 3A), and miR-93-5p, with a fold change more than 3, was further examined. [score:3]
However, miR-93-5p was also shown to inhibit cell expansion in human colon cancer [30]. [score:3]
miR-93-5p expression during hBMSCs differentiation was also assessed. [score:3]
Online software was used to screen potential miR-93-5p targets. [score:3]
Q-PCR was used to further confirm the miR-93-5p expression and showed that miR-93-5p was significantly increased in the peripheral blood of TIONFH patients (Fig 3B), consistent with the microarray analysis. [score:3]
In vivo, BMP-2 protein expression was low in tissues of femoral head necrosis patients and had an inverse correlation with the miR-93-5p level. [score:3]
At the same time, of bone tissue showed that the expression of miR-93-5p in necrotic areas was significantly higher than that in normal areas (Fig 3C). [score:3]
do) was used to predict potential target genes of miR-93-5p. [score:3]
Comparison of the tissue samples showed that miR-93-5p expression increased, and alkaline phosphatase (ALP) and osteopontin (OPN) levels decreased, suggesting miR-93-5p may be involved in osteogenic differentiation. [score:3]
Our study showed that increased miR-93-5p in TIONFH patients inhibited osteogenic differentiation, which may be associated with BMP-2 reduction. [score:3]
Therefore, miR-93-5p may be a potential target for prevention of TIONFH. [score:3]
A 580 bp region of BMP-2 3' untranslated region (UTR), containing the potential miR-93-5p binding site cloned into the luciferase assay plasmid PSICHECK2.0, was obtained from Ginbio (Guangzhou, China) to demonstrate a direct interaction between miR-93-5p and BMP-2. All primers were synthesised by Sangon Biotech (Shanghai, China) (Table 1). [score:2]
In this study, we demonstrated that miR-93-5p posttranscriptionally regulates BMP-2 by binding to the 3'UTR. [score:2]
The miR-93-5p mimic/inhibitor, negative control mimics (NC) and primers were obtained from RiboBio Biotechnology (Guangzhou, China). [score:2]
Two cell groups (miR-93-5p NC and miR-93-5p mimic groups) were seeded on 96-well plastic dishes (Costar) to examine mineralisation. [score:1]
miR-93-5p mimics bind specifically to BMP2-3'UTR WT. [score:1]
In this study, we only assessed the effects of miR-93-5p in TIONFH, and more detailed examinations are required. [score:1]
We found that many miRNAs were significantly different, including miR-93-5p; the increase in this miRNA was verified by Q-PCR. [score:1]
These results prompted us to assess whether hBMSCs have multi-lineage differentiation potential [38] and whether miR-93-5p induces hBMSCs toward other types of differentiation. [score:1]
Q-PCR analysis 1: miR-93-5p. [score:1]
Association of miR-93-5p and osteoblast differentiation. [score:1]
Three groups of cells (miR-93-5p NC group, miR-93-5p mimic group and miR-93-5p inhibitor group) were harvested 72 h after transfection in cell lysis buffer and then assayed for luciferase activity using the Dual-Luciferase Reporter Assay System (Beyotime, Shanghai) and a luminometer according to the manufacturer’s protocol. [score:1]
To address this issue, we further verified the impact of miR-93-5p on osteoblast differentiation in cell experiments. [score:1]
The BMP2 3′-UTR with either the predicted WT miR-93-5p binding site or a mutant binding site was inserted into the psiCHECK2 vector (Fig 5A). [score:1]
These results demonstrated that miR-93-5p can bind to the 3′-UTR of the BMP-2. 10.1371/journal. [score:1]
After 7 days in differentiation medium with osteogenic induction, two cell groups (miR-93-5p NC group and miR-93-5p mimic group) were seeded on 60 mm plastic dishes (WHB) and cultured for 7 days in osteogenic differentiation medium. [score:1]
Luciferase gene reporter experiments showed that in the WT group, luciferase activity was significantly reduced due to binding to the miR-93-5p mimic, while the miR-93-5p mimic did not affect the luciferase activity of the MUT group (Fig 5C). [score:1]
We screening genes between TIONFH patients and patients without necrosis and demonstrated that miR-93-5p was increased in the subjects, which was verified by Q-PCR. [score:1]
1: miR-93-5pQ-PCR for miR-93-5p was performed according to standard protocols using a Bio-Rad CFX96 Touch [™] Deep Well Q-PCR Detection System. [score:1]
Whether increased miR-93-5p in TIONFH patients is involved in bone metabolism abnormalities caused by bone necrosis remains to be elucidated. [score:1]
However, increased miR-93-5p will have broad pleiotropic effects. [score:1]
These results demonstrated that miR-93-5p can bind to the 3′-UTR of the BMP-2. 10.1371/journal. [score:1]
Based on the complementarity with the miR-93-5p seed sequence, we focused BMP-2 (Fig 5A). [score:1]
The expression of miR-93-5p was evaluated using a mirVana [™] qRT-PCR miRNA Detection Kit (RiboBio Biotechnology, Guangzhou). [score:1]
Q-PCR for miR-93-5p was performed according to standard protocols using a Bio-Rad CFX96 Touch [™] Deep Well Q-PCR Detection System. [score:1]
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[+] score: 166
RUNX1 directly regulated miR-93 through suppressing expression and function of miR-93, miR-93 functions as a suppress gene in pancreatic cancer and regulates its target such as HMGA2. [score:12]
Overexpression of RUNX1 reduced miR-93 expression, whereas RUNX1 downregulation resulted in a marked increased miR-93 level, indicating that miR-93 is inhibited by RUNX1. [score:10]
While RUNX1 was overexpressed, miR-93 expression was downregulated (Figure 3A). [score:8]
RUNX1 overexpression promoted cell migration and invasion, while overexpression of miR-93 expression resulted in reduced cell migration and invasion. [score:7]
RT-PCR showed that when RUNX1 was knocked down, miR-93 expression was upregulated (Figure 3B). [score:7]
Despite these experimentally testified oncogenic protein-coding target genes regulated by miR-93, our study found that miR-93 also targets HMGA2. [score:6]
This result was shown in two pancreatic cell lines, suggesting a negatively regulated expression of RUNX1 and miR-93 expression in pancreatic cells. [score:6]
We examined TargetScan [22], miRanda, CLIP-Seq [23] and miRDB and found almost 100 potential miR-93 target genes. [score:5]
Moreover, restoration of miR-93 expression results in suppressing EMT process. [score:5]
Taken together, these results all indicate that overexpression of MIR-93 suppress migration and invasiveness of PDAC cells. [score:5]
Western blot of PANC-1 cells showed that miR-93 overexpression significantly decreased HMGA2 protein expression (Figure 5B). [score:5]
In triple negative breast cancer cells in vitro, miR-93 overexpression also inhibit the invasive ability [33]. [score:5]
Our study also identified that increased RUNX1 level is responsible, at least in part, for the decreased level of miR-93, explaining the downregulation of miR-93 in pancreatic cancer tissues. [score:4]
We first overexpressed and knock down RUNX1 in pancreatic cell lines (PANC-1, MIA PaCa -2) to see whether the miR-93 level can be altered. [score:4]
Quantitative PCR of miR-93 and HMGA2 were done in fresh pancreatic samples of cancers and normal tissues (Supplementary Figure 1), combining the opposite result of miR-93 and HMGA2 expression and function, we anticipated that miR-93 could work through regulating HMGA2 in PANC1 cells. [score:4]
The mechanism mediating this oncogenic effect was that by directly interacting with its promoter; subsequently miR-93 induced degradation of miR-93 target gene HMGA2 was abrogated. [score:4]
Further, we showed that overexpression of miR-93 or knocking down of HMGA2 both can decrease the invasive ability of pancreatic cancer. [score:4]
So an independent transcription factor can be an important regulator of miR-93 expression. [score:4]
We further examined the role of MIR-93 in PDAC cells by overexpressing miR-93 using MIR-93 mimics. [score:3]
Wound healing assay in PANC-1and transwell invasive assays in PANC-1and MIA PaCa-2 showed that enhanced expression of MIR-93 suppressed cell invasion in both PANC-1 and MIA PaCa-2 PDAC cells (Figure 4B and 4D). [score:3]
But our relative expression of mRNA in pancreatic tissues showed a positive correlation between miR-93 and RUNX1, this may be due to lower amount of samples. [score:3]
This possible axis of RUNX1-miR-93-HMGA2 was never reported except for one article HMGA2 was directly regulated by RUNX1 during expansion of myeloid progenitors in mice in a cell type -dependent manner instead of binding to the HMGA2 promotor [37]. [score:3]
This phenomenon was more obvious in PANC-1 cells that have higher endogenous E-cadherin and HMGA2 expression; this also explains MIA PaCa -2 didn’t show the same behavior as pobble-like cell typed PANC-1. MiR-93 was once reported to slow cell proliferation and migration through direct degradation of p21 and CCNB1 in colon cancer, thus arresting the cell cycle in G2 stage [32]. [score:3]
However, RUNX1 and miR-93 expression in pancreatic cancers and corresponding normal tissues was positive correlated (P=0.04) (Figure 3E). [score:3]
Figure 3 (A) miR-93 expression decreased in pcDNA3. [score:3]
Additionally, our data added deeper evidence that RUNX1 regulates miR-93 by directly binding to the DNA -binding sites in the miR-93 promotor. [score:3]
Epithelial-mesenchymal transition (EMT) is an early event migration of cancer, also indicating an invasive potential of tumor cells, here, we found that in PANC-1 cell lines, epithelial marker E-cadherin was greatly increased and mesenchymal markers vimentin, N-cadherin were drasticlly decreased after MIR-93 transfection on both protein and mRNA level (Figure 4E–4F), suggesting an EMT suppression status. [score:3]
MiR-93 has also been found to inhibit EMT process of breast cancer cells through regulating MKL-1 and STAT3 [34]. [score:3]
Using bioinformatics methods, we found that miR-93 is a potential target of RUNX1. [score:3]
RUNX1 negatively regulated miR-93. [score:2]
Here, we have defined an innovative oncogenic pathway downstream of RUNX1 involving miR-93 -dependent regulation of HMGA2 in PDAC. [score:2]
MiR-93 suppress migration, invasion and epithelial and mesenchymal transformation in pancreatic tumor cells. [score:2]
MiR-93 expression in normal pancreas is higher than that in pancreatic cancer tissues. [score:2]
MiR-93 target HMGA2. [score:2]
Here, we demonstrated that RUNX1 can regulate miR-93 in pancreatic cancers. [score:2]
Reporter gene plasmids harboring either the 3’-UTR region wild-type HMGA2 or a 3’-UTR mutant (Figure 5C) and either miR-93 or NC were co -transfected into PANC-1. MiR-93 overexpression reduced luciferase activity compared with the reporter gene plasmid group containing the HMGA2 3’-UTR wildtype sequence (40% reduction) (Figure 5D). [score:2]
The promotor region of miR-93 covering the binding sites were each cloned in between KpnI and BagI of pGL3 basic vector (Promega). [score:1]
This study didn’t mention miR-93 and didn’t show TF-miRNA interaction or a TF-gene interaction. [score:1]
The white box indicates miR-93 precurser. [score:1]
MiRNA mimics (for miR-93) were also synthesized and purified by Genepharma. [score:1]
This possible repression of miR-93 and HMGA2 interpreted TF-miRNA interaction and miRNA-gene interaction and provide us with insights into mechanisms underlying high levels of RUNX1 in PDAC progression. [score:1]
Other EMT related transcription factor in PANC-1 after MIR-93 transfection were also tested. [score:1]
Seq1 and Seq1, Seq2 (sequence of miR-93 promoter containing RUNX1 binding sites) and Mut1, Mut2 (sequence of miR-93 promoter containing mutated RUNX1 binding sites) (Figure 3C) were cloned into pGL3-basic named as pGL3-miR93 Seq1-WT or pGL3-miR93 Seq1-Mut1. [score:1]
The black boxes indicate the two predicted RUNX1 binding site in the promoter region of miR-93, respectively. [score:1]
Using bioinformatics methods ChIPBase [19] and GTRD [20] and review of the literature [21], we found that there are two binding sites of the miR-93 promoter region that can potentially act as a transcript site (name as Seq1 and Seq2) for RUNX1 (Figure 3C). [score:1]
Besides, miR-93 was showed to play important roles in promoting mesenchymal-epithelial (re-)transition (MET) in normal and malignant breast stem cells [35]. [score:1]
First, we analysed the relative MIR-93 levels in all pancreatic cell lines (Figure 5A) and we choose PANC-1 as the tool cells for protein degradation experiment. [score:1]
Cells with 80% confluence were transfected with pGL3-prom-miR93 (800ng per well), pRL-TK vector (200 ng per well) and pcDNA3.1 construct (RUNX1 or empty). [score:1]
This may due to the different binding ability of these two sites and also indicated that these two binding sites actually work in pancreatic cell lines and they may partially contribute to the interaction between RUNX1 and miR-93. [score:1]
First, we identified a potential negative correlation of function between RUNX1 and miR-93 in pancreatic cells. [score:1]
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[+] score: 120
Other miRNAs from this paper: hsa-mir-21, hsa-mir-345
Furthermore, miR-93 overexpression in full-length HCV replicon -expressing cells but not in full-length core -overexpressing cells suggested that other HCV proteins might up-regulate miR-93 expression. [score:12]
MicroRNA-345 down-regulates p21 [Waf1/Cip1] gene expression in human hepatoma cellsIt has been reported that human p21 [Waf1/Cip1] gene expression can be inhibited by miR-345 and miR-93 in HEK 293 cells [33]. [score:10]
The results showed that miR-345 and miR-93 were overexpressed with more than 3.5- and 2-fold changes, respectively, in mature core (173 a. a. ) and more truncated core (153 a. a. ) -overexpressing Huh7 cells but not in full-length core (191 a. a. ) -overexpressing cells compared to cells nonexpressing core (Fig. 1C, left upper panel). [score:8]
0061089.g002 Figure 2MicroRNA-345 down-regulates p21 [Waf1/Cip1] gene expression through targeting its 3′UTR but not microRNA-93 in human hepatoma cells. [score:8]
In this study, we indicated that two p21 [Waf1/Cip1] -targeting microRNAs, microRNA-345 (miR-345) and microRNA-93 (miR-93), were up-regulated in core -overexpressing Huh7 cells (Fig. 1B). [score:8]
MicroRNA-345 down-regulates p21 [Waf1/Cip1] gene expression through targeting its 3′UTR but not microRNA-93 in human hepatoma cells. [score:8]
Together, these results demonstrate that truncated HCV core proteins (amino acids 1–173 and 1–153) up-regulate cellular miR-345 and miR-93 expression in human hepatoma cells. [score:6]
HCV core protein up-regulates miR-345 and miR-93 expression in human hepatoma cells. [score:6]
It has been reported that human p21 [Waf1/Cip1] gene expression can be inhibited by miR-345 and miR-93 in HEK 293 cells [33]. [score:5]
Figure S1 MicroRNA-93 cannot down-regulate endogenous p21 [Waf1/Cip1] gene expression in human hepatoma cells. [score:5]
MicroRNA-345 and microRNA-93 are overexpressed in HCV core -overexpressing human hepatoma cells. [score:5]
Three core gene -expressing vectors for core protein with amino acids 1–191, 1–173 and 1–153, respectively, were transfected into cells, and then the relative expression of miR-345 and miR-93 was determined at 48 hours after transfection. [score:5]
The relative expressions of miR-345 and miR-93 were determined by TaqMan real-time qPCR in full-length HCV replicon -expressing Huh7 cells (right lower panel). [score:5]
As the results in luciferase reporter assay (Fig. 2A, left lower and right lower panels), p21 [Waf1/Cip1] expression at protein level was not suppressed by miR-93 in HepG2 cells (Fig. S1). [score:4]
The results showed that the treatment with miR-345 mimic led to a significant reduction of luciferase activity in p21 3′UTR Sense construct -transfected Huh7 cells, but treatment with miR-93 mimic had no significant inhibition in luciferase activity (Fig. 2A, left lower panel, second bar cluster). [score:3]
Because the different cell types might generate different results, the effects of miR-345 and miR-93 on human p21 [Waf1/Cip1] gene expression were examined in human hepatoma cells. [score:3]
At 48 hours after transfection, relative expression of miR-345 or miR-93 was determined by TaqMan real-time qPCR in Huh7 cells (left upper panel) and HepG2 cells (right upper panel). [score:3]
We further examined human p21 [Waf1/Cip1] gene expression at protein level when miR-93 mimic was transfected into HepG2 cells. [score:3]
These results demonstrated that human p21 [Waf1/Cip1] gene may be not a target of miR-93 in human hepatoma cells. [score:3]
The relative expressions of miR-345 and miR-93 were determined by TaqMan real-time qPCR. [score:3]
As expected, miR-345 and miR-93 mimics had no effect in control vector -transfected and p21 3′UTR Antisense construct -transfected Huh7 cells (Fig. 2A, left lower panel, first and third bar clusters). [score:1]
Human microRNA-345 (hsa-miR-345, MIMAT0000772) and microRNA-93 (hsa-miR-93, MIMAT0000093) mimics were purchased from Thermo Scientific. [score:1]
Three microRNAs, miR-21, miR-345 and miR93, of thirty-one microRNAs were indicated. [score:1]
Huh7 cells and HepG2 cells were transfected with p21 3′UTR sense or antisense luciferase reporter vector in combination with miR-345 mimic, miR-93 mimic or a mixture of miR-345 and miR-93 mimics. [score:1]
Moreover, treatment with a mixture of miR-345 and miR-93 mimics also had no double reduction of luciferase activity (Fig. 2A, left lower panel, second bar cluster). [score:1]
Array data were further confirmed by TaqMan real-time qPCR for miR-345 and miR-93 in Huh7 and HepG2 cells. [score:1]
HepG2 cells were transiently transfected with the increased amount of miR-93 mimic for 24 hours. [score:1]
[1 to 20 of 27 sentences]
14
[+] score: 120
The downregulated miRNAs include miR-93 (16.7 times lower), and the upregulated expression level of miR-93 significantly inhibits cell growth and coloning efficiency of colon CSCs by negatively regulating mRNA and protein expression of HDAC8 and TLE4. [score:14]
There was a 16.7 fold drop of miR-93 expression in SW1116csc, and the growth and coloning efficiency of SW1116csc were obviously inhibited by elevated expression of miR-93 [16]. [score:7]
The expression of the FLUC reporter was inhibited by endogenous LOCCS, as was the expression of miR-93 from pGLM-miR-93. [score:7]
CT: control group; OX: LOCCS-ox plasmid -transfected group; SI: siLOCCS plasmid -transfected group To determine whether the tumor inhibition of LOCCS knockdown were mediated by miR-93, miR-93 upregulation by LOCCS knockdown was rescued using Lv-si-miR-93 transfection before the evaluation of cell proliferation. [score:6]
This hypothesis is supported by our study elucidating that LOCCS serves as a competitive endogenous RNA (sponge) for miR-93, thus releasing miR-93 inhibition of target molecules, including MSI1, HDAC8 and TLE4, in CSCs. [score:5]
Taken together, our study elucidate that the lncRNA LOCCS may be a new modulator of human colon CSCs, which can exercise its functions by inhibiting miR-93 expression. [score:5]
In this study, we revealed that knockdown of LOCCS induced the upregulation of miR-93. [score:5]
In the former research, we found that miR-93 acts as a cancer suppressor in CSCs by targeting the HDAC8 and TLE4 genes [16]. [score:5]
d The inhibition ratio of three pENTR/U6-siLOCCS plasmids (Z1, Z2, Z3) of the expression of miR-93. [score:5]
Furthermore, microRNA-93 (miR-93) and Musashi-1 mediated the tumor suppression of LOCCS knockdown. [score:4]
This result suggested that miR-93 mediates the suppressive effects of LOCCS knockdown on colon cancer stem cell proliferation. [score:4]
From the above results, we conceive that LOCCS may regulate the expression of HDAC8 and TLE4 through miR-93, and it may also take part in the Notch and Wnt signaling pathways through MSI1. [score:4]
Furthermore, miR-93 and MSI1 mediated the tumor suppression of LOCCS knockdown. [score:4]
MiR-93 and MSI1 mediated the tumor-suppressive effects of LOCCS knockdown on CD133+/CD166+/CD44+ spheroid cells. [score:3]
Additional file 1: Sequences of long intergenic non-protein-coding RNA 1567 (LOCCS) (DOC 27 kb) Additional file 2: Sequences of pENTR/U6-Z1, pENTR/U6-Z2, pENTR/U6-Z3, pGL3M-miR-93, pcDNA-LOCCS and pcDNA-LOCCS-T plasmids (DOC 245 kb) CRC colorectal carcinoma CR-CSCs CRC stem cells CSCs cancer stem cells FITC fluorescein isothiocyanate FLUC firefly luciferase GAPDH glyceraldehyde-3-phosphate dehydrogenase IC50 50% reduction in cell viability LINC01567 long intergenic non-protein-coding RNA 1567 lncRNAs long non-coding RNAs LOCCS lncRNA overexpressed in colon cancer stem cells miR-93 microRNA-93 MSI1 musashi-1 PBS phosphate-buffered saline PCC primary cultured cells PE phycoerythrin SFM serum-free medium siRNA small interfering RNA SSM serum-supplemented medium TPSC triple positive spheroid cells Not Applicable. [score:3]
However, when the endogenous LOCCS was degraded by siLOCCS transcribed from the pENTR/U6-siLOCCS plasmid, the inhibition of miR-93 by LOCCS was weakened, and thus FLUC levels were higher (A vs. [score:3]
The transfected cells were harvested 48 h later, and expression level of miR-93 was mensurated using quantitative PCR. [score:3]
Fig. 2Expression of LINC01567 and its interaction with miR-93. [score:3]
When cells were cotransfected with pcDNA-LOCCS, miR-93 was inhibited by both endogenous and exogenous LOCCS, and, thus, the levels of miR-93 in this group were the lowest among the groups (D vs. [score:3]
In the Lv-si-LOCCS + Lv-si-miR-93 group, Lv-si-miR-93 rescued the suppression of Lv-si-LOCCS on cell growth (Fig. 6). [score:3]
Research on mechanisms suggested direct binding, as a predicted miR-93 binding site was identified in LOCCS. [score:2]
However, lncRNAs that may competitively regulate miR-93 in CR-CSCs have not yet been identified. [score:2]
The observation that if the binding sequence in LOCCS was mutated, it could not combine with miR-93 confirmed that LOCCS acts on miR-93 directly. [score:2]
Searching for lncRNAs with an miR-93 binding site, we found LOCCS. [score:1]
In contrast, LOCCS-T transcribed from the pcDNA-LOCCS-T plasmid could not combine with miR-93. [score:1]
b Colony formation rates of TPSC transfected with the Lv-si-LOCCS + Lv-si-miR-93 or Lv-si-LOCCS + Lv-si-MSI1 plasmids. [score:1]
The human LOCCS, miR-93, and MSI1-specific siRNA sequences were designed and synthesized by Shanghai Haike Corporation. [score:1]
Construction of the pGL3M-miR-93 luciferase reporter plasmid. [score:1]
a Growth curves of TPSC transfected with the Lv-si-LOCCS + Lv-si-miR-93 or Lv-si-LOCCS + Lv-si-MSI1 plasmids. [score:1]
The interaction between LOCCS and miR-93 in CD133+/CD166+/CD44+ spheroid cells. [score:1]
LINC01567 MicroRNA-93 Colon Cancer stem cells Regulation Colorectal carcinoma (CRC) is in the third of malignant tumor in men and second in women, and in 2008, there were about 1.2 million new patients and 600,000 death cases [1]. [score:1]
For endogenous LOCCS analysis (no exogenous LOCCS transfection), A: 400 ng pGL3M-miR-93 + 400 ng pENTR/U6-si-LOCCS +500 ng pRL-CMV; B: 400 ng pGL3M-miR-93 + 500 ng pRL-CMV; C: 400 ng pGL3M + 500 ng pRL-CMV; for exogenous LOCCS analysis, D: 400 ng pGL3M-miR-93 + 400 ng pcDNA-LOCCS +500 ng pRL-CMV; E: 400 ng pGL3M-miR-93 + 400 ng pcDNA-LOCCS-T + 500 ng pRL-CMV; F: 400 ng pGL3M-miR-93 + 500 ng pRL-CMV. [score:1]
We hypothesized that LOCCS may act as a competitive RNA for miR-93 in CSCs. [score:1]
For the luciferase reporter vector construction, the pre-miR93 sequence was synthesized with added XbaI sites by Shanghai Haike Corporation. [score:1]
Using lncRNA interaction analysis software (Starbase v2.0), we confirmed that LOCCS could bind with miR-93, and the binding region is shown in Fig. 2c. [score:1]
Using bioinformatics analysis and luciferase reporter assays, we elucidated the direct binding site of miR-93 in LOCCS. [score:1]
48 h later, the lentiviral particles were harvested using 50,000 × g ultracentrifugation for 2 h, and they are named as Lv-si-LOCCS, Lv-si-miR-93, Lv-si-MSI1 and Lv-si-NC (negative control). [score:1]
c The predicted binding sites between LOCCS and miR-93. [score:1]
To confirm the interaction between LOCCS and miR-93, we synthesized pGL3M-miR-93, pcDNA-LOCCS and pcDNA-LOCCS-T plasmids and transfected them into TPSC. [score:1]
There was reciprocal repression between LOCCS and miR-93. [score:1]
The miR-93 levels were similar to those of the group without exogenous LOCCS (E vs. [score:1]
[1 to 20 of 41 sentences]
15
[+] score: 112
Other miRNAs from this paper: mmu-let-7g, hsa-let-7g, mmu-mir-93
However, the miRNAs whose expression levels changed may play crucial roles in the regulation of target gene expression without affecting transcript expression levels, for example, targeting of the MICA protein by miRNA93, whose expression levels were downregulated by HBV replication. [score:17]
Since the expression levels of miRNA93 were downregulated by HBV replication (Figure 1d and Supplementary Table 4), we delivered miRNA93 via BNCs into HBV replicating human hepatocytes to rescue the downregulation of miRNA93 levels and examine the effects of decreased miRNA93 on transcript levels (Figure 3b). [score:9]
The rescue of miRNA93 expression, recovered the baseline-level expression of some genes, such as 17-beta-hydroxysteroid dehydrogenase 14 (HSD17B14) and tripartite motif-containing protein 31 (TRIM 31), which were increased by HBV replication (Supplementary Table 1), suggesting that the mRNA levels of these genes may be directly or indirectly regulated by miRNA93. [score:8]
Thus, the decreased expression of miRNA93 by HBV suggested that the regulation of MICA expression by miRNA93 has biological significance. [score:6]
Although it was found that miRNA93 expression levels decreased during HBV replication in primary hepatocytes (Figure 1d and Supplementary Table 4), MICA transcript levels were not affected (GEO accession number: GSE55928), suggesting that the effects of miRNA93 on MICA may be mediated by translational repression and not by mRNA decay, as we reported previously [23]. [score:5]
The increased levels of MICA protein expression agreed with the decreased miRNA93 expression. [score:5]
Among these miRNAs, miR93-5p was significantly downregulated during HBV replication by more than 50%. [score:4]
We previously identified miRNA93 as a critical regulator of MICA protein expression [23], which plays a role in the susceptibility to HBV -induced HCC [25]. [score:4]
Since miRNA93 regulates the expression levels of the MICA protein [23, 29], which is involved in the susceptibility to hepatocellular carcinoma in chronic hepatitis patients [24, 25], we focused on this miRNA in further analyses. [score:4]
Flow cytometric analysis of membrane-bound MICA protein expression in cells delivered BNC -mediated control (green line), let7g (blue line), or miRNA93 (red line). [score:3]
We report that HBV replication in human hepatocytes decreases miRNA93 expression and increases soluble MICA levels. [score:3]
However, these miRNA93 delivery results may not be accurate due to direct or indirect effects of miRNA93. [score:3]
d, miRNA93 expression in Huh7 cells after the delivery of miRNA93 via BNCs. [score:3]
MiRNA93 is a critical regulator of MICA protein expression [23, 29]. [score:3]
Modulation of MICA protein expression levels by delivery of miRNA93 using BNCs. [score:3]
Only a small increase in miRNA93 expression levels was observed 24 hours after transfer into 293T cells, based on Northern blots (Figure 2c), indicating that the BNCs had high specificity for hepatocyte-derived cells. [score:3]
These results suggest that miRNA93 delivery into HBV-replicating hepatocytes using BNC methods may enhance HBV immune clearance or suppress HCC by altering miRNA93 levels in HBV-infected cells. [score:3]
We found that BNCs carrying synthesized miRNA93 could efficiently restore deregulated soluble MICA protein levels in the supernatant of HBV-replicating primary hepatocytes. [score:2]
Soluble MICA protein levels were regulated by miRNA93 in human primary hepatocytes. [score:2]
were sequentially collected after incubation with BNCs containing miRNA93 and subjected to Northern blotting. [score:1]
Increased soluble MICA levels in the serum are strongly associated with HBV-related HCC [25], and the increased soluble MICA levels could be antagonized by the delivery of miRNA93 into hepatocytes using BNCs. [score:1]
The day after delivery of the BNCs, cells were collected and subjected to Northern blotting against let-7g, miRNA93, and U6, the loading control, and the results showed successful delivery of miRNAs into all cell lines tested (Figure 2a). [score:1]
Twenty-four hours after transfection, cells were mixed with BNCs containing let7g (black bar), miRNA93 (gray bar), or negative control (white bar). [score:1]
After 24 hours, cells were subjected to Northern blotting for miRNA93. [score:1]
Liver cancer cell lines, Huh7 and Hep3B, and primary hepatocytes immortalized by SV40, Fa2N4, were incubated with BNCs harboring the indicated miRNAs (miRNA93 or let7g) or BNCs without miRNAs (NC). [score:1]
MiRNA93 regulates MICA protein levels, but not transcript levels [23, 29]. [score:1]
Thus, we examined the effects of delivery of BNCs carrying miRNA93 into HBV-infected hepatocytes. [score:1]
Based on these results, BNCs carrying miRNA93 can be used to eliminate HBV-infected hepatocytes, which may be a novel approach for the prevention of subsequent virus -induced HCC. [score:1]
Thus, BNCs carrying miRNA93 may be used to prevent HCC in patients with chronic HBV infection. [score:1]
Hybridization was performed overnight at 42°C in ULTRAhyb-Oligo Buffer (Ambion) containing a biotinylated probe specific for miRNA93 (CTA CCT GCA CGA ACA GCA CTT TG) and let-7g (AAC TGT ACA AAC TACT ACC TCA), which was heated to 95°C for 2 min prior to hybridization. [score:1]
These results suggested that miRNA93 delivery into the liver decreases soluble MICA levels in the serum, which may be used to prevent HCC in chronic hepatitis B patients. [score:1]
Membranes were re-probed for let7g, miRNA93, and U6 as a loading control. [score:1]
The arrow indicates the result for miRNA93. [score:1]
b, A scatter plot reflecting the transcriptome results between the control and primary human hepatocytes treated with BNCs containing miRNA93. [score:1]
The 293T cells (human embryonic kidney cells) were incubated with BNCs containing let7g, miRNA93, or negative control (NC). [score:1]
To confirm the efficiency of delivery of miRNAs into liver-derived cells by BNCs, we delivered BNCs carrying let-7g or miRNA93 to the human hepatocellular carcinoma cell lines, Huh7 and HepG2 cells, and to human normal hepatocytes immortalized with SV40 large T antigen, Fa2N4 cells [28]. [score:1]
The incorporation of miRNAs (miRNA93 or let-7g) into the hollow space and the delivery of miRNAs into human liver cells were performed as described previously [31]. [score:1]
The firefly luciferase reporter plasmid was used to examine let7g and miRNA93 function. [score:1]
Even though the MICA mRNA levels were not significantly affected by miRNA93 delivery based on microarray results (GEO accession: GSE55928), soluble MICA protein in the supernatant significantly decreased according to ELISA (Figure 4b). [score:1]
b, Soluble MICA protein levels in the supernatants of primary hepatocytes after delivery of the indicated miRNAs (let7g or miRNA93) or negative control (NC) with or without HBV replication. [score:1]
Human primary hepatocytes isolated from chimeric mice were incubated with BNCs containing the indicated miRNAs (miRNA93 or let7g) or BNCs without miRNAs (NC), with or without Polybren. [score:1]
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[+] score: 110
Furthermore, we showed that targeted inhibition of miR-93-5p in human Hep3B HCC cells with anti-miR-93-5p reduced the oncogenic cancer cell phenotype, as evidenced by decreased cell viability, decreased colony formation, and increased sensitivity of the cells to the multikinase inhibitor Sorafenib, the only current effective systemic chemotherapeutic agent for the treatment of advanced HCC [14, 46]. [score:7]
Importantly, transfection of Hep3B cells with anti-miR-93-5p reduced the expression of the MCM7 gene (Figure 5C), cell viability (Figure 5D), increased the sensitivity to Sorafenib, a multikinase inhibitor used to treat HCC (Figure 5D), and inhibited colony formation (Figure 5E). [score:7]
This suggestion corresponds to the results of our study showing that over -expression of the miR-106b∼25 cluster in NASH-derived HCC, especially miR-93-5p, is accompanied by substantial decrease in the levels of E2F1, PTEN, and CDKN1A proteins, direct targets of miR-93 [19, 42, 43]. [score:6]
Figure 5B shows that after the 72-hour transfection, the level of miR-93 in Hep3B cells was greatly reduced, whereas the level of tumor suppressor protein PTEN, a direct target of miR-93 [19], significantly increased. [score:6]
Importantly, several of the identified miRNAs, including miR-34a, miR-93-5p, miR-221-3p, and miR-222-3p, were also significantly over-expressed in human HCC suggesting that aberrant expression of these miRNAs may serve as an indicator of the hepatocarcinogenic process. [score:5]
Mechanistically, the over -expression of miR-25-3p, miR-93-5p, and miR-106b-5p in NASH-derived HCC may be attributed to an increased expression of the Mcm7 gene, which harbors the miR-106b∼25 cluster [16– 18]. [score:5]
In particular, we identified 10 over-expressed miRNAs (miR-17-5p, miR-221-3p, miR-93-5p, miR-25-3p, miR-181b-5p, miR-106b-5p, miR-186-5p, miR-222-3p, miR-15b-5p, and miR-223-3p; Figure 2A) that are involved in the activation of major liver carcinogenesis-related gene expression networks, especially the TGF-β- and Wnt/β-catenin signaling pathways, the roles of which are well-established in hepatocarcinogenesis [14]. [score:5]
Among the miRNAs distinctively over-expressed in NASH-derived HCC, miR-221-3p and miR-222-3p, which exhibited a carcinogenesis stage -dependent increase in expression, and miR-25-3p, miR-93-5p, and miR-106b-5p, which are members of the oncogenic miR-106b∼25 cluster, are of special interest. [score:5]
Additionally, it has been demonstrated that miR-221-3p and miR-222-3p also target PTEN and CDKN1A [44, 45], indicating that the oncogenic activity of miR-93-5p, miR-221-3p, and miR-222-3p may be attributed to the silencing of these key cancer-related genes and consequent impairment in cell-cycle arrest and inhibition of apoptosis. [score:5]
We also demonstrated that over -expression of miR-25-3p, miR-93-5p, miR-106b-5p, miR-221-3p, and miR-222-3p was accompanied by the reduced protein levels of their targets, including E2F1, PTEN, and CDKN1A. [score:5]
The expression of the MCM7 and MIR93 genes was determined by quantitative reverse-transcription PCR (qRT-PCR) using TaqMan gene expression assays (Life Technologies, Grand Island, NY). [score:4]
Figure 3 shows that the expression of four miRNAs, miR-34a-5p, miR-93-5p miR-221-3p, and miR-222-3p, that were over-expressed in mouse NASH-derived HCC was also significantly greater in human HCC (n = 358) as compared to non-tumor liver tissue samples (n = 50). [score:4]
This suggestion is based on the results of the present study showing a concomitant up-regulation of the Mcm7 gene and the miR-106b∼25 cluster in NASH-derived HCC, and a positive correlation between the level of miR-93-5p and the Mcm7 transcript during mouse liver carcinogenesis and in human HCC. [score:4]
Among the differentially expressed miRNAs in NASH-derived HCC, three miRNAs, miR-106b, miR-93, and miR-25, are members of the oncogenic miR-106b∼25 intragenic cluster [15, 16]. [score:3]
Figure 5 (A) Expression of miR-93 in human liver cancer cell lines. [score:3]
Furthermore, Figure 4C shows that Mcm7 expression strongly positively correlated (r = 0.70, P = 0.03) with the miR-93-5p level. [score:3]
The expression of miR-93 in α-fetoprotein -positive PLC/PRF/5, Hep3B, and HepG2 cells is presented as an average fold relative to that in α-fetoprotein -negative SK-Hep-1 cells, which were assigned a value 1. * - denotes a significant (P < 0.05) difference from SK-Hep-1 cells. [score:3]
The statistical analyses of miR-34a-5p, miR-93-5p, miR-221-3p, and miR-222-3p expression datasets in human HCC samples were conducted by the Mann-Whitney Rank Sum test. [score:3]
Importantly, the expression of the MCM7 gene was also increased in human HCC (Figure 4D) and positively correlated (r = 0.69, P = 4.25 x 10 [-6]) with the level of miR-93-5p (Figure 4E). [score:3]
To determine whether or not inhibition of miR-93-5p decreases the oncogenic phenotype of cancer cells, four human liver cancer cell lines (SK-Hep-1, PLC/PRF/5, Hep3B, and HepG2) were screened for miR-93 levels (Figure 5A). [score:3]
Importantly, miR-93-5p, miR-221-3p, and miR-222-3 were significantly over-expressed in human HCC. [score:3]
Pearson product-moment correlation coefficients were used to determine the relationship between miR-93-5p levels and the Mcm7 gene expression. [score:3]
Among these miRNAs, the over -expression of ten miRNAs (miR-15b-5p, miR-17-5p, miR-25-3p, miR-93-5p, miR-106b-5p, miR-181b-5p, miR-186-5p, miR-221-3p, miR-222-3p, and miR-223-3p) was associated with the activation of major hepatocarcinogenesis-related pathways, including the TGF-β, Wnt/β-catenin, ERK1/2, mTOR, and EGF signaling. [score:3]
For example, miR-17-5p, located in the miR-17∼92 cluster, and miR-93-5p and miR-106b-5p, located in the miR-106b∼25 cluster, belong to the same miR-17 family [33]. [score:1]
Since Hep3B cells exhibited the highest level of miR-93 among the human liver cancer cell lines, this cell line was transfected with 50 nM of anti-miR-93-5p. [score:1]
Transfection of Hep3B cells with anti-miR-93-5p. [score:1]
Effect of anti-miR-93 transfection on human Hep3B hepatocellular carcinoma cells. [score:1]
After 48 hours of incubation, 7.5 × 10 [3] viable Hep3B cells that had been transfected with either anti-miR-93 or scrambled RNA oligonucleotide were seeded onto 0.6% noble agar in growth media. [score:1]
Importantly, five of these miRNAs (miR-34a-5p, miR-93-5p, miR-106b-5p, miR-221-3p, and miR-222-3p) were in common with those in the 10-miRNA set in our study. [score:1]
Levels of miR-34a-5p, miR-93-5p, miR-221-3p, and miR-222-3p in human HCC samples. [score:1]
To investigate the functional consequences of the miR-106b∼25 cluster over -expression with respect to the hepatocarcinogenic process, the levels of E2F1, PTEN, and CDKN1A proteins, experimentally confirmed targets of miR-106b, miR-93-5p, and miR-25, were evaluated. [score:1]
Transient transfection of Hep3B HCC cells with anti-miR-93 increases cell death and drug sensitivity and decreases colony formation. [score:1]
[#] - denotes a significant (P < 0.05) difference from Hep3B cells transfected with anti-miR-93 and treated with Sorafenib alone (n = 3). [score:1]
The Hep3B cells were seeded in 100 mm dishes at a density of 1 × 10 [6] cells/dish, and transfected with 50 nM of either anti-miR-93-5p (n = 3; Life Technologies) or scrambled RNA oligonucleotide (n = 3) using Lipofectamin™ 3000 transfection reagent (Life Technologies) according to the manufacturer’s instructions. [score:1]
To determine drug sensitivity, 72 hours after being transfected with scrambled RNA oligonucleotide or anti-miR-93, adherent Hep3B cells were harvested by mild trypsinization, washed in PBS, plated at a density of 7.5 x 10 [3] cells/well in 96-well plates, and the transfection was repeated. [score:1]
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17
[+] score: 110
Other miRNAs from this paper: hsa-mir-17, hsa-mir-20a, hsa-mir-25, hsa-mir-106b, hsa-mir-375
We identified miRNAs targeting CIC from the miRNAs known to be overexpressed in prostate cancer tissues, and proposed that miR-93, miR-106b, and miR-375 could potentially contribute to the down-regulation of CIC levels in the process of prostate cancer progression. [score:8]
Moreover, disruption of the putative miRNA binding sites in the 3′UTR of CIC abrogated suppression of luciferase activity by the three miRNAs (Figure 6E), demonstrating that miR-93, miR-106b, and miR-375 directly target the 3′UTR of CIC to regulate CIC levels. [score:7]
Overexpression of miR-93, miR-106b, and miR-375 increased cell proliferation and invasion (Figures 7B and 7C and Supplementary Figure 16), accompanied with down-regulation of CIC levels (Figure 7A), suggesting the cancer promoting property of these three miRNAs in prostate cancer cells. [score:6]
The miRNAs -mediated increases in cell proliferation and invasion were partially abolished by recovery of CIC levels in PC-3 cells (Figures 7B and 7C and Supplementary Figure 16), suggesting that miR-93, miR-106b, and miR-375 promote prostate cancer progression in part by down-regulation of CIC expression. [score:6]
We also measured CRABP1 levels in the same set of cells, and found that co -expression of miR-93, miR-106b, and miR-375 resulted in up-regulation of CRABP1, which was restored by overexpression of CIC in PC-3 cells (Figure 7D). [score:6]
In fact, among the two putative miR-17 family miRNA binding sites in the CIC 3′UTR, the first site is predicted to be more preferentially targeted by miR-93, whereas the second one by miR-106b, according to the miRNA target prediction databases (Supplementary Figure 15B). [score:5]
Comparative analysis on the selected miRNAs identified five miRNAs, miR-20a, miR-25, miR-93, miR-106b, and miR-375, which not only potentially target CIC, but are also known to be frequently overexpressed in prostate cancer cells (Supplementary Figure 15A). [score:5]
However, co-transfection with all three miRNA duplexes markedly down-regulated CIC levels in PC-3 cells (Figures 6B and 6C), indicating that miR-93, miR-106b, and miR-375 cooperatively regulate CIC levels. [score:5]
For clonogenic assay of PC-3 cells treated with miRNA duplexes and CIC-S expressing lentivirus, 5 × 10 [3] PC-3 cells were seeded in six well plates a day before transfection, and then co -transfected with miR-93, miR-106b, and miR-375 duplexes using Dhamafect 2. After 24 h, the cells were infected with lentivirus expressing CIC-S for 3 sequential days. [score:4]
miR-93, miR-106b, and miR-375 cooperatively down-regulate CIC levels. [score:4]
All error bars show s. e. m. To determine the impact of the miRNAs -mediated down-regulation of CIC on prostate cancer progression, we assessed cell proliferation and invasion in PC-3 cells transfected with either control, miR-93/miR-106b/miR-375 or siCIC duplexes. [score:4]
Of the five miRNAs, we initially chose to evaluate miR-93, miR-106b, and miR-375, considering the number of putative binding sites for each miRNA in the 3′UTR of CIC and their frequency of overexpression in prostate cancer patients, and tested whether these miRNAs can down-regulate CIC levels. [score:4]
miR-93, miR-106b, and miR-375 cooperatively down-regulate CIC levels in PC-3 cells. [score:4]
Inhibition of miR-375, but not miR-93 and miR-106b, significantly increased CIC levels (Figure 6D), suggesting that, among the three miRNAs, miR-375 is the most critical endogenous miRNA for regulation of CIC levels in PC-3 cells. [score:4]
Co-transfection with three miRNAs decreased luciferase activity in PC-3 cells (Figure 6E), suggesting that miR-93, miR-106b, and miR-375 down-regulate CIC levels through the 3′UTR of CIC. [score:4]
We also examined whether CIC expression is regulated by endogenous miR-93, miR-106b, and miR-375 in PC-3 cells. [score:4]
All error bars show s. e. m. D. analysis for changes in CIC levels by inhibition of endogenous miR-93, miR-106b or miR-375 in PC-3 cells and its quantification. [score:3]
On the other hand, the three miRNAs still slightly repressed luciferase activity derived from the pGL3-CIC 3′UTR Mut compared with control vector (Figure 6E), implying that there might be other binding sites for miR-93, miR-106b, and miR-375 in the 3′UTR of CIC, or that the three miRNAs might also be able to repress CIC expression indirectly. [score:3]
B. analysis for changes in CIC levels by overexpression of miR-93, miR-106b, and miR-375 in PC-3 cells. [score:3]
To verify that miR-93, miR-106b, and miR-375 directly target the 3′UTR of CIC, we constructed luciferase reporter gene linked to the CIC 3′UTR (pGL3-CIC 3′UTR WT), and carried out dual luciferase assays. [score:3]
We did not observe such effect when different combinations of two miRNA duplexes were co -transfected (Figure 6B), suggesting that miR-93 and miR-106b may not function redundantly to regulate CIC levels, although they share the same seed sequences. [score:2]
miR-93, miR-106b, and miR-375 cooperatively regulate CIC-CRABP1 axis to promote prostate cancer progression. [score:2]
Taken together, these data demonstrate that miR-93, miR-106b, and miR-375 function cooperatively to regulate the CIC-CRABP1 axis in promoting prostate cancer progression. [score:2]
For cell growth assay of PC-3 cells treated with miRNA duplexes and CIC-S expressing lentivirus, 1 × 10 [3] PC-3 cells were seeded in 24 well plates a day before transfection, and then co -transfected with miR-93, miR-106b, and miR-375 duplexes using Dhamafect 2 and set as day “0”. [score:2]
miR-93, miR-106b, and miR-375 co-regulate CIC-CRABP1 axis to promote cancer progression in PC-3 cells. [score:2]
In sum, our findings suggest that miR-93/miR-106b/miR-375-CIC-CRABP1 is a novel regulatory axis in prostate cancer progression (Figure 7E). [score:2]
There are two putative binding sites for miR-20a/miR-93/miR-106b, one for miR-25 and another for miR-375 in the 3′UTR of CIC (Supplementary Figure 15B). [score:1]
Figure 6 in PC-3 cells A. analysis for CIC levels in PC-3 cells transfected with control, miR-93, miR-106b, or miR-375 duplexes. [score:1]
Comparative miRNA profiling of prostate carcinomas with increasing tumor stages has revealed that levels of miR-375 and miR-106b gradually increase from normal to lymph node metastasizing tumors, whereas miR-93 increases from normal to extracapsular growing tumors [36], suggesting that these miRNAs are likely to participate in the gradual decrease in CIC levels during prostate cancer progression. [score:1]
Since the seed sequences of miR-20a, miR-93, and miR-106b are identical, they are classified as the same miRNA family (miR-17 family). [score:1]
All error bars show s. e. m. A. analysis for CIC levels in PC-3 cells transfected with control, miR-93, miR-106b, or miR-375 duplexes. [score:1]
Sequences are as follows: miR-93 sense; 5′-CAAAGUGCUGUUCGUGCAGGUAG-3′, and antisense; 5′-ACCUGCACGAACAGCACUUUAUU-3′. [score:1]
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18
[+] score: 110
Other miRNAs from this paper: hsa-mir-17
We hypothesized that miRNA may regulate leptin -mediated OSM expression, using online computational algorithms (TargetScan) and filtering out seven candidate miRNAs that target OSM, to find that miR-93 was mostly downregulated by leptin treatment (Figure 3A); leptin concentration -dependently decreased miR-93 expression (Figure 3B). [score:13]
Treatment with the Akt inhibitor or transfection of Akt siRNA reversed leptin -inhibited miR-93 expression (Figure 5), i. e., leptin increases OSM production by inhibiting miR-93 expression via the Akt signaling pathway. [score:11]
We hypothesized that miRNA mediated leptin-increased OSM production, finding that leptin decreased miR-93 expression most, but only slightly affecting the expression of miR-17, -20a, -20b, -106a, -106b and -519d, which target OSM. [score:7]
Figure 3Leptin increases OSM expression through inhibition of miR-93 expression. [score:7]
The results show that leptin increases OSM expression by downregulating miR-93 through the Akt signaling pathway. [score:6]
Although some molecular targets are documented, the role of miR-93 in OSM expression is largely unknown. [score:5]
Leptin Increases OSM Production in Osteoblasts by Inhibition of miR-93 Expression. [score:5]
We tested to see if Akt is upstream in leptin -inhibited miR-93 expression. [score:5]
Leptin enhances OSM production in human osteoblasts by inhibition miR-93 expression through the Akt signaling pathway. [score:5]
We also showed the potentiation of OSM activated by leptin through inhibiting miR-93 expression via Akt signal pathway in osteoblasts. [score:5]
By contrast, cell incubation with the Akt inhibitor or siRNA abolished leptin-reduced miR-93 expression. [score:5]
Data suggest that leptin increases OSM production by inhibiting miR-93 expression. [score:5]
These indicate that leptin increased OSM yield by inhibiting miR-93 expression through the Akt pathway. [score:5]
Figure 5Leptin increases OSM expression by inhibition miR-93 through the Akt pathway. [score:5]
Xu Y. Jin H. Yang X. Wang L. Su L. Liu K. Gu Q. Xu X. MicroRNA-93 inhibits inflammatory cytokine production in LPS-stimulated murine macrophages by targeting IRAK4 FEBS Lett. [score:4]
Osteoblasts were pretreated with Akt inhibitor (10 µM) (A) for 30 min or transfected with Akt siRNA (B) for 24 h followed by stimulation with leptin (30nM) for 24 h; miR-93 expression was measured by qPCR. [score:3]
The results showed that leptin increases OSM production by binding to the OBRl receptor and activating Akt signaling, which reduces miR-93 expression and leads to the transactivation of OSM production (Figure 6). [score:3]
We also used a miR-93 mimic to confirm the role of miR-93, finding diminished leptin-enhanced OSM expression. [score:3]
To affirm miR-93 involvement in leptin-increased OSM production, an miR-93 mimic of cells reversed leptin-increased OSM mRNA and protein expression (Figure 3C,D). [score:3]
We obtained control miRNA, the miR-93 mimic and Lipofectamine 2000 from Life Technologies (Carlsbad, CA, USA), as well as the Akt inhibitor (sc-394003), rabbit polyclonal antibodies for p-Akt and Akt; both Akt and the control siRNA were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). [score:3]
In addition, miR-93 has been reported to be a negative regulator of the immune response [24]. [score:2]
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[+] score: 84
b miRNA RT-qPCR analysis showing the complete regulation of Hsa-miR-93, Hsa-miR-20a, Hsa-miR-125b and Hsa-miR-27b and, significant increase in the expression of Hsa-miR-1260 and Hsa-miR-1224-3p in metastatic tumors as compared to the non-metastatic xenograft To validate the altered expression levels observed with whole genome miRNA array, we examined the expression levels of select miRNAs including hsa-miR-1260, hsa-miR-1224-3p (showing significant upregulation; see Fig.   2), hsa-miR-93, hsa-miR-20a, hsa-miR-125b, hsa-miR-27b (showing significant downregulation; see Fig.   3a), using individual miRNA QPCR analysis. [score:13]
b miRNA RT-qPCR analysis showing the complete regulation of Hsa-miR-93, Hsa-miR-20a, Hsa-miR-125b and Hsa-miR-27b and, significant increase in the expression of Hsa-miR-1260 and Hsa-miR-1224-3p in metastatic tumors as compared to the non-metastatic xenograftTo validate the altered expression levels observed with whole genome miRNA array, we examined the expression levels of select miRNAs including hsa-miR-1260, hsa-miR-1224-3p (showing significant upregulation; see Fig.   2), hsa-miR-93, hsa-miR-20a, hsa-miR-125b, hsa-miR-27b (showing significant downregulation; see Fig.   3a), using individual miRNA QPCR analysis. [score:13]
In order to validate the miRNA expression obtained from whole genome profiling, expression of selected metastamiRs, including hsa-miR-1224-3p, hsa-miR-1260 (both significantly upregulated), hsa-miR-125b, hsa-miR-27b, hsa-miR-93,and hsa-miR-20a (all significantly downregulated) were confirmed using QPCR. [score:11]
To define the effect of characterized metastamiRs on the putative target proteins, we adopted two approaches: (i) inhibited hsa-miR-1224-3p or hsa-miR-1260 (both significantly upregulated) and (ii) functionally mimicked hsa-miR-125b, hsa-miR-27b, hsa-miR-93 or hsa-miR-20a (all significantly downregulated) and examined for the miRNA -dependent modulations in protein targets. [score:11]
First, MSDACs transiently transfected with mimics for hsa-miR-125b, hsa-miR-27b, hsa-miR-93 or hsa-miR-20a (those exhibited complete suppression in aggressive disease) and examined for the regulation of their corresponding target proteins (Fig.   6a). [score:8]
Thus, we validated our microarray results with RT-qPCR for upregulation (Hsa-miR-1260; Hsa-miR-1224-3p) and downregulation (Hsa-miR-20a, Hsa-miR-27b, Hsa-miR-125b, Hsa-miR-93) profiles (see Fig.   3b). [score:7]
miRNA mimic (hsa-miR-125b, hsa-miR-27b, hsa-miR-93, hsa-miR-20a) and inhibitor (hsa-miR-1224-3p, hsa-miR-1260) approach for select miRNAs revealed the direct influence of the altered metastamiRs in the regulation of identified protein targets. [score:7]
Transient transfection of MSDACs with hsa-miR-125b-, hsa-miR-27b-, hsa-miR-93- or hsa-miR-20a- mimics (MISSION® microRNA Mimics, Sigma-Aldrich) as well as hsa-miR-1224-3p- and hsa-miR-1260 -inhibitors (MISSION® Synthetic miRNA Inhibitors, Sigma-Aldrich) were carried out by using either TurboFectin 8.0 reagent (Origene) or Neon electroporation transfection system (Life Technologies). [score:5]
b Histograms of mean cell–Alexa Fluor intensity obtained from Columbus automated batch analysis showing alterations in the expression (i) GRB10, MMP2, p38, STAT3, TNFα and VEGF in cells with hsa-miR-125b mimic, (ii) EGFR FOSB, kRAS, p38, PTPN3 and VEGF in hsa-miR-27b mimic transfected cells, (iii) ASK1, CREB, MMP2, MMP3/10, PTPN3, STAT3and VEGF in MSDACs with hsa-miR-20a mimic and, (iv) MMP2, MMP3/10, PTPN3 and STAT3 with hsa-miR-93 mimic in MSDACs. [score:3]
Moreover, hsa-miR-93 mimic exhibited statistically significant inhibition of MMP2, MMP3/10, PTPN3 and STAT3 in this setting (Fig.  6b iv). [score:3]
Compared to the non-metastatic xenograft, we observed a complete (P < 0.001) decrease in the expression of hsa-miR-93, hsa-miR-20a, hsa-miR-125b, and hsa-miR-27b (Fig.   3b). [score:2]
Of the 74 metastamiRs identified in this study, we found an overlap of 16 metastamiRs, including Hsa-miR-148b, Hsa-miR-23a, Hsa-miR-100, Hsa-miR-93, Hsa-miR-125b, Hsa-miR-98, Hsa-miR-92a, Hsa-miR-29b, Hsa-miR-30c, Hsa-let-7a, Hsa-let-7b, Hsa-let-7c, Hsa-let-7e, Hsa-let-7f, and Hsa-let-7g, with the findings of other researchers. [score:1]
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20
[+] score: 79
Upregulation of p21 [WAF1] gene expression in UVB -treated cells was correlated with the downregulation of miRNAs 20a/b and 93, known to target p21 [WAF1][27, 30, 44- 46], and the identification of E2F1 and VEGFA as relevant targets for miR-93 in the context of UVB -induced senescence is consistent with the available data. [score:13]
Moreover, miR-93 increases survival in cisplatin-resistant ovarian cancer cells, by directly targeting PTEN and upregulation of the AKT signaling pathway [31]. [score:7]
miR-93 is known to inhibit angiogenesis by suppressing VEGF release [29], and contributes to silencing of p21 [WAF1] gene expression after DNA damage [30]. [score:7]
To further validate the bioinformatics -based target selection, regulation of several newly identified candidate genes by specific microRNAs was addressed in HDF overexpressing miR-15a, miR-20a, and miR-93, respectively. [score:6]
However, miR-93 was also shown to i) promote tumor growth and angiogenesis by targeting integrin-β8 [42], ii) to contribute to silencing of p21 [WAF1] gene expression after DNA damage [30], and iii) to promote cell proliferation and clonogenicity of HepG2 Cells [43], suggesting that effects of miR-93 depend on cellular context. [score:5]
Expression levels for miR-15a, miR-20a, miR-20b, miR-93, and miR-101 are shown in Figure  3, along with their established target mRNAs. [score:5]
Although frequently overexpressed in human malignancies, miR-93 may actually function as a tumor suppressor gene. [score:5]
Accordingly, miR-93 abrogated VEGF protein secretion, suggesting that miR-93 interferes with angiogenesis [29], blocked tumor development in mammary fat pads [40], and suppressed proliferation of human colon cancer stem cells [41]. [score:4]
Thereby, eight miRNAs (miR-15a, miR-17, miR-20a, miR-20b, miR-34, miR-93, miR-101, miR-155) were identified for which regulated mRNA targets were found with high confidence. [score:4]
To achieve the overexpression of microRNA in HDFs, cells were reverse transfected with Pre-miR™ miRNA Precursor for miR-15a, miR-20a, miR-93, miR-101, and Pre-miR™ miRNA Precursor Molecules-Negative Control #2 for negative control (Applied Biosystems, Austria) using siPORT™ NeoFX™ Transfection Agent (Ambion, Austria) according the manufacturer’s protocol. [score:3]
of the bioinformatic analysis suggested DRAM, PIK3IP1, DKK2, Serpin G1, ADAMTS5, TIMP3, BTG2, RUNX1 and EGR2 as potential targets for miR-93 (Figure  5A). [score:3]
miRNA expression levels for miR-20a, miR-20b, miR-15a, and miR-93 were determined by. [score:3]
Together the results obtained in this study suggest important roles for microRNAs miR-15, miR-20a/b, miR-93 and miR-101, and their mRNA targets, during UVB -induced senescence of human diploid fibroblasts. [score:3]
Bioinformatic analysis of miRNA-mRNA networks was performed to identify new functional mRNA targets with high confidence for miR-15a, miR-20a, miR-20b, miR-93, and miR-101. [score:3]
Figure 5 Correlation networks of miR-93/miR-15 and their high confidence target genes. [score:3]
In these experiments, data obtained by the miRNA array for miR-15a, miR-20a, miR-20b, miR-93, and miR-101 were confirmed (Figure  2); whereas miR-17, miR-34 and miR-155 were also regulated in UVB -treated cells in accordance with the miRNA array results, the observed differences did not reach statistical significance (data not shown). [score:2]
In addition, 7 other microRNAs (hsa-miR-155, hsa-miR-15a, hsa-miR-17, hsa-miR-20a, hsa-miR-20b, hsa-miR-34a, hsa-miR-93) were chosen for qPCR confirmation of array data using the Taqman qPCR platform (Life Technologies). [score:1]
of the analysis are presented here for miR-93 (A), and miR-15 (B). [score:1]
Our analysis in UVB -induced senescence confirmed a high confidence interaction of miR-93 with CDKN1A and vEGF-A (Figure  5A), thereby validating the analytical procedure. [score:1]
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21
[+] score: 61
In other studies, miR-93 expression was found to promote tumor angiogenesis and metastasis by suppressing large tumor suppressor homology 2 expression [21]– [22]. [score:9]
Levels of miR-29c and miR-93 expression were upregulated in NSCLC tissues, while serum levels of miR-29c were also upregulated, but levels of serum miR-429 were decreased in NSCLC. [score:9]
Our data showed that levels of miR-29c and miR-93 expression were upregulated in NSCLC tissues compared to the corresponding noncancerous lung tissues. [score:5]
We found that increased miR-93 expression was strongly associated with NSCLC histology (P = 0.031, Table 2), whereas serum miR-29c expression was associated with abnormal CEA levels (P = 0.030, Table 2). [score:5]
Our data showed that levels of miR-29c and miR-93 expression were upregulated in NSCLC tissues compared to the corresponding noncancerous lung tissues (P = 0.0408 and P = 0.0444, respectively), whereas miR-429 levels were not significantly different between NSCLC and noncancerous lung tissues (P = 0.3903, Figure 1A). [score:5]
The serum level of miR-429 expression was significantly correlated with that in NSCLC tissues (r = 0.3578, P = 0.0024, Figure 1E), whereas serum levels of miR-29c and miR-93 expression were not associated with those in NSCLC tissues (r = −0.07877, P = 0.5169 and r = 0.1515, P = 0.2105, respectively, Figure 1C and D). [score:5]
We found that miR-29c and miR-93 expression was upregulated in NSCLC tissues compared to the corresponding noncancerous lung tissues. [score:5]
miRNA array was used to profile differentially expressed miRNAs and Taqman -based quantitative RT-PCR assays were used to analyze levels of miR-29c, miR-93, and miR-429 expression in NSCLC tissue samples, corresponding normal tissue samples, and serum samples from 70 NSCLC patients as well as in serum samples from 48 healthy controls. [score:4]
However, expression of miR-29c, miR-93, and miR-429 in NSCLC tissues and serum levels of miR-29c and miR-93 were not associated with the overall survival of NSCLC patients. [score:3]
Pearson correlation test showed that miR-429 expression in serum was significantly associated with that in NSCLC tissues (r = 0.3578, P = 0.0024), whereas serum levels of miR-29c and miR-93 were not associated with those in NSCLC tissues (r = −0.07877, P = 0.5169 and r = 0.1515, P = 0.2105, respectively). [score:3]
We found that serum miR-93 expression did not differ between NSCLC patients and healthy controls (P = 0.3530). [score:3]
Thus, in this study, we analyzed levels of three miRNAs (miR-29c, miR-93, and miR-429) in non-small cell lung cancer (NSCLC) tissues and compared them to those in serum samples of NSCLC patients and healthy controls, particularly, their expression levels in early stage NSCLC patients. [score:2]
P-values for miR-29c, miR-93, and miR-429 were 0.0408, 0.0444, and 0.3903, respectively, using a paired sample t-test. [score:1]
P-values of serum miR-29c, miR-93, and miR-429 were 0.0012, 0.9291, and 0.0001, respectively, using an unpaired sample t-test. [score:1]
Based on our miRNA array (Agilent) and validation data, we selected three miRNAs (miR-29c, miR-93, and miR-429) for further study in NSCLC samples. [score:1]
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22
[+] score: 51
Overexpressed miR-93 directly inhibits glucose transporter isoform 4 (GLUT4) expression, influencing glucose metabolism. [score:8]
In addition to our findings that miR-223 and miR-93 regulate IR in AT by targeting GLUT4, these two miRNAs have also been found to suppress proinflammatory activation of macrophages by targeting IRAK4 (for miR-93) [24] and Pknox1 (for miR-223) [25]. [score:8]
Both miRNAs act as antiangiogenesis regulators, but miR-93 directly targets vascular endothelial growth factor A (VEGF-A) [39], while miR-223 targets β1 Integrin [40]. [score:7]
As we previously noted that PCOS women with IR had the lowest expression of GLUT4 [11], it is possible that miR-93 and miR-223 may have additive effects on the regulation of GLUT4 expression. [score:6]
We previously reported that miR-93 is upregulated in adipose tissue (AT) from PCOS and non-PCOS women who display IR [11]. [score:4]
Macrophage activation is associated with IR [26]; therefore, these data suggest that miR-223 and miR-93 could also regulate IR by regulating inflammation. [score:3]
We previously reported that the expression of miR-93 was significantly increased in the subcutaneous abdominal AT of all PCOS patients studied and non-PCOS women with IR [11]. [score:3]
Together, these data suggest that miR-93 expression is associated with both IR and PCOS, whereas miR-223 is not involved in PCOS per se but is related to IR. [score:3]
miR-223 and miR-93 have been found to have similar functions yet they may or may not target the same genes. [score:3]
However, to promote cancer activity, they also target different genes including C/EBP β, FOXO1, NFI-A, STAT5A, ARTN, FBXW7, and SEPT6 (for miR-223) [29– 33] and FUS1, RhoC, PTEN, CDKN1A, TGF βR2, and NRF2 (for miR-93) [34– 38]. [score:3]
Both miR-93 and miR-223 regulated GLUT4 protein content in adipocytes [11]. [score:2]
These data suggest that miR-223 and miR-93 may also have additive effects on these functions. [score:1]
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[+] score: 44
In multiple human cancers, PTEN expressions are downregulated by miRNAs, which are shown in Table 1. Table 1 miRNA Locus Expression status Tumor type Reference MiR-21 17q23.1 Upregulated Colorectal, bladder, and hepatocellular cancer[112– 114] MiR-19a 13q31.3 Upregulated Lymphoma and CLL[87, 115] MiR-19b Xq26.2 Upregulated Lymphoma[87] MiR-22 17p13.3 Upregulated Prostate cancer and CLL[116, 117] MiR-32 9q31.3 Upregulated Hepatocellular carcinoma[118] MiR-93 7q22.1 Upregulated Hepatocellular carcinoma[119] MiR-494 14q32.31 Upregulated Cervical cancer[120] MiR-130b 22q11.21 Upregulated Esophageal carcinoma[121] MiR-135b 1q32.1 Upregulated Colorectal cancer[122] MiR-214 1q24.3 Upregulated Ovarian cancer[123] MiR-26a3p22.2 (MIR26A1)12q14.1(MIR26A2) Upregulated Prostate cancer[113] MiR-23b 9q22.32 Upregulated Prostate cancer[114] Abbreviations: CLL, chronic lymphocytic leukemia. [score:44]
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24
[+] score: 41
Therefore, we speculate here that downregulation of miR-93 and miR-130b can result in elevated expression of tumor suppressor TP53INP1, which in turn can cause growth arrest of AR42J-B13 cells leading to transdifferentiation. [score:8]
Using these hepatocyte and non-hepatocyte cell lines and primary tissues, we performed unsupervised clustering analysis by selecting 7 down-regulated miRNAs (miR-17-5p, miR-18a, miR-93, miR-106a, miR-106b, miR-130b and miR-375) and 4 up-regulated miRNAs (miR-21, miR-22, miR-122a and miR-182). [score:7]
Both up-regulated miRNAs (miR-21, miR-22, miR-122a and miR-182) and down-regulated miRNAs (miR-17-5p, miR-18a, miR-93, miR-106a, miR-106b, miR-130b and miR-375) were chosen as a parameter for comparison. [score:7]
Total RNA extracted from Dex/OSM treated AR42J-B13 cells (7 Days) and mock controls were used for Northern blot analysis using antisense probes against down-regulated miRNAs (miR-93, miR-106b and miR-130b) and up-regulated miRNAs (miR-21, miR-22, miR-122a and miR-182). [score:7]
Interestingly, Yeung et al. reported that miR-93 and miR-130b have a potential to target tumor suppressor protein p53 -induced nuclear protein 1 (TP53INP1) in HTLV-1 infected/transformed cells [39]. [score:5]
Mature miRNA of miR-93, miR-106b, miR-130b, miR-21, miR-22 and miR-182 were differentially expressed after transdifferentiation. [score:3]
The pattern of coordinated reduction in expression of miR-25, miR-93, and miR-106b (Table 1) is due to the clustering of these three miRNA genes at intron 12 of Mcm7 on chromosome 12. [score:3]
As shown in Table 1, both miR-93 and miR-130b were reduced in transdifferentiated hepatocytes. [score:1]
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25
[+] score: 36
Other miRNAs from this paper: hsa-mir-25, hsa-mir-106b
In order to verify the alteration in expression level of this cluster, we used quantitative real-time PCR (qRT-PCR) to detect these three miRNAs in total RNA isolated from cultured ECC-1 and HEC-1A cells with or without TSA (100 ng/mL) for 24 h. The expressions of miR-106b, miR-93, and miR-25 were shown to be downregulated significantly in cells treated with TSA compared to control (Fig. 1D & C), consistent with our microarray results (Fig. 1C). [score:7]
The expression of miR-106b, miR-93, miR-25 and their host gene MCM7 were upregulated in EMC tissues compared to the normal adjacent tissues. [score:5]
This result suggested that TSA may regulate the expressions of miR-106b, miR-93, and miR-25. [score:4]
The miR-106b-93-25 cluster consists of three miRNAs, miR-106b, miR-93 and miR-25, and is located in the 13th intron of the minichromosome maintenance protein 7 (MCM7) gene of human chromosome 7. This miRNA cluster is upregulated in many human cancers, such as gastric, prostate, and pancreatic neuroendocrine tumors, The miRNAs of the miR-106b-93-25 cluster are co-transcribed in the context of the MCM7 primary transcript. [score:4]
miR-106b and miR-93 gain of function led to an increase in cells in S-phase, while the G1-phase population was increased in the cells transfected with their inhibitors. [score:3]
miR-106b, miR-93, miR-25 promoted the growth of ECC-1 cells, while their inhibitors significantly decreased proliferation in ECC-1 cells. [score:3]
The miR-106b, miR-93 and miR-25 duplexes promoted cell proliferation compared with a control duplex, whereas their inhibitors significantly decreased proliferation of ECC-1 cells (Fig. 2A). [score:2]
We found the expressions of miR-106b, miR-93 and miR-25 were decreased in the group treated with TAM and TSA, while they were unchanged in the group treated with TAM only, compared with the control (Fig. 1C). [score:2]
We also constructed plasmids containing the p21-3′UTR with mutated seed regions for the predicted miR-106b/miR-93 binding sites (p21-mut-3′UTR), along with plasmids containing the BIM-3′UTR with mutated seed regions for the predicted miR-25 binding sites (BIM-mut-3′UTR). [score:1]
Cell numbers in G1 phase were reproducibly increased by anti- miR-106b and anti- miR-93 but were unchanged by anti- miR-25 (Fig. 2B). [score:1]
The proportion of cells transfected with miR-106b, or miR-93 in the G1 phase fell, while that of cells transfected with miR-25 remained unchanged (Fig. 2B). [score:1]
miR-106b and miR-93 had no effect on the apoptosis levels of the cells. [score:1]
However, miR-106b and miR-93 had no appreciable apoptotic effects on the cells (Fig. 2C), suggesting that these miRNAs from the cluster had more subtle effects. [score:1]
The miR-106b-93-25 cluster is composed of the highly conserved miR-106b, miR-93, and miR-25 that have been shown to accumulate in different types of cancer, including gastric cancer [37], prostate cancer [38], and esophageal adenocarcinoma [39], hepatocellular carcinoma [40], and multiple myeloma [41]. [score:1]
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26
[+] score: 34
To test the effect of alternative EC on target miRNA detection, the expression of miRNA targets (miR-181a, miR-652 and miR-93) with previously documented expression in the circulation of breast cancer patients were measured using the ECs. [score:7]
The relative expression of three target miRNAs (miR-93, miR-181a and miR-652) is presented following normalization using each of four distinct EC strategies: U6 alone, miR-16 alone, miR-425 alone and finally miR-16 and miR-425 in combination. [score:5]
This study advocated the use of miR-16 and miR-93, the most stably expressed candidate ECs, for normalization of miRNA expression in serum for gastric cancer. [score:5]
However, when U6 was applied as the normalizer miR-93 appeared to be upregulated in the cancer group. [score:4]
MiR-93, which was previously shown not to be dysregulated in breast cancer (McDermott et al., unpublished data) was overexpressed in the cancer group when U6 (p = 0.017) was used as an EC but was unaltered with other candidate ECs. [score:3]
However, when the other EC candidates are used for normalization there is no difference in miR-93 expression between the cancer and control group. [score:3]
Target miRNA (miR-93, miR-181a and miR-652) expression levels were estimated using qBase software (Biogazelle, Belgium) to calculate amplification efficiency-corrected relative quantities following normalization to each candidate EC. [score:3]
A. MiR-93 expression. [score:2]
MiR-93 has been shown to be stably expressed in blood of women with breast cancer compared to healthy controls. [score:1]
Song et al focused on gastric cancer, examining 6 miRNAs (let-7a, miR-16, miR-93, miR-103, miR-192, and miR-451) and one small nucleolar RNA, RNU6B for suitability as candidate ECs [29]. [score:1]
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27
[+] score: 34
Other miRNAs from this paper: hsa-mir-21, hsa-mir-25, hsa-mir-31, hsa-mir-155, hsa-mir-106b
Over-expressed miR-106b, miR-93, and miR-25 inhibit the synthesis of p21 [CIP1] and Bim (TGF-β downstream effectors) and therefore prevent cell cycle inhibition and apoptosis Mutations in TβRII that lead to insensivity of cell lines to TGF-β mediated growth inhibition have been previously described [194]. [score:10]
Over-expressed miR-106b, miR-93, and miR-25 inhibit the synthesis of p21 [CIP1] and Bim (TGF-β downstream effectors) and therefore prevent cell cycle inhibition and apoptosisMutations in TβRII that lead to insensivity of cell lines to TGF-β mediated growth inhibition have been previously described [194]. [score:10]
In turn, miR-106b and miR-93 regulate E2F1 expression, establishing a miRNA negative feedback loop. [score:4]
The relationship between TGF-β resistance and up-regulated level of miR-106b-25 cluster (miR-106b, miR-93, and miR-25) has been recently elucidated [193]. [score:4]
Conversely, miR-106b and miR-93 control E2F1 expression thus establishing negative feedback that prevents E2F1 self-activation. [score:3]
Over -expression of miR-106b, miR-93 and miR-25 decreases response of gastric cancer cells to TGF-β since they interfere with synthesis of TGF-β downstream effectors that promote cell cycle arrest and apoptosis, such as p21 [CIP1] and BIM, respectively [193] (Figure 5). [score:3]
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[+] score: 32
One hundred seven human placenta samples were analyzed for the expression of candidate miRNA previously shown to be expressed in the placenta and involved in regulating cell growth and developmental processes by targeting genes in a variety of cell growth and cell functioning pathways, specifically, miR-16, miR-21, miR-93, miR-135b, miR-146a, and miR-182. [score:9]
Since miRNA have been described as playing important roles in development and are susceptible to the environment, we sought to further characterize the expression of six candidate miRNA previously shown to be expressed in the placenta and previously reported to target genes in pathways crucial for regulating key cell processes – miR-16 [9], [14], miR-2 1 [9], [15], miR-93 [12], [13], miR-135b [11], miR-146a [9], [16], and miR-182 [10] – in a large series of human placentas for associations with fetal growth. [score:7]
We analyzed 107 primary, term, human placentas for expression of 6 miRNA reported to be expressed in the placenta and to regulate cell growth and development pathways: miR-16, miR-21, miR-93, miR-135b, miR-146a, and miR-182. [score:7]
Median (amol) Range (amol) miR-16 18.64 3.53–399.50 miR-21 54.86 5.25–434.74 miR-93 4.26 0.14–118.39 miR-135b 4.72 0.13–216.92 miR-146a 0.1 0.002–3.95 miR-182 0.23 0.001–3.63 Observing that expression in the lowest quartiles of miR-16 and miR-21 was associated with reduced birthweight percentile, we more specifically examined the association between low expression (≤median vs. [score:5]
Expression of miR-16, miR-21, miR-93, miR-135b, miR-146a, and miR-182 determined through qRT-PCR in 107 primary human term placenta samples. [score:3]
0021210.g001 Figure 1(A) miR-16 (p = 0.04), (B) miR-21 (p = 0.02), (C) miR-93 (p = 0.88), (D) miR-135b (p = 0.84), (E) miR-146a (p = 0.46), and (F) miR-182 (p = 0.55). [score:1]
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[+] score: 27
Previous study reported that miR-17-92 cluster has been reported to be upregulated during the clonal expansion stage of adipocyte differentiation, posi­tively regulating adipogenesis by targeting the tumor suppressor RB2/p130 [27] and knockdown of miR-106b and miR-93 significantly induced the expression of brown fat-specific genes and promoted the accumulation of lipid-droplet in differentiating brown adipocytes. [score:12]
miR-93 is a highly expressed PCOS patients' adipocyte tissue and downregulates GLUT4 gene expression, suggesting that it plays a role in the IR of PCOS [15]. [score:8]
In addition, ectopic expression of miR-106b and miR-93 suppressed the mRNA level of Ucp1 [28]. [score:5]
Additionally, miR-93 and miR-720 were lowly expressed with 5-fold in mature adipocytes compared to SVCs/hMSCs-Ad. [score:2]
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[+] score: 27
Overexpression of miR-93 could inhibit the expression of p21 by direct binding to promote cell proliferation and G [1]/S transformation and thereby accelerate tumor formation [34]. [score:8]
As previously mentioned, high expression of miR-93 leads to resistance to bromocriptine treatment by inhibiting the expression of p21 in patients with PRL adenomas, thereby stimulating tumor growth. [score:7]
MiR-128a, miR-155, and miR-516-3p are highly expressed in NFPAs, whereas miR-155 and miR-93 are highly expressed in GH adenomas. [score:5]
This indicates that the overexpression of miR-93 is positively correlated with resistance to bromocriptine. [score:3]
In PRL adenomas, expression of miR-93 is higher in patients with bromocriptine resistance than in patients who are sensitive to bromocriptine. [score:3]
Silencing of miR-93 strongly increases the drug sensitivity of MMQ cells. [score:1]
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[+] score: 26
Additional inhibitors of Wnt ligands, or β-catenin -mediated function, include miR-29b, miR-29c, and miR-93, which target BCL9L [a β-catenin co-activator and miR-29b target (Subramanian et al., 2014; Zhang et al., 2014b; Tang et al., 2015)], GNA13 and PTP4A [miR-29c targets that negatively regulate GSK3β (Zhang et al., 2014b), a kinase that phosphorylates β-catenin and triggers degradation] and SMAD7 [a miR-93 target that promotes nuclear accumulation of β-catenin (Subramanian et al., 2014; Zhang et al., 2014b; Tang et al., 2015)]. [score:12]
The miRNA miR-93 can stimulate the TGF-β pathway by repressing the inhibitory SMAD7, although the effect of miR-93 is inhibitory of Wnt signaling through inhibition of SMAD7, which can augment nuclear accumulation of β-catenin. [score:7]
MicroRNA-93 suppress colorectal cancer development via Wnt/beta-catenin pathway downregulating. [score:6]
The human miR-17 family consists of eight miRNAs (miR-17, miR-18a/b, miR-20a/b, miR-93, and miR-106a/b), with three of these (miR-17, miR-18a, and miR-20a) transcribed from the miR-17-92 miRNA locus. [score:1]
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[+] score: 25
In addition, overexpression of miR-93 increases expression of type II collagen by targeting MMP3 and might thereby promote cartilage repair [38]. [score:7]
The mRNA expression of HLA-DRB1 was positively correlated with cartilage repair in vivo, and the mRNA expression of TMEM155 and expressions of miR-486-3p, miR-148b, miR-93, and miR-320b were negatively correlated with cartilage repair. [score:7]
On the other hand, the expression of HLA-DRB1 was higher and that of TMEM155, miR-486-3p, miR-148b, miR-93, and miR-320b lower in hMSCs from the three donors that showed good cartilage repair in vivo than in hMSCs from the other two donors (Table 6). [score:3]
On the other hand, the expression levels of HLA-DRB1, TMEM155, miR-486-3p, miR-148b, miR-93, and miR-320b might be used for the quality assessment of MSCs based on the efficacy of cartilage repair. [score:3]
On the other hand, we found expression of HLA-DRB1, TMEM155, miR-486-3p, miR-148b, miR-93, and miR-320B in hMSCs to be related to the capacity for cartilage regeneration. [score:3]
Jing and Jiang reported that miR-93 is lower in human degenerative nucleus pulposus tissues and that its level is associated with disc degeneration grade. [score:1]
Of 768 miRNAs analyzed by TLDA, we detected miR-486-3p, miR-148b, miR-93, and miR-320B in cartilage repair in vivo (Table 4). [score:1]
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[+] score: 24
Figure 3miRNAs (miR-17, miR-18a, miR-20a, miR-93) negatively regulate RUNX1 targets, including miR-223 and miR-222/221, in blocking myeloid differentiation by increasing KIT expression and enabling KIT -mediated proliferation Both gain- and loss-of-function in vivo studies of miR-126 in mouse mo dels demonstrated that either enforced expression or knockout of miR-126 substantially promoted development of t(8;21) AML in mice [75]. [score:10]
miRNAs (miR-17, miR-18a, miR-20a, miR-93) negatively regulate RUNX1 targets, including miR-223 and miR-222/221, in blocking myeloid differentiation by increasing KIT expression and enabling KIT -mediated proliferation. [score:6]
miR-18a, miR-20a and miR-93 are frequently upregulated in distinct subtypes of non-CBF-AML [74]. [score:4]
Fischer et al identified that miR-17, miR-18a, miR-20a and miR-93 all function as the CBF-AML fusion proteins in negative regulating their target RUNX1 and the RUNX1-miR-221-KIT axis [74]. [score:4]
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hy926 cells Not shown HIF1A Angiogenesis[123] miR-200b HMVECs Downregulated upregulated ETS1 Angiogenesis hypoxia[135, 136] HUVECs KLF2 miR-210 HUVECs Upregulated EFNA3 Angiogenesis hypoxia[131, 132, 150] HIF3A miR-424 HUVECs, hMVECs, hBOECs and hMBECs Upregulated CUL2 Angiogenesis[137] HIF1A* miR-429 HUVECs Upregulated HIF1A Hypoxia[12, 109] HIF3A miR-433 HUVECs Downregulated HIF1A Proliferation and migration[138] miRNAs proven to directly bind HIF mRNAs are in bold, and indirect effects are marked with “*” ARNT aryl hydrocarbon receptor nuclear translocator, CUL2 cullin-2, EFNA3 ephrin A3, EGLN1 prolyl hydroxylase domain-containing protein 2 (PHD2), ETS1 ETS Proto-Oncogene 1, Transcription Factor, HIF1A hypoxia-inducible factor 1-alpha, HIF1AN hypoxia-inducible factor 1-alpha inhibitor, HIF3A hypoxia-inducible factor 3 alpha, KLF2 Kruppel-like factor 2 The miR-17 family includes miR-17, miR-18a/b miR-20a/b, miR-93 and miR-106a/b. [score:23]
With the exception of miR-93, these miRNAs are produced from several miRNA gene clusters, which apparently arose from a series of ancient evolutionary genetic duplication events, and also include members of the miR-19 and miR-25 families [111]. [score:1]
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[+] score: 24
Other miRNAs from this paper: hsa-mir-572
By doing so, we obtained the p21 response after genotoxic stress in four scenarios: (1) endogenous miRNA expression (Control); (2) overexpression of miR-572; (3) overexpression of miR-93; and (4) both miRNAs moderately overexpressed. [score:9]
The individual overexpression of miR-572 or miR-93 led to the reduction of the upregulation of p21 response after genotoxic stress induction. [score:6]
In Figure 3(a), the network module contains an incoherent FFL composed by AF2 α, miR-93 and p21, and a cascaded regulation in which p21 is repressed by FOXF2 via miR-572; in Figure 3(b), the same cascaded regulation together with a coherent FFL composed of TP53, MYC, miR-93 and p21 forms another regulatory module of p21. [score:4]
For the network module of an incoherent FFL plus the cascaded regulation (Figure 3(c), bottom left), the saturation pattern appears in two distinct regions: one with weakened transcriptional strength of miR-93 and the other with enhanced transcriptional strength of miR-93; for the other module (Figure 3(c), bottom right), the saturation pattern only appears in the region, in which the transcriptional strength of miR-93 is enhanced. [score:2]
To experimentally validate the capability of our mo del to predict the relative p21 concentrations regulated by cooperative miRNAs, we selected miR-572 and miR-93 as a case study. [score:2]
To do so, two network modules including both miR-93 and miR-572, and their TFs were exemplified. [score:1]
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[+] score: 23
Here, we took a closer look at members of the tumorigenic miR-17~92 cluster, since (i) miR-17–5p was among the top ten hits of abundant miRNAs of which the expression was maintained by the E6/E7 oncogenes, (ii) all other members of this cluster, as well as of the paralog cluster miR-106b~25, were also downregulated by E6/E7 silencing when applying less stringent selection criteria (S3 Table), (iii) several members of the miR-17~92 cluster are well-decumented to be overexpressed in cervical cancer tissues, including the tested miR-17–5p [27– 30, 34, 37] and miR-20a-5p [23, 30, 31, 34, 35] (also see S2 Dataset), and (iv) four of these miRNAs (miR-17–5p, miR-20a-5p, miR-93–5p, and miR-106b-5p) possess the same seed sequence and can bind to the 3’ UTR of the p21 mRNA [18]. [score:8]
HPV E6/E7 increase intracellular levels of members of the oncogenic miR-17~92 cluster that reduce p21 expression in HPV -positive cancer cellsThe 52 most abundant cellular miRNAs that were downregulated > 1.5-fold upon E6/E7 silencing in both deep sequencing and qRT-PCR analyses encompassed miR-17–5p and miR-19b-3p, two members of the miR-17~92 cluster, and miR-93–5p, a member of the paralog miR-106b~25 cluster (Fig. 2D/E). [score:6]
The 52 most abundant cellular miRNAs that were downregulated > 1.5-fold upon E6/E7 silencing in both deep sequencing and qRT-PCR analyses encompassed miR-17–5p and miR-19b-3p, two members of the miR-17~92 cluster, and miR-93–5p, a member of the paralog miR-106b~25 cluster (Fig. 2D/E). [score:4]
Four miRNAs encoded by the miR-17~92 and miR-106b~25 clusters (miR-17–5p, miR-20a-5p, miR-106b-5p, miR-93–5p) are grouped into the miR-17 family, according to their identical seed sequence, and target two binding sites in the 3’ UTR of p21 [66, 67]. [score:3]
The oncogenicity of miRNAs has been particularly well demonstrated for members of the miR-17~92 cluster (also called “oncomir-1”; coding for miR-17, miR-20a, miR-18a, miR-19a, miR-19b and miR-92a) and of its paralog cluster miR-106b~25 (coding for miR-106b, miR-93 and miR-25) [18]. [score:1]
The 23 miRNAs comprise several family members of the miR-378 family (miR-378a-3p, miR-378c, miR-378d, miR-378f), as well as members of the miR-17~92 and miR-106b~25 clusters (miR-17–5p, miR-19b-3p, miR-93–5p). [score:1]
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Hsa-miR-93 and -31 were also upregulated in liver metastasis, but not in lymph nodes positive for metastasis, whereas hsa-miR-21 over -expression was evident in both, liver and lymph nodes metastasis. [score:6]
ISH of hsa-miR-93 confirmed its upregulation in adenocarcinomas and liver metastasis, showing a significant difference between normals and either adenocarcinomas (P-value  = 0.0345) or liver metastasis (P-value  = 0.007) and between adenocarcinomas and liver metastasis (P-value  = 0.043). [score:4]
Induced over -expression of hsa-miR-93 was shown: to be tumorigenic in vitro and in vivo, to promote survival but not proliferation, and to enhance neoangiogenesis [87]. [score:3]
Thus, hsa-miR-93 expression pattern could differentiate normal colon from primary adenocarcinoma, and normal colon and primary adenocarcinoma from liver metastasis, but not from secondary tumoral lymphnodal invasion. [score:3]
0096670.g001 Figure 1Normal colon tissue (NORM), colon adenocarcinomas (ADENOCA), lymphonodal metastasis (POSLYM) and colon liver (LIVERMET) for miR-21(A), miR-31 (B), miR-93 (C), miR-103 (D) and miR-566 (E). [score:1]
The ISH data for hsa-miR-93 completely overlapped the qRT-PCR results: an intense staining was detected in primary colonic adenocarcinomas and liver metastasis. [score:1]
Moreover, increased hsa-miR-93 levels, correlate to progression and decreased patients survival in serous ovarian carcinoma [88] and, can predict relapse breast cancer [89]. [score:1]
miR-93 In Situ Hybridization of Normal Colon (5X), Primary Tumor (5X) and lymph node metastasis (5X). [score:1]
with normal colon tissue (NORM), colon adenocarcinomas (ADENOCA), lymphonodal metastasis (POSLYM) and colon liver (LIVERMET) for miR-21 (A), miR-103(B), miR-93(C), miR-566(D) and miR-200c(E). [score:1]
Normal colon tissue (NORM), colon adenocarcinomas (ADENOCA), lymphonodal metastasis (POSLYM) and colon liver (LIVERMET) for miR-21(A), miR-31 (B), miR-93 (C), miR-103 (D) and miR-566 (E). [score:1]
0096670.g002 Figure 2with normal colon tissue (NORM), colon adenocarcinomas (ADENOCA), lymphonodal metastasis (POSLYM) and colon liver (LIVERMET) for miR-21 (A), miR-103(B), miR-93(C), miR-566(D) and miR-200c(E). [score:1]
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[+] score: 22
Seven miRNAs (downregulated: miR-29c, miR-93, miR-101 and miR-130a; upregulated: miR-9, miR-182 and miR-221) were identified as differentially expressed (≥2-fold) in both A172-TR and U251-TR cell lines (Figure 1A). [score:9]
Upregulated miRNAs (miR-9, miR-182 and miR-221) were shown in red, downregulated miRNAs (miR-29c, miR-93, miR-101 and miR-130a) were shown in green. [score:7]
Among them, three miRNAs (miR-29c, miR-93 and miR-101) were downregulated and two (miR-9 and miR-182) were upregulated in the TMZ-refractory samples as compared with that in the primary tumor samples (p<0.05, Figure 1D). [score:6]
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Leukemia inhibitory factor (LIF), which is known to be a major regulator of trophoblast functions, has been reported to upregulate miR-21 and miR-93, but down-regulate miR-141, expression in JEG-3 cells [109]. [score:12]
Further analysis using real-time PCR confirms that miR-93, -205, -224, -335, -451, and -491 are upregulated while miR-424 is down-regulated when cells were cultured under 1% oxygen for 48 h [57]. [score:7]
Similarly, miR-93 promotes tumor angiogenesis in U87 cells by targeting integrin-β8 [141]. [score:3]
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[+] score: 22
We used miScript Primer Assays for 9 miRNAs (miR-744-5p, miR-648, miR-193b-3p, miR-212-3p, miR-143-3p, miR-93-5p, miR-222-3p, miR-423-3p and miR-766-3p) and QuantiTect Primer assays for 9 target genes (CDKN1A, MYC, PTEN, ESR1, ETS1, SOD2, MGMT, KRAS and HNF4A) (Qiagen, Hilden, Germany) to validate the different expression levels of the miRNA and their target genes, which are determined by miRTargetLink prediction software. [score:5]
Out of the eight miRNAs, miR-648 was up-regulated and the seven remaining miRNAs were down regulated (miR-744-5p, miR-193b-3p, miR-212-3p, miR-143-3p, miR-93-5p, miR-423-3p and miR-766-3p). [score:5]
In addition, we selected six miRNAs (miR-328-5p, miR-210-5p, miR-423-3p, miR-143-3p, miR-874-5p and miR-93-5p) with low or moderate expression levels based on the array analysis and with known association with cardiac pathologies and its related process [26– 33]. [score:3]
The blue and orange lines indicate the two main clusters of samples To validate the data obtained from the miRNA microarray, RT-qPCR was performed to re-examine the expression level of nine miRNAs, namely miR-328-5p, miR-4750-5p, miR-210-5p, miR-423-3p, miR-143-3p, miR-564, miR-770-5p, miR-874-5p and miR-93-5p. [score:3]
The significance of the differences in the expression was confirmed for seven miRNAs namely miR-210-5p, miR-423-3p, miR-143-3p, miR-564, miR-770-5p, miR-874-5p and miR-93-5p) (P < 0.05) in the atrial myocardial samples from patients with CHDs after CPB compared to before CPB by RT-qPCR (Fig.   3). [score:2]
Similarly, reduced expression of miR-93 was observed in the left atrium compared to right atrial tissue of patients with sinus rhythm [50] indicating that miR-93 promotes angiogenesis in the ischemic tissue by coordinating the functional pathways of cell proliferation and apoptosis [51]. [score:2]
By including two out of the seven validated miRNAs namely miR-143-3p and miR-93-5p, a clear distinction between the groups based on the clustering dendrogram was, however, not possible (Additional file 2: Figure S1). [score:1]
In addition, we found two miRNAs with border-line P-values (miR-423-3p, P-value = 0.060 and miR-93-5p, P-value = 0.070) (Fig.   4). [score:1]
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[+] score: 21
Other miRNAs from this paper: hsa-mir-25, hsa-mir-106b
Both miR-106b and miR-93 have also been shown to target E2F1 effectively inhibiting its translation [17]. [score:7]
Consistent with our observation that overexpression of miR-106b and miR-93 predict sensitivity to CHOP, E2F1 expression has previously been associated with poor survival of breast cancer patients treated with FEC [18]. [score:5]
The miR-106b-25 cluster (miR-106b, miR-93, and miR-25) is involved in E2F1 posttranscriptional regulation and Targets PTEN. [score:4]
miR-106b, miR-93 and miR-25 form a cluster, all expressed from the same intron. [score:3]
miR-93 is part of the miR-106b-25 cluster, where all members are part of the CHOP microRNA profile. [score:1]
That is miR-93. [score:1]
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[+] score: 21
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7e, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-20a, hsa-mir-21, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-101-1, hsa-mir-106a, hsa-mir-107, hsa-mir-192, hsa-mir-34a, hsa-mir-204, hsa-mir-205, hsa-mir-214, hsa-mir-215, hsa-mir-222, hsa-mir-223, hsa-mir-1-2, hsa-mir-15b, hsa-mir-125b-1, hsa-mir-141, hsa-mir-191, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-127, hsa-mir-149, hsa-mir-184, hsa-mir-186, hsa-mir-200c, hsa-mir-1-1, hsa-mir-200a, hsa-mir-101-2, hsa-mir-34b, hsa-mir-34c, hsa-mir-339, hsa-mir-146b, hsa-mir-548a-1, hsa-mir-548b, hsa-mir-548a-2, hsa-mir-548a-3, hsa-mir-548c, hsa-mir-624, hsa-mir-650, hsa-mir-651, hsa-mir-548d-1, hsa-mir-548d-2, hsa-mir-449b, hsa-mir-1185-2, hsa-mir-1283-1, hsa-mir-1185-1, hsa-mir-708, hsa-mir-548e, hsa-mir-548j, hsa-mir-1285-1, hsa-mir-1285-2, hsa-mir-548k, hsa-mir-548l, hsa-mir-548f-1, hsa-mir-548f-2, hsa-mir-548f-3, hsa-mir-548f-4, hsa-mir-548f-5, hsa-mir-548g, hsa-mir-548n, hsa-mir-548m, hsa-mir-548o, hsa-mir-548h-1, hsa-mir-548h-2, hsa-mir-548h-3, hsa-mir-548h-4, hsa-mir-548p, hsa-mir-548i-1, hsa-mir-548i-2, hsa-mir-548i-3, hsa-mir-548i-4, hsa-mir-1283-2, hsa-mir-548q, hsa-mir-548s, hsa-mir-548t, hsa-mir-548u, hsa-mir-548v, hsa-mir-548w, hsa-mir-548x, hsa-mir-548y, hsa-mir-548z, hsa-mir-548aa-1, hsa-mir-548aa-2, hsa-mir-548o-2, hsa-mir-548h-5, hsa-mir-548ab, hsa-mir-548ac, hsa-mir-548ad, hsa-mir-548ae-1, hsa-mir-548ae-2, hsa-mir-548ag-1, hsa-mir-548ag-2, hsa-mir-548ah, hsa-mir-548ai, hsa-mir-548aj-1, hsa-mir-548aj-2, hsa-mir-548x-2, hsa-mir-548ak, hsa-mir-548al, hsa-mir-548am, hsa-mir-548an, hsa-mir-548ao, hsa-mir-548ap, hsa-mir-548aq, hsa-mir-548ar, hsa-mir-548as, hsa-mir-548at, hsa-mir-548au, hsa-mir-548av, hsa-mir-548aw, hsa-mir-548ax, hsa-mir-548ay, hsa-mir-548az, hsa-mir-548ba, hsa-mir-548bb, hsa-mir-548bc
An overall down-regulation of the Rb pathway was apparent, with mutations in RB1 in three samples and global up-regulation of MDM2 (RB1 repressor) and of RB1 -targeting miRNAs (miR-215, miR-106a, miR-17, miR-20a, miR-93, miR-215, miR-21). [score:10]
The top five up-regulated oncogenic miRNAs were miR-93, miR-20a, miR-214, miR-146b and miR-92a (percentile rank of overexpressed miRNAs 92.9, 91.4, 84.3, 75.8 and 68, respectively). [score:6]
Tumor associated miRNAs present in this network included miR-18a, miR-93, miR-146b, miR-200c, miR-214, miR-223, miR-650 and miR-1285, collectively repressing 9 tumor suppressors genes (including ARID2, ATM, CYLD, KLF6, NBN, SDHD, SMAD4, SMARCA4, TNFAIP3), nine oncogenes (including BCL2, CREB1, CREBBP, FOXO1, MYC, MYH11, MYH9, NOTCH1), growth inhibitors (BMPR2, BTG2, BTG3, LITAF and PA2G4), chromatin remo delers, six claudin genes (CLDN11, CLDN14, CLDN5, CLDN7, CLDN8, CLDND1) 11 DNA-repair/ATM pathway genes (ATF4, CREB3, CREB5, DCLRE1C, ERCC1, PARP1, POLR2C, RAD23B, RAD51, RPA1, XRCC5) and six proapoptotic genes (BAK1, BCL2L1, MCL1, SMAD3, SMAD5, SMAD7). [score:5]
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[+] score: 19
For example, miR-106b, miR-107, miR-130a, miR-34 [9], miR-93, miR-155, miR-181a, miR-21, miR-23a, miR-320a [8], miR-193b, miR-320b [13] are significantly up-regulated and miR-148a [11, 14], miR-330-5p [15], miR-373 [16] significantly down-regulated. [score:7]
Interestingly, among the miRNAs participating in the 50 most significant miRNA-mRNA interactions we can find: miR-106b, miR-93, miR-148a, miR-330-5p that could be interacting with more than 4 different targets at the same time. [score:3]
The same occurs for miR-93 and miR-106b that belong to the same family, for miR-320a and miR-320b and for miR-19a and miR-19b (right part Figure 3A) that are clustered together, respectively, according to their miRComb targets. [score:3]
Interestingly, most of these miRNAs are coincident with those appearing in Table 1 (miR-374b, miR-148a, miR-181a, miR-373, miR-320a, miR-93, miR-106b, miR-497, miR-23a, miR-19b, miR-107, miR-15a, miR-330-5p, miR-144), indicating that, apart from being targeting many mRNAs, these miRNAs are participating in the most reliable interactions. [score:3]
It is worth noting that these 10 miRNAs together (miR-374b, miR-148a, miR-181a, miR-373, miR-320a, miR-448, miR-93, miR-106b, miR-217, miR-539) could potentially be regulating 41% of the mRNAs significantly altered in PDAC. [score:2]
These miRNA-mRNA interactions are miR-106b-LRRC55, miR-21-PDCD4, miR-148a-YWHAB, miR-93-FAM129A, miR-330-5p-GPI, miR-330-5p-BHLHE40, miR-93-LRIG1, miR-23a-LRIG1, miR-148a-ARF4, miR-106b-FAM129A, miR-148a-ACVR1, miR-148a-CTTNBP2NL. [score:1]
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[+] score: 19
Expression changes were also validated by q-RT-PCR for seven miRNAs: 5 up-regulated (hsa-miR-21, hsa-miR-29a, hsa-miR-29b, hsa-miR-221 and hsa-miR-222) and 2 down-regulated (hsa-miR-93 and hsa-miR-106a) (Figure 3B-H). [score:9]
For instance, the paralog miRNA clusters miR-106a/363 (integrated by miR-106a, miR-363, miR-92-2, miR-19b-2, miR-20 and miR-18b), miR-106b/25 (compound of miR-106b, miR-25 and miR-93) and miR-17/92 (comprising miR-17, miR-18a, miR-19a, miR-20a, miR-19b-1 and miR-92a-1) are down-regulated upon differentiation, while clusters miR-29a/29b and miR221/222 are strongly up-regulated, suggesting an important role for coordinate regulatory miRNA networks during GIC differentiation. [score:8]
Validation of the differential expression of miR-29a (B), miR-29b (C), miR-221 (D), miR-222 (E), miR-21 (F), miR-93 (G) and miR-106a (H) was carried out by q-RT-PCR using specific TaqMan microRNA assays, normalizing their expression values with respect to RNU6B levels and to the NS state by calculating 2 [-ΔΔCt]. [score:2]
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[+] score: 17
In the case of miR-342miR-93 one may presume that before sponging, this edge was not present because of the lower reaction rates between these two miRNAs and the common target compared to the significantly higher reaction rate between the miRNAs of group S with this specific target. [score:4]
ATP13A3 ATPase 13A3 N/A CDKN1A cyclin dependent kinase inhibitor 1A[33] GSK3B glycogen synthase kinase 3 beta[34] RAPH1 Ras association (RalGDS/AF-6) and pleckstrin homology domains 1 N/A SLC38A1 solute carrier family 38 member 1 N/A TET3 tet methylcytosine dioxygenase 3 N/A TP53INP1 tumor protein p53 inducible nuclear protein 1 N/A TSPAN14 tetraspanin 14 N/A Group C (miR-21, miR-23, miR-26a, miR-26b, miR-93, miR-223). [score:3]
After sponging, despite the lower reaction rates, the target becomes more available for miR-342 and miR-93, thus increasing their competition over it. [score:3]
To better illustrate our findings, we applied the principle of predator-prey like mathematical mo del to our groups of miRNAs, S = {miR-182, miR-146, miR-155, miR-16, miR-29a} and C = {miR-23, miR-26a, miR-26b, miR-93, miR-223, miR-21}. [score:1]
The C group is composed of six miRNAs–miR-23, miR-26a, miR-26b, miR-93, miR-223 and miR-21. [score:1]
New edges appear between miR-93 and miR-342 and between miR-16 and miR-486. [score:1]
In the control patient network, miR-23, miR-26a and miR-26b are strongly connected each through 11 edges; miR-93 and miR-155 are also strong connected to 10 other nodes through 10 edges. [score:1]
These nodes are represented by miR-26a, miR-26b, miR-23, miR-93 miR-146 and miR-155 for the control group. [score:1]
Also, we observed that only two new edges appear (red edges), an edge that connects miR-342 with miR-93 and an edge that connects miR-16 with miR-486. [score:1]
This was reflected by the appearance of a new edge between miR-342 and miR-93 (red line in Fig 4). [score:1]
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[+] score: 17
MiR-29 and miR-30e have been shown to influence Wnt signaling; miR-23b has been shown to inhibit FZD7 translation; miR93 has been shown to down-regulate expression of genes encoding β-catenin, Axin, Myc and Cyclin D; and miR-135a/5 have been shown to suppress APC expression [4]. [score:14]
In our data miR-29b-3p and miR-93-5p had a seed region match with CCND1, and miR-29a-3p was also associated with SFRP5 and ROCK2; while miR-30a-5p was associated with DAAM2, suggesting regulation of both the canonical and non-canonical Wnt-signaling pathways. [score:2]
While 33s were associated with only one gene, manys were associated with multiple genes: miR-7i-5p, miR-133b, miR-221-3p, miR-27a-3p, miR-29a-3p, miR-6515-5p, miR-663a, and miR-93-5p were associated with two genes; miR-145-5p, miR-17-5p, miR-193-3p, miR-19b-30, miR-203a, miR-20a-5p, miR-21-5p, miR-3651, miR-650, and miR-663b with three genes, and miR-92a-3p, miR-150-5p, and miR-20b-5p with four genes. [score:1]
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[+] score: 16
Divergent molecular patterns of microRNA expression were observed in alternate lung lobes, specifically noted was disparate expression of miR-93 and miR-4454 in alveolar macrophages along with altered expression of miR-451a and miR-663a in bronchoalveolar lavage fluid. [score:7]
The one overlap we noted between our protein data and microRNA data was the observation of the increase of miR-93-5p expression seen in lower lobe AM is consistent with our observed decreased of IL-8 secretion by AM in vitro. [score:3]
Potential targets identified via miRTarBase [21] of miR-93-5p include PTEN [26] and Il-8 [27], of miRNA-22-5p include PPARA and BMP7 [28] and HMGB1 [29], and of miR-4454 include TRAM2 and CCR4. [score:3]
However, in the lower lobe, four microRNAs were significantly altered including: miR-22-3p (P = 0.007), miR-30e-5p (P = 0.015), miR-4454 (P = 0.019), and miR-93-5p (P = 0.036). [score:1]
As a first approach, we examined the microRNA content of AM and identified a number of microRNAs that play roles related to innate immunity, inflammation or extracellular matrix remo deling including: miR-93-5p, miR-22-3p and miR-4454. [score:1]
Ultimately, it remains to be determined if the specific microRNAs identified in this study, miR-93, miR-4454, miR-451a or miR-663a, function intracellularly, in a paracrine or endocrine manner in cell-to-cell communication, or are involved in trans-kingdom crosstalk with potential pathogens, or some combination of these processes. [score:1]
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[+] score: 16
Other miRNAs from this paper: hsa-mir-17, hsa-mir-25, hsa-mir-125b-1, hsa-mir-125b-2, hsa-mir-106b
Here we would like to suggest that this intricate regulatory circuit might have another layer to it; in addition to being targeted by the miR-17/92 cluster, E2F family genes might also be targeted by miR-93 cluster members, which share similar seeds. [score:6]
Moreover, here the architecture is more complex, as it also includes a set of mutual target genes, through which E2F and the miR-93 cluster may exert their regulatory roles. [score:4]
In turn, the miR-93 cluster is transcribed from an intron of the MCM7 host gene, which is a verified target of the E2F family [46]. [score:3]
miR-93 cluster members are also homologous to two other genomic miR clusters, one of which is miR cluster 17/92 [41]. [score:1]
miR-93 is part of a cluster of three miRs, miR-106b, miR-93, and miR-25, which lie in close proximity to each other inside an intron of the MCM7 gene. [score:1]
The circuit consists of the TF E2F and miR-93. [score:1]
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49
[+] score: 16
Different groups have previously reported down and up-regulation of miRNAs in Tax -expressing cells such as miR93, 144, 146a, 155, 451 [13, 31, 32] however, it is not clear whether the observed up- or downregulation of miRNA expression simply reflects malignant weakening of the tumor or is a direct cause for initiation and tumoral progression [13, 20, 33, 34]. [score:12]
Among those upregulated miRNAs by Tax, we focused on miRNAs known to play a key role in oncogenesis and chemoresistance such as miRlet7-a, miR16, miR20, miR 21, miR31, miR93, miR125a, miR132, miR143, miR155,miR200 and miR873 [22, 23]. [score:4]
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50
[+] score: 15
We analyzed the number of validated target genes that were differentially expressed in SCZ individuals and revealed a lower frequency compared with the respective miR-137 targets (Additional file  1: Table S6, differentially expressed targets: let-7a 14%, miR-21-5p 11%, miR-93-5p 14%, miR-451a 0%, and miR-675-5p 0%). [score:10]
We selected the following five microRNAs that are expressed in the DLPFC but are not differentially expressed in SCZ individuals: let-7a, miR-21-5p, miR-93-5p, miR-451a, and miR-675-5p. [score:5]
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[+] score: 15
In our study, miR-93 was underexpressed in the serum of women who developed HER-2 -overexpressing breast tumors; interestingly, miR-93 expression was recently shown to induce a more differentiated cell phenotype in breast cancer cell lines, and expression of miR-93 in mouse mammary fat pads blocked tumor development and metastases [44]. [score:10]
Of the seven miRNAs differentially expressed in the serum of women who developed HER-2 -overexpressing breast tumors, miR-93, miR-183, and miR-29a have been reported to be associated with breast cancer in previous studies [20, 43, 44]. [score:5]
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[+] score: 15
miR-93, miR-141, miR-200 and miR-214 are frequently upregulated whereas miR-100, miR-143, miR-145 and let-7 are downregulated in ovarian carcinomas compared with normal counterparts (5– 9). [score:6]
Our miRNA hits did not always correspond to dysregulated miRNAs reported in ovarian cancer (5– 10), but included pro-proliferative miR-93 that was upregulated in primary ovarian carcinomas (6), supporting an oncogenic role of this miRNA in ovarian cancer. [score:5]
We discovered pro-proliferative miRNAs (miR-9 [*], miR-93, miR-130a, miR-130b, miR-301, miR-302b, miR-302d, miR-363, miR-372, miR-373), and anti-proliferative miRNAs (miR-7, miR-124a, miR-192, miR-193a, miR-193b, miR-199a [*], miR-432 [*], miR-497, miR-506, miR-517c) in A2780 cells. [score:1]
For example, miR-93/miR-302/miR-372/mir-373 seed family miRNAs (miR-93, miR-302b, miR-302d, miR-372, miR-373) were pro-proliferative, while miR-193 seed family miRNAs (miR-193a, miR-193b) were anti-proliferative (Table I). [score:1]
By the same miRNA mimics library screening, we found that miR-93/miR-372/miR-373 and miR-124a were pro-proliferative and anti-proliferative, respectively, in DLD-1 colorectal cancer cells (23), suggesting consistent roles of these miRNAs on cell proliferation in ovary and colorectal cancer cells. [score:1]
pro-proliferative miR-93, miR-302b, miR-302d, miR-372, miR-373 and anti-proliferative miR-193a, miR-193b), supporting the importance of the seed region of miRNA on its function. [score:1]
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[+] score: 14
The category of genes matching the keyword ‘development’ is significantly enriched in antisense (complementary) Alu-localized target sites for hsa-mir-367, -25, and 92, and at the same time shows decrease in sites localized in sense (direct) Alu insertions as hsa-mir-93, -17-5p, -20, -106. [score:5]
A little more is known about the targets for hsa-mir-93, -17-5p, -20, -106. [score:3]
Beyond their non-homogenous distribution, hsa-mir-367/-25/92 and hsa-mir-93/17-5p,/20/106 target sites occur in too many genes with quite different functions. [score:3]
Not regarding their non-homogenous distribution, hsa-mir-367/-25/92 and hsa-mir-93/17-5p,/20/106 target sites occur in too many genes with quite different functions. [score:3]
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[+] score: 14
Furthermore, we tested two more inhibitors of miRNAs expressed in HaCaT cells that are predicted to target TGFBR2: miR-93 and miR-423 (miR-20a and miR-93 have the same seed sequence). [score:7]
We therefore went beyond the siRNA screen and looked for evidence for repression of TGFBR2 by expressed miRNAs (in HaCaT cells), such as miR-20a, miR-93, miR-34a and miR-423, and non-expressed miRNAs, such as miR-373. [score:5]
The result of the miR-423 assay was not consistent with expectation, but transfection of the miR-93 inhibitor resulted in increased TGFBR2 mRNA levels relative to a LacZ siRNA control (Figure 10B). [score:2]
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[+] score: 14
Figure 4 in 9 patients with obvious coherent dysregulations of the 5 validated miRNAsExpression levels of these miRNAs (2 [−ΔCt] scale at Y-axis) were normalized by the mean Ct value of miR-30d and miR-93. [score:4]
Expression levels of these miRNAs (2 [−ΔCt] scale at Y-axis) were normalized by the mean Ct value of miR-30d and miR-93. [score:3]
Expression levels of the miRNAs (2 [−ΔCt] scale at Y-axis) were normalized by the mean Ct value of miR-30d and miR-93. [score:3]
Figure 3Expression levels of these miRNAs (2 [−ΔCt] scale at Y-axis) were normalized by the mean Ct value of miR-30d and miR-93. [score:3]
Suggested endogenous miRNA controls (miR-30d, miR-93) were selected to normalizing miRNA profiles of seminal plasma [36, 37]. [score:1]
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[+] score: 14
Interestingly, SIRT7 is a metabolic target for miR-93, a negative regulator of adipogenesis, whose expression is decreased in genetically obese ob/ob mice [55]. [score:6]
As mentioned above, SIRT -targeting miRNAs were found to be crucial for the regulation of adipogenesis and determination of MSCs differentiation towards preadipocytes (e. g., miR-34a, miR-22, miR-93, miR-146b, miR-181a) as well as lipid metabolism (miR-33, miR-34a), insulin secretion (miR-15b) and sensitivity (miR-125a), and their expression profile differs between tissues obtained from obese and normal-weight individuals [26, 41, 44, 53, 85, 86, 87, 88]. [score:6]
Cioffi M. Vallespinos-Serrano M. Trabulo S. M. Fernandez-Marcos P. J. Firment A. N. Vazquez B. N. Vieira C. R. Mulero F. Camara J. A. Cronin U. P. MiR-93 Controls Adiposity via Inhibition of Sirt7 and Tbx3 Cell Rep. [score:2]
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[+] score: 14
Taken together, these data indicate that up-regulation of miR-27a/b, -125b, -146a and down-regulation of miR-93 may contribute to the stress-related vascular and inflammatory components of AD pathology. [score:7]
In this context, prime examples of the latter include the up-regulation of miR-27a/b [99], miR-125b [98] and down-regulation of miR-93 [96], as observed in AD brain. [score:7]
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[+] score: 13
Results from microarrays (Figure 3) and qRT-PCR (Figure 7) suggested that levels of expression of all members of the miR-106b-25 cluster (miR-106b, miR-25, and miR-93) were decreased in cells transfected with miR-20b mimic or with miR-363-5p mimic (Figures 7A,B). [score:3]
Likewise, the more potent anti-proliferative miR-20b decreased expression miR-19a and miR-93 (Figures 6C, 7C) as well as increasing the level of miR-106a by about 60% (Figure 8C); changes consistent with an anti-proliferative effect (Qin et al., 2010; Fang et al., 2011). [score:3]
MicroRNA miR-93 promotes tumor growth and angiogenesis by targeting integrin-beta8. [score:3]
from microarrays (Figure 3) and qRT-PCR (Figure 7) suggested that levels of expression of all members of the miR-106b-25 cluster (miR-106b, miR-25, and miR-93) were decreased in cells transfected with miR-20b mimic or with miR-363-5p mimic (Figures 7A,B). [score:3]
The miR-106b-25 cluster, located on human chromosome 7, encodes three miRNAs: miR-106b, miR-93, and miR-25. [score:1]
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[+] score: 13
Figure 4 A. Histograms of individual miRNA QPCR analysis showing circulating levels of randomly selected miRNAs (miR-20a, miR-27b, miR-1224-3p, miR-1260, and miR-93) in the serum of animals with non-metastatic primary disease or with high-risk metastatic disease. [score:5]
Compared with the non-metastatic favorable disease animals, we observed marginal variations in the expression of miR-20a, miR-27b, miR-93, miR-1260, and miR-1224 (Figure 4A). [score:4]
For the present study, we used QPCR to confirm expression of selected miRNAs, including hsa-miR-1224-3p, hsa-miR-1260, hsa-miR-27b, hsa-miR-93, and hsa-miR-20a. [score:3]
B. Correlation analysis of the serum-circulating profiles of miR-20a, miR-27b, miR-1224-3p, miR-1260, and miR-93 observed using the miRnome approach. [score:1]
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60
[+] score: 13
This downregulation of Pur-α in monocytes was attributed to the robust expression of a group of miRNAs that included miR-15a, miR-15b, miR-16, miR-20a, miR-106b, and miR-93, which all directly targeted the 3′UTR of Pur-α [229]. [score:9]
Taking advantage of the availability of Dicer1 -deficient mice, Otsuka et al. [210] determined that cellular miRNAs miR-24 and miR-93 display antiviral activity against vesicular stomatitis virus (VSV) by targeting the viral genes encoding large protein (L protein) and phosphoprotein (P protein). [score:3]
RT-PCR analysis of a selective group of four miRNAs from this list—miR-93, miR-148b, miR-221 ad miR-16, supported the microarray data. [score:1]
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[+] score: 13
Because of the previous study of Stern-Ginossaret al. reported that miRNAs (miR-20a, miR-93 and miR-106b) could downregulate MICA and MICB expression [16], two types of reporter mutants were generated. [score:6]
The studies of Stern-Ginossa et al. reported that miR-20a, miR-93 and miR-106b could control MICA and MICB expressions [16] and recent study of Tsukerman et al. also showed that MICB expressions were regulated by miR-10b [31]. [score:6]
One type of constructs contained the mutated binding sites of both known miRNAs (miR-20a, miR-93 and miR-106b) and nine novel miRNAs (our candidate miRNAs, miR-320c, miR-320a, miR-320b, miR-320c, miR-320d, miR-542-3p, miR-641, miR-661 and miR-940) and another type contained only the mutated binding sites of known miRNAs as a positive control (Figure 3A). [score:1]
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[+] score: 12
Other miRNAs from this paper: hsa-mir-135b
healthy) SOLiD sequencing Twenty microRNAs downregulated[32•] BeWo and JAR cells miR-93 inhibitors Reduced invasion[33] Feto-placental endothelial cells derived from the third trimester Conditioned media derived from trophoblasts isolated from patients with gestational diabetes mellitus (GDM)Reduced migrationIncreased tube formation[57] JEG-3 cells hCG-H neutralising antibody reduced invasion, no effect on migration[18] JEG-3 cells shRNA MTA-3 stable knockdownhCG secretion reducedMigration increased[36] HTR-8/SV. [score:7]
Furthermore, Pan et al. have shown that miR-93 inhibitors can stimulate trophoblast migration and invasion [33]. [score:3]
Placental choriocarcinoma-derived cell lines, BeWo and JAR cells, in a transwell chamber also displayed reduced motility when transfected with miR-93 mimetics [33]. [score:1]
The functional role of miR-93 has yet to be elucidated in pre-eclampsia; however, the levels of this microRNA are increased within the plasma of patients who developed pre-eclampsia. [score:1]
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[+] score: 12
BBR inhibited miR-93 expression and increase the expression of the PTEN tumor suppressor which is a target of miR-93. [score:11]
miR-93 levels were higher in cisplatin resistant A2780 cells [285]. [score:1]
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The downregulation of ULBP2 and MICA expression by miR-519a-3p has been implicated in the inhibition of NK cell -mediated cytotoxicity of breast cancer cells (35), whereas miR-93 mimics decreased cell surface expression of MICA, MICB, and ULBP3 by translational repression, thus contributing to the immune evasion of glioma cells (36). [score:12]
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[+] score: 12
A similar auto-regulatory feedback mechanism has also been described for the regulation of miR-93 and downstream Osterix expression during osteoblast mineralization [3] as well as for miR-140 on TGF-β signaling through SMAD3 suppression [44]. [score:7]
Foetal epiphyseal cells expressed higher levels of miRs including miR-140 and miR-138 reported to promote chondrogenesis [20]– [22] while diaphyseal cells expressed miRs including miR-210 and miR-93 previously reported to promote osteogenesis [3], [23], [39]. [score:5]
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[+] score: 12
Similarly, through the c-Met/PI3K/Akt pathway, miR-93 promotes cell proliferation, migration, and invasion and also allows liver cancer cell to avoid apoptosis by directly inhibiting PTEN and CDKN1A expression [40]. [score:6]
By downregulating the antitumor genes tensin homology protein (PTEN) and cyclin -dependent kinase inhibitor 1 (CDKN1A), miR-93 can promote cell proliferation, migration, and invasion by activating the c-Met/PI3K/Akt signaling pathway; additionally, anti-miR-93 transfection makes HCC cells more sensitive to sorafenib and tivantinib treatment [40]. [score:6]
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67
[+] score: 11
miRNA-33b and miR-93 could both regulate the MYC gene and promote apoptosis in tumors, whereas circCCDDC66 could inhibit this process by inhibiting both miRNAs [62] (Fig.   3 ). [score:6]
CircCCDC66 can promote apoptosis by inhibiting miR-93, thereby promoting the expression of c-MYC. [score:5]
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[+] score: 11
The expression of miR-106a, miR-106b, miR-17-5p, miR-92, miR-93, miR-130a, miR-20a and miR-190 were much higher in EB than in either hES cells or adult cells (Figure 6, panel B). [score:3]
The expression of miR-106a, miR-106b, miR-17-5p, miR-92, miR-93, miR-190, miR-20a and miR-130 were highest in EB (panel B). [score:3]
For miR-106b, miR-92, miR-93, miR-130a and miR-190, the difference in their expression between EB and hES cells and between EB and adult cells were significant (P < 0.05). [score:3]
The members of miR-17-92 cluster and its paralogs such as miR-106a, miR-106b, miR-93, and miR-17-5p are related to DNA replication and cell mitosis in cancer cells [60- 62], moreover, miR-17-5p and miR-20a can induce heterochromatic features in promoters that undergo overlapping transcription and possess sequence complementarity to the miRNA seed region [63]. [score:1]
The second was located on chromosome 7 and includes miR-25, miR-93 and miR-106b. [score:1]
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Li N. Muthusamy S. Liang R. Sarojini H. Wang E. Increased expression of miR-34a and miR-93 in rat liver during aging, and their impact on the expression of Mgst1 and Sirt1 Mech. [score:5]
Mir-93* targets AKT1, SESN2, NOX4, NFE2 and MGST1, which could possibly contribute to a pro-apoptotic function in the context of PDT. [score:2]
Fu X. Tian J. Zhang L. Chen Y. Hao Q. Involvement of microrna-93, a new regulator of pten/akt signaling pathway, in regulation of chemotherapeutic drug cisplatin chemosensitivity in ovarian cancer cells FEBS Lett. [score:2]
Long J. Wang Y. Wang W. Chang B. H. Danesh F. R. Identification of microrna-93 as a novel regulator of vascular endothelial growth factor in hyperglycemic conditions J. Biol. [score:1]
Espinosa-Parrilla Y. Munoz X. Bonet C. Garcia N. Vencesla A. Yiannakouris N. Naccarati A. Sieri S. Panico S. Huerta J. M. Genetic association of gastric cancer with mirna clusters including the cancer-related genes mir29, mir25, mir93 and mir106: Results from the epic-eurgast study Int. [score:1]
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Other miRNAs from this paper: hsa-mir-32
These are AATK (apoptosis associated tyrosine kinase) and SLC27A1 (solute carrier family 27 fatty acid transporter member 1), which are targeted by miR32 and mir93 [*], and CDKAL1 (CDK5 regulatory subunit associated protein 1-like 1), which is targeted by miR32 (Figure 5(b)). [score:6]
We also identified the mRNAs for three proteins, AATK, SLC27A1, and CDKAL1, to be preferentially secreted out in exosomes from Nef -expressing cells and their targeting miRNAs (miR32 and miR93 [*]) to be retained in these cells. [score:5]
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In ovarian cancer, 11 miRs were upregulated (miR-16, miR-20a, miR-21, miR-23a, miR-23b, miR-27a, miR-93, miR-141, miR-200a, miR-200b, and miR-200c) and 12 were downregulated (miR-10b, miR-26a, miR-29a, miR-99a, miR-100, miR-125a, miR-125b, miR-143, miR-145, miR-199a, miR-214, and let-7b). [score:7]
MiR-21, miR-29a, miR-92, miR-93, and miR-126 were significantly overexpressed in the serum of ovarian cancer patients compared to controls, while miR-99b, miR-127, and miR-155 were significantly underexpressed. [score:4]
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[+] score: 11
Both miR-93 and miR-106b directly target p21, resulting in its transcriptional silencing and impairment of its tumor-suppressing activity [14]. [score:6]
miR-25, miR-93, miR-106b, and miR-130 inhibit apoptosis by preventing the expression of the pro-apoptotic protein, Bim (Figure 2) [14]. [score:5]
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73
[+] score: 11
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-18a, hsa-mir-19a, hsa-mir-19b-1, hsa-mir-20a, hsa-mir-21, hsa-mir-22, hsa-mir-23a, hsa-mir-26a-1, hsa-mir-26b, hsa-mir-27a, hsa-mir-29a, hsa-mir-30a, hsa-mir-31, hsa-mir-33a, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-96, hsa-mir-29b-1, hsa-mir-29b-2, hsa-mir-103a-2, hsa-mir-103a-1, hsa-mir-16-2, hsa-mir-197, hsa-mir-199a-1, hsa-mir-208a, hsa-mir-148a, hsa-mir-30c-2, hsa-mir-30d, hsa-mir-7-1, hsa-mir-7-2, hsa-mir-7-3, hsa-mir-34a, hsa-mir-181a-2, hsa-mir-181b-1, hsa-mir-181c, hsa-mir-199a-2, hsa-mir-204, hsa-mir-210, hsa-mir-181a-1, hsa-mir-221, hsa-mir-222, hsa-mir-223, hsa-mir-224, hsa-mir-200b, hsa-let-7g, hsa-let-7i, hsa-mir-1-2, hsa-mir-15b, hsa-mir-27b, hsa-mir-30b, hsa-mir-122, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-125b-1, hsa-mir-130a, hsa-mir-132, hsa-mir-133a-1, hsa-mir-133a-2, hsa-mir-138-2, hsa-mir-140, hsa-mir-141, hsa-mir-142, hsa-mir-143, hsa-mir-145, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-138-1, hsa-mir-146a, hsa-mir-193a, hsa-mir-194-1, hsa-mir-195, hsa-mir-206, hsa-mir-320a, hsa-mir-200c, hsa-mir-1-1, hsa-mir-155, hsa-mir-181b-2, hsa-mir-194-2, hsa-mir-106b, hsa-mir-29c, hsa-mir-30c-1, hsa-mir-200a, hsa-mir-34b, hsa-mir-34c, hsa-mir-130b, hsa-mir-30e, hsa-mir-26a-2, hsa-mir-363, hsa-mir-365a, hsa-mir-365b, hsa-mir-369, hsa-mir-370, hsa-mir-371a, hsa-mir-375, hsa-mir-378a, hsa-mir-133b, hsa-mir-423, hsa-mir-448, hsa-mir-429, hsa-mir-486-1, hsa-mir-146b, hsa-mir-181d, hsa-mir-520c, hsa-mir-499a, hsa-mir-509-1, hsa-mir-532, hsa-mir-33b, hsa-mir-637, hsa-mir-320b-1, hsa-mir-320c-1, hsa-mir-320b-2, hsa-mir-378d-2, hsa-mir-509-2, hsa-mir-208b, hsa-mir-509-3, hsa-mir-103b-1, hsa-mir-103b-2, hsa-mir-320d-1, hsa-mir-320c-2, hsa-mir-320d-2, hsa-mir-378b, hsa-mir-320e, hsa-mir-378c, hsa-mir-378d-1, hsa-mir-378e, hsa-mir-378f, hsa-mir-378g, hsa-mir-378h, hsa-mir-378i, hsa-mir-371b, hsa-mir-499b, hsa-mir-378j, hsa-mir-486-2
[192] miR-33b Decrease lipogenesis via early B cell factor 1 (EBF1) targeting C/EBPα and PPARγ signaling[193] miR-93 Sirt7 and Tbx3[194] miR-125a ERRα[195] miR-130 Inhibition of adipogenesis by inhibiting PPARγ[66] miR-138 Inhibition of adipocyte differentiation via EID-1. Lipid droplet reduction[196] miR-145 Preadipocyte differentiation by targeting IRS1[197] miR-155 C/EBPβ pathway[198] mirR-193a/b Adiponectin production in the adipose tissue. [score:11]
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74
[+] score: 10
Cellular miRNAs miR-24 and miR-93 were reported to target viral large protein (L protein) and phosphoprotein (P protein) genes and their decreased expression increased vesicular stomatitis virus (VSV) replication [7]. [score:5]
The selected segment 5 (NP) of Influenza A H1N1 virus in Bangkok with selected human miRNA hsa-miR-93 and default binding parameters setting (e. g., minimum score, maximum energy, and alignment length is equal to 80, -15 KCal/Mol, and 10 bp, respectively), resulted in five distinct NP genes, each with two binding positions. [score:1]
The top ranked MREs of human microRNA hsa-miR-93 is consistent with previous literature while other results waited to be experimentally verified. [score:1]
Nine distinct binding sites of six human miRNAs, hsa-miR-25, hsa-miR-92a, hsa-miR-93, hsa-miR-99b, hsa-miR-191, and hsa-miR-503 were top ranked by the prioritization with binding energy lower than -37 KCal/Mol and having at most two amino acid substitutions without effects on physical properties (see Additional file 3). [score:1]
From Example 1 (see Figure 2A), with selected human miRNA hsa-miR-93 and two input sequences of segment 5 (NP), gi|8486129:A/Puerto Rico/8/34(H1N1) and gi|89779327: A/Puerto Rico/8/34(H1N1) of Influenza A virus with minimum score and maximum energy as 80 and -15 KCal/Mol, we got 18 records of input sequences with nine binding sites (see Figure 2B). [score:1]
Among them, an MRE of hsa-miR-93 with binding position 772-794 nt (from the start codon) was identified within segment 5 (NP) of the Influenza A H1N1 virus and consistent with Site 2 in [9]. [score:1]
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75
[+] score: 10
Other miRNAs from this paper: hsa-mir-96, hsa-mir-155, hsa-mir-20b
In a separate report, the downregulation of miR-93 expression in ovarian cancer cells caused an increase in Pol η levels. [score:6]
This negative regulation was validated using both a miR-93 mimic and an inhibitor [72]. [score:4]
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76
[+] score: 10
MiR-203 was indicated by a study [52] as an anti-metastatic miRNA in PC, intervening the advancement of the cancer via repressing a cohort of premetastatic targets; miR-93 was commonly overexpressed in PC patients and worked collectively with miR-106b and miR-375 to attenuate Capicua levels and facilitate PC progression [53]; a reduction or loss of miR-146b expression was suggested as an omen of PC invasion by the literature [54]; miR-486-5p, the 5p arm of the pre-miRNA for miR-486, stagnated the migration and invasion of PC by lowering the protein expression of Snail, a key regulator of the epithelial–mesenchymal transition for cancer metastasis [55]. [score:10]
[1 to 20 of 1 sentences]
77
[+] score: 9
In addition, Fu et al. demonstrated that miR-93*(miR-93-3p) was the most upregulated in active TB serum [27]; however, our analysis indicated that miR-93-3p was downregulated, making it a potential diagnostic marker to distinguish latent TB from active TB. [score:7]
01720Xhsa-miR-652-3p0.1640.00214Xhsa-miR-1273 g-3p0.3820.005491hsa-miR-21-5p0.1650.0005917hsa-miR-4668-5p0.3860.000139hsa-miR-142-5p0.1750.0005617hsa-miR-20b-3p0.3900.01073Xhsa-miR-36530.1780.0011722hsa-miR-148a-3p0.3910.000757hsa-miR-27b-3p0.1880.001339hsa-miR-483-3p0.3921.4E-0511hsa-miR-299-3p0.1910.0011214hsa-miR-44500.3930.000684hsa-miR-1260a0.1937.5E-0514hsa-miR-93-5p0.4000.007367hsa-miR-4445-5p0.2028.2E-053hsa-miR-56840.4050. [score:1]
0013219hsa-miR-301a-3p0.2070.0048517hsa-miR-45000.4130.0096213hsa-miR-451b0.2100.0055917hsa-miR-36540.4150.004007hsa-miR-1070.2160.0001010hsa-miR-223-3p0.4160.00199Xhsa-miR-196b-3p0.2260.000837hsa-miR-3607-5p0.4210.004125hsa-miR-5581-3p0.2299.8E-051hsa-miR-93-3p0.4220.001297hsa-miR-44170.2300.001241hsa-miR-24-3p0.4270.037889hsa-miR-185-5p0.2390.0136722hsa-miR-365a-3p0.4330.0003016hsa-miR-12750.2400.013796hsa-miR-1260b0.4340. [score:1]
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78
[+] score: 9
They observed upregulation of miR-21, miR-93, miR-205, and miR-708, and downregulation of miR-125b and miR-145 [18]. [score:7]
Additional dysregulated miRNAs contribute to LSCC progression, such as miR-93, miR-27a, and miR-375 [22– 24]. [score:2]
[1 to 20 of 2 sentences]
79
[+] score: 9
A. of microRNA expression identifies six members of the miR-17 (miR-17, miR-20a, miR-20b, miR-93, miR-106a and miR 106b) in normal liver and human HCC tumor tissue. [score:3]
Figure 1 A. of microRNA expression identifies six members of the miR-17 (miR-17, miR-20a, miR-20b, miR-93, miR-106a and miR 106b) in normal liver and human HCC tumor tissue. [score:3]
The miR-17 family (miR-17, miR-20a, miR 20-b, miR 106-a, miR 106-b, miR-93) and MYC expression was examined in human HCC from the cancer genome atlas (TCGA) [16]. [score:3]
[1 to 20 of 3 sentences]
80
[+] score: 9
The top four most significantly associated miRNAs - hsa-miR-130a (11q12.1), hsa-miR-22 (17p13.1), hsa-miR-93 (7q22.1) and hsa-miR-383 (8p22) - with DNA CNVs in breast cancer cell lines are shown in Figure 6. Figure 6 Association of miRNA expression with genomic copy number variation in breast cancer cell lines. [score:3]
The top four most significantly associated miRNAs - hsa-miR-130a (11q12.1), hsa-miR-22 (17p13.1), hsa-miR-93 (7q22.1) and hsa-miR-383 (8p22) - with DNA CNVs in breast cancer cell lines are shown in Figure 6. Figure 6 Association of miRNA expression with genomic copy number variation in breast cancer cell lines. [score:3]
Another group of 17 miRNAs (hsa-miR-575, hsa-miR-155, hsa-miR-26b, hsa-miR-200a, hsa-miR-200b, hsa-miR-141, hsa-miR-200c, hsa-miR-190b, hsa-miR-492, hsa-miR-640, hsa-miR-196a, hsa-miR-29c, hsa-miR-93, hsa-miR-193a-3p, hsa-miR-191, hsa-miR-26a, hsa-miR-182) showed significantly higher expression in the major cluster compared with the other miRNAs (fold change ≥ 1.5) (Figure 2, bottom red box). [score:2]
The majority of these miRNAs (hsa-miR-130a, hsa-miR-93, hsa-miR-383, hsa-miR29c, hsa-miR-382, hsa-miR-31) were already found to be located in regions that exhibited DNA copy number abnormalities in breast cancer tumors [65]. [score:1]
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81
[+] score: 9
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-15a, hsa-mir-16-1, hsa-mir-17, hsa-mir-18a, hsa-mir-20a, hsa-mir-21, hsa-mir-22, hsa-mir-23a, hsa-mir-24-1, hsa-mir-24-2, hsa-mir-25, hsa-mir-27a, hsa-mir-29a, hsa-mir-30a, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-101-1, hsa-mir-106a, hsa-mir-107, hsa-mir-16-2, hsa-mir-192, hsa-mir-196a-1, hsa-mir-199a-1, hsa-mir-129-1, hsa-mir-148a, hsa-mir-10b, hsa-mir-34a, hsa-mir-181b-1, hsa-mir-181c, hsa-mir-196a-2, hsa-mir-199a-2, hsa-mir-203a, hsa-mir-210, hsa-mir-212, hsa-mir-214, hsa-mir-215, hsa-mir-217, hsa-mir-218-1, hsa-mir-218-2, hsa-mir-221, hsa-mir-222, hsa-mir-223, hsa-mir-200b, hsa-let-7g, hsa-let-7i, hsa-mir-1-2, hsa-mir-15b, hsa-mir-27b, hsa-mir-122, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-125b-1, hsa-mir-130a, hsa-mir-141, hsa-mir-142, hsa-mir-143, hsa-mir-145, hsa-mir-153-1, hsa-mir-153-2, hsa-mir-9-1, hsa-mir-9-2, hsa-mir-9-3, hsa-mir-125a, hsa-mir-125b-2, hsa-mir-126, hsa-mir-129-2, hsa-mir-146a, hsa-mir-150, hsa-mir-185, hsa-mir-195, hsa-mir-206, hsa-mir-200c, hsa-mir-1-1, hsa-mir-155, hsa-mir-181b-2, hsa-mir-106b, hsa-mir-29c, hsa-mir-200a, hsa-mir-101-2, hsa-mir-34b, hsa-mir-34c, hsa-mir-130b, hsa-mir-376c, hsa-mir-375, hsa-mir-378a, hsa-mir-148b, hsa-mir-338, hsa-mir-335, hsa-mir-423, hsa-mir-20b, hsa-mir-429, hsa-mir-449a, hsa-mir-433, hsa-mir-451a, hsa-mir-193b, hsa-mir-520d, hsa-mir-503, hsa-mir-92b, hsa-mir-610, hsa-mir-630, hsa-mir-650, hsa-mir-449b, hsa-mir-421, hsa-mir-449c, hsa-mir-378d-2, hsa-mir-744, hsa-mir-1207, hsa-mir-1266, hsa-mir-378b, hsa-mir-378c, hsa-mir-378d-1, hsa-mir-378e, hsa-mir-378f, hsa-mir-378g, hsa-mir-378h, hsa-mir-4512, hsa-mir-378i, hsa-mir-203b, hsa-mir-451b, hsa-mir-378j
miR-106b and miR-93 abrogate TGFβ -induced apoptosis in GC cells by targeting the expression of BIM, encoding the pro-apoptotic protein BCL2-like 11, and thereby prevent apoptosis and cause tumor progression [26]. [score:5]
In addition, both miR-106b and miR-93 down-regulate p21, whereas miR-222 and miR-221 both control p27 and p57. [score:4]
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82
[+] score: 8
Translation of E2F1 is inhibited by miR-106b and miR-93 (derived from this cluster). [score:5]
The cluster encodes three miRNAs: miR-106b, miR-93, and miR-25 [35, 39]. [score:1]
Both the evolutionary sequence analysis and the seed-sequence -based grouping partition these miRNAs into four families: the miR-106 family (miR-17, miR-20a/b, miR-106a/b, and miR-93), the miR-18 family (miR-18a/b), the miR-19 family (miR-19a/b-1/2), and the miR-92 family (miR-25, miR-92a-1/2, and miR-363). [score:1]
This creates a negative feedback loop between E2F1 activity, miR-106b/miR-93, and transcription of the Mcm7 gene. [score:1]
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83
[+] score: 8
Four miRNAs (miR-93, miR-669c, miR-214, and miR-709) were especially upregulated, and proteomic profiling of the same samples demonstrated a significant correlation between the aforementioned miRNAs and expression of the corresponding gene targets associated with mitochondrial function, oxidative stress, and proliferation (Maes et al., 2008). [score:8]
[1 to 20 of 1 sentences]
84
[+] score: 8
Some of these miRNAs (e. g. miR-18a, miR-20a, miR-93) are upregulated in a high proportion of non-CBF-AML, and are associated with distinct AML subtypes (Additional file 1: Figure S3). [score:4]
The expression of miR-17, miR-18a, miR-20a, miR-93, and miR-181 in was evaluated from published gene expression datasets [24, 25]. [score:3]
Specifically, 52 non-CBF-AML and 31 CBF-AML were analyzed for miR-17, 31 non-CBF-AML and 18 CBF-AML were analyzed for miR-18a, 53 non-CBF-AML and 34 CBF-AML were analyzed for miR-20a, 34 non-CBF-AML and 18 CBF-AML were analyzed for miR-93 and miR-181. [score:1]
[1 to 20 of 3 sentences]
85
[+] score: 8
Among the miRNAs whose expression was suppressed by butyrate, members of the miR-106b family, including miR-17, miR-20a/b, miR-93 and miR-106a/b, regulate p21 translation and cancer cell proliferation [15, 16]. [score:8]
[1 to 20 of 1 sentences]
86
[+] score: 8
Other miRNAs from this paper: hsa-mir-17, hsa-mir-106a, hsa-mir-106b
Studies showed that DAB2 protein as a tumor suppressor inhibits cell growth in lung cancer, and miR-93, which is homologous to miR-106b, can inhibit its function in regulating the DAB2 protein level [27]. [score:8]
[1 to 20 of 1 sentences]
87
[+] score: 8
In an in-vitro study using H-69 lung cancer cell line, Pak et al. (2014) reported upregulation of miR-16-2, miR-93, miR-95, mir-153, mir-195, miR-199a-3p, and down-regulation of miR let7a, let7i, miR-124a in the presence of excretory secretory protein of C. sinensis. [score:7]
Among the miRNAs, seven of them (miR-16-2, miR-93, miR-95, miR-136, miR-153, miR-195, and miR-199a-3p) were mainly associated with esophageal adenocarcinoma, breast cancer and colorectal carcinoma, suggesting the role of these miRs in cell-proliferation and cell-signaling. [score:1]
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88
[+] score: 7
However, only a small number of microRNAs, including miR-28, miR-93, miR-144, miR-153, miR-27a, miR-132, and miR142-5p, have been confirmed experimentally to directly bind to the 3′UTR of Nrf2 and consequently downregulate gene expression [104– 108]. [score:7]
[1 to 20 of 1 sentences]
89
[+] score: 7
In human melanoma cells, miR-93 and miR-572 displayed cooperative inhibitory effects on the expression of cyclin -dependent kinase inhibitor p21 [20]. [score:7]
[1 to 20 of 1 sentences]
90
[+] score: 7
c-Myc downregulates TGFβ1 and TGFβ receptor 2 (TGFBR2), which is also a target of the miR-302 and miR-93 families [119, 121, 123, 133]. [score:6]
For instance, miR-93 and 106b (that belong to the same family and share 5/6 of the miR-302 seed) and the unrelated miR-138 enhance iPSC generation [118, 122]. [score:1]
[1 to 20 of 2 sentences]
91
[+] score: 7
Notably, several of the top expressed miRNAs (miR-93, cluster 520, cluster 302 and cluster 372-3) contain the same seed sequence and are thus predicted to target the same mRNAs (according to TargetScan 4.0). [score:7]
[1 to 20 of 1 sentences]
92
[+] score: 7
In particular, several endogenous miRNAs, miR-155, miR-181b and miR-93 have recently been shown to regulate AID at the level of translation via interaction with its 3′UTR [50]– [53]. [score:4]
Relative expression of miR-93, miR-155 and miR-181b was assessed via qRT-PCR analysis. [score:3]
[1 to 20 of 2 sentences]
93
[+] score: 7
We found that IFN-γ and TNF-α resulted in up-regulation of miR-155–5p, miR-4485–3p, miR-218–5p, and miR-146a–5p and down-regulation of miR-582–5p, miR-582–3p, miR-93–5p, miR-217 and miR-125b–5p (supplemental Tables S1 and S2). [score:7]
[1 to 20 of 1 sentences]
94
[+] score: 7
Patients with high miR-93/19a expression had a more favorable outcome compared to patients with high miR-221/222 expression (Figure  7C). [score:4]
In GBM, a high miR-93/miR-221 or a high miR-19a/miR-222 ratio was predictive of better overall survival. [score:1]
Specifically, we analyzed the effect of the two most significantly inversely correlated miRNA pairs (miR-93/miR-221 and miR-19a/miR-222, see Additional file 13: Table S12-2). [score:1]
For OvCa this was miR-222/miR-20a; for GBM it was miR-222/miR-19a, and for KIRC it was miR-221/miR-93. [score:1]
[1 to 20 of 4 sentences]
95
[+] score: 7
Additionally, host cell miRNAs were described targeting HDFs important for HIV-1 Tat -mediated LTR activation: PCAF (P300/CBP -associated factor), a histone acetylase is target by hsa-miR-20a and hsa-miR-17-5p (Triboulet et al., 2007); purine-rich element binding protein alpha (PUR-α), is targeted by hsa-miR-15a, hsa-miR-15b, hsa-miR-16, hsa-miR-20a, hsa-miR-93, and hsa-miR-106b (Shen et al., 2012); and cyclin T1 is repressed by hsa- miR-198 and hsa-miR-27b (Sung and Rice, 2009; Chiang et al., 2011). [score:7]
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96
[+] score: 7
We recently revealed that OBP-301 20 and tumor suppressor p53 -expressing OBP-301 (OBP-702) 21 increase the expression of cellular microRNAs (miRNAs) miR-7 and miR-93/106b, respectively, via activation of transcription factor E2F1 in human cancer cells. [score:7]
[1 to 20 of 1 sentences]
97
[+] score: 7
Other miRNAs from this paper: hsa-mir-17, hsa-mir-155, hsa-mir-520b
Fabbri E. Borgatti M. Montagner G. Bianchi N. Finotti A. Lampronti I. Bezzerri V. Dechecchi M. C. Cabrini G. Gambari R. Expression of microRNA-93 and Interleukin-8 during Pseudomonas aeruginosa -mediated induction of proinflammatory responses Am. [score:3]
Artificially increasing the levels of miR-93 (with pre-miR-93 transfection) could decrease IL-8 expression in these CF airway epithelial cells in vitro [70]. [score:3]
Fabbri et al. identified miR-93 as a miRNA that is decreased in IB3-1 and Cufi-1 cells infected with P. aeruginosa. [score:1]
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98
[+] score: 7
Patients with high expression of either miR-155, miR-17-3p, miR-106a, miR-93 or miR-21 and low expression of either let-7a-2, let-7b or miR-145 were found to have a significantly worse prognosis. [score:5]
A univariate Cox proportional hazard regression mo del with global permutation test in BRB-Array-Tools indicated that eight miRNAs (miR-155, miR-17-3p, miR-106a, miR-93, let-7a-2, miR-145, let-7b, and miR-21) were related to the adenocarcinoma patients survival. [score:2]
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99
[+] score: 7
The microRNAs upregulated with increasing emphysema severity included broadly conserved and poorly conserved microRNAs, with the most upregulated including miR-520e and miR-302d from the miR-93 family, miR-92a, miR-638, miR-211, and miR-150. [score:7]
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100
[+] score: 7
Selected pNL4-3 -downregulated miRNAs [miR-93 (B), miR-148b (C), miR-221 (D) and miR-16 (E)] were validated by real-time PCR. [score:4]
Using these assays, we checked the RAKE results in HeLa cells for four HIV-1 downregulated miRNAs (miR-93, miR-148b, miR-221 and miR-16) (Fig 5B, C, D and 5E). [score:3]
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