RNPC1 inhibits non-small cell lung cancer progression via regulating miR-181a/CASC2 axis

Objectives

To study the roles and mechanisms of RNA binding protein RNPC1 in non-small cell lung cancer progression.

Results

RNPC1 and long non-coding RNA CASC2 expression levels were significantly downregulated in lung cancer tissues compared with normal adjacent tissues, and their expression levels were positively correlated. Functionally, overexpression of RNPC1 or CASC2 inhibited non-small cell lung cancer cells proliferation, migration and invasion, and promoted cells apoptosis. Mechanistically, RNPC1 was found to harbor binding sites on CASC2 and directly bound to CASC2, and increased CASC2 mRNA stability and expression. Notably, the promotive effects of RNPC1 on CASC2 expression were attenuated by miR-181a overexpression. Moreover, CASC2 3′UTR with mutated miR-181a binding sites did not respond to RNPC1 alteration. Finally, the inhibitory effects of RNPC1 overexpression were attenuated or even reversed by CASC2 knockdown or miR-181a overexpression.

Conclusions

RNA bind protein RNPC1 could inhibit non-small cell lung cancer progression by competitively binding to CASC2 with miR-181a.

     

Bioinformatics prediction of miR-30a targets and its inhibition of cell proliferation of osteosarcoma by up-regulating the expression of PTEN

Background

MiRNAs are frequently abnormally expressed in the progression of human osteosarcoma. Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is one of the tumor suppressors in various types of human cancer. In the present study, we detected how hsa-miR-30a-3p regulated PTEN and further tested the role of hsa-miR-30a-3p in the cell proliferation of osteosarcoma cells.

Methods

The levels of miR-30a were determined by real time PCR. The expression of PTEN was tested by western blotting analysis. Cell distribution of PTEN was observed with confocal laser scanning microscope. Cell viability was determined by MTT assay.

Results

The expression of miR-30a and PTEN was obviously decreased in MG-63, 143B and Saos-2 cells compared with primary osteoblasts. TargetScan analysis data showed miR-30a might bind with position 30-57 of 3’UTR of PTEN. Transfection with miR-30a-3p increased the level of PTEN in MG-63 cells, while transfection with miR-30a-3p inhibitor significantly decreased the expression of PTEN in osteosarcoma cells. Transfection with miR-30a-3p significantly inhibited cell proliferation of osteosarcoma cells, while miR-30a inhibitor obviously promoted cell viability of MG63 cells and Saos-2 cells. Inhibition of PTEN eliminated the proliferation inhibitory effect of miR-30a-3p.

Conclusion

Thus, all these findings revealed the anti-tumor effects of miR-30a in human osteosarcoma cells, which could be mediated by regulating the level of PTEN.

     

Circ_0027599/PHDLA1 suppresses gastric cancer progression by sponging miR-101-3p.1

Background

Pleckstrin homology-like domain family A member 1 (PHLDA1) is a tumor suppressor gene in gastric cancer, but its role regulated by circular RNAs (circRNAs) is not known. CircRNAs are important regulators in cancer growth and progression, however, the molecular roles of circRNAs in gastric cancer are rarely known. The study was aimed to investigate the role of circRNAs in regulating PHLDA1 expression in gastric cancer.

Results

The circRNA expression profile in the gastric cancer tissues by circRNA microarray showed that hsa_circ_0027599 (circ_0027599) was significantly down-regulated in gastric cancer patients and cells when comparing with the controls. Circ_0027599 overexpression suppressed gastric cancer cell proliferation and metastasis. By using bioinformatics tools and luciferase reporter assays, circ_0027599 was verified as a sponge of miR-101-3p.1 (miR-101) and suppressed cancer cell survival and metastasis. It was also verified that PHLDA1 was regulated by circ_0027599 in gastric cancer cells.

Conclusions

The study uncovered that PHLDA1 was regulated by circ_0027599/miR-101, which suppressed gastric cancer survival and metastasis in gastric cancer.

     

Non-small cell lung cancer: miR-30d suppresses tumor invasion and migration by directly targeting NFIB

Objective

To evaluate the role and the molecular mechanism of miR-30d in non-small cell lung cancer (NSCLC).

Results

qRT-PCR was used to detect miR-30d expression in NSCLC tissues and cell lines. miR-30d was frequently down-regulated in NSCLC and its expression was associated with clinicopathological features of NSCLCC patients. Over-expression of miR-30d notably inhibited cell migration and invasion in NSCLC cell lines. miR-30d could negatively regulate Nuclear factor I B (NFIB) by directly targeting its 3′-UTR, which was confirmed by luciferase assay. NFIB also reversed miR-30d-mediated suppression on the migration and invasion in NSCLC cell lines.

Conclusion

miR-30d suppressed cell migration and invasion by directly targeting NFIB in NSCLC, and NFIB could partially abrogated the inhibition of biological functions by miR-30d.

     

Silencing CDK4 radiosensitizes breast cancer cells by promoting apoptosis

Background

The discovery of molecular markers associated with various breast cancer subtypes has greatly improved the treatment and outcome of breast cancer patients. Unfortunately, breast cancer cells acquire resistance to various therapies. Mounting evidence suggests that resistance is rooted in the deregulation of the G1 phase regulatory machinery.

Methods

To address whether deregulation of the G1 phase regulatory machinery contributes to radiotherapy resistance, the MCF10A immortalized human mammary epithelial cell line, ER-PR-Her2+ and ER-PR-Her2- breast cancer cell lines were irradiated. Colony formation assays measured radioresistance, while immunocytochemistry, Western blots, and flow cytometry measured the cell cycle, DNA replication, mitosis, apoptosis, and DNA breaks.

Results

Molecular markers common to all cell lines were overexpressed, including cyclin A1 and cyclin D1, which impinge on CDK2 and CDK4 activities, respectively. We addressed their potential role in radioresistance by generating cell lines stably expressing small hairpin RNAs (shRNA) against CDK2 and CDK4. None of the cell lines knocked down for CDK2 displayed radiosensitization. In contrast, all cell lines knocked down for CDK4 were significantly radiosensitized, and a CDK4/CDK6 inhibitor sensitized MDA-MB-468 to radiation induced apoptosis. Our data showed that silencing CDK4 significantly increases radiation induced cell apoptosis in cell lines without significantly altering cell cycle progression, or DNA repair after irradiation. Our results indicate lower levels of phospho-Bad at ser136 upon CDK4 silencing and ionizing radiation, which has been shown to signal apoptosis.

Conclusion

Based on our data we conclude that knockdown of CDK4 activity sensitizes breast cancer cells to radiation by activating apoptosis pathways.

     

Long non-coding RNA H19 regulates E2F1 expression by competitively sponging endogenous miR-29a-3p in clear cell renal cell carcinoma

Background

Numerous recent studies indicate that the long non-coding RNAs (lncRNAs) are frequently abnormal expressed and take critical roles in many cancers. Renal cell carcinoma is the secondary malignant tumors in the urinary system and has high mortality and morbidity. Around 80% of RCCs is clear cell renal cell carcinoma (ccRCC) and is characterized by high metastasis and relapse rate. However, the clinical significances of lncRNAs in ccRCC are still unknown.

Methods

The human cancer lncRNA PCR array (Yingbio) was performed to detect the differentially expressed lncRNAs in human ccRCC samples. Real-time PCR (RT-PCR), dual-luciferase assay, RNA binding protein immunoprecipitation (RIP) assay, transwell assay, CCK-8 assay, and western blot were performed to explore the molecular mechanism of lncRNAs in ccRCC cell migration and invasion.

Results

In this study, lncRNA-H19 was high expressed and negatively correlated with miR-29a-3p in ccRCC. By bioinformatics software, dual-luciferase reporter and RIP assays, we verified that miR-29a-3p was identified as a direct target of lncRNA-H19. RT-PCR and western blot demonstrated that down-regulated lncRNA-H19 could affect the expression of miR-29a-3p targeting E2F1 with competitively binding miR-29a-3p. Furthermore, transwell assays indicated that lncRNA-H19 knockdown inhibited cells migration and invasion, but this effect was attenuated by co-transfection of lncRNA-H19 siRNA and miR-29a-3p inhibitor. Over expression of E2F1 could rescue lncRNA-H19 siRNA induced suppression on cell migration and invasion in ccRCC cells.

Conclusions

These results show a possible competing endogenous RNAs regulatory network involving lncRNA-H19 regulates E2F1 expression by competitively sponging endogenous miR-29a-3p in ccRCC. This mechanism may contribute to a better understanding of ccRCC pathogenesis, and lncRNA-H19 may be further considered as a potential therapeutic target for ccRCC intervention.

     

miR-130b-3p/301b-3p negatively regulated Rb1cc1 expression on myogenic differentiation of chicken primary myoblasts

Objectives

To explore the roles of miR-130b-3p and miR-301b-3p which may regulate Rb1-inducible coiled-coil 1 (Rb1cc1) expression during myogenic differentiation of chicken primary myoblasts.

Results

After 4 days of myogenic differentiation, myotubes appeared and after 6 days the cells fused to each other and expression of MyHC could be detected by immunofluorescence staining. TargetScan and RNAhybrid 2.2 showed miR-130b-3p and miR-301b-3p were well complementary with the target site of Rb1cc1 3′-untranslated region (3′-UTR). Using the dual-luciferase assay, we found miR-130b-3p and miR-301b-3p could inhibit Rb1cc1 expression by binding to its 3′-UTR. Real-time PCR showed Rb1cc1 mRNA expression level was almost reciprocal to that of miR-130b-3p or miR-301b-3p during myogenic differentiation. Furthermore, over-expression of miR-130b-3p or miR-301b-3p down-regulated the expression levels of Rb1cc1, myoblast determination protein, myogenin and myosin heavy chain.

Conclusions

miR-130b-3p or miR-301b-3p negatively regulate Rb1cc1 expression to affect myogenic differentiation.

     

MicroRNA-190b confers radio-sensitivity through negative regulation of Bcl-2 in gastric cancer cells

Objectives

To determine the role of miR-190b in radio-sensitivity of gastric cancer (GC).

Results

In radio-resistant GC cells, down-regulation of miR-190b and up-regulation of Bcl-2 were observed. The protein expression of Bcl-2 was negatively regulated by miR-190b. Overexpression of miR-190b significantly decreased cell viability and enhanced radio-sensitivity of GC cells. Of note, these effects of miR-190b on GC cells radio-sensitivity were abolished by Bcl-2.

Conclusion

miR-190b confers radio-sensitivity of GC cells, possibly via negative regulation of Bcl-2.

     

The lncRNA MEG3 downregulation leads to osteoarthritis progression via miR-16/SMAD7 axis

Background

Osteoarthritis (OA) is a chronic joint disease and there is no a definitive cure at present. Long non-coding RNAs (lncRNAs) have been confirmed to play important roles in the development of OA. However, the underlying mechanism of lncRNA maternally expressed gene 3 (MEG3) in OA has not been well elucidated.

Methods

The rat OA model and interleukin-1β (IL-1β)-induced rat chondrocytes were constructed. The expression pattern of lncRNA MEG3 and miR-16 was detected by RT-qPCR assay in cartilage tissues of rat OA model. The effect of MEG3 and miR-16 on IL-1β-induced chondrocytes was evaluated on the basis of cell viability and apoptosis. Then, the interaction among MEG3, miR-16 SMAD7 was explored by dual-luciferase reporter assay and RIP assay.

Results

It is found that lncRNA MEG3 was down-regulated and miR-16 was up-regulated in rat OA cartilage tissues. MEG3 knockdown promoted proliferation and inhibited apoptosis, while miR-16 knockdown suppressed proliferation and promoted apoptosis in IL-1β-induced rat chondrocytes. Moreover, MEG3 was involved in miR-16 pathway and MEG3 suppressed miR-16 expression. Additionally, SMAD7 was a target gene of miR-16 and miR-16 suppressed SMAD7 expression in IL-1β-induced chondrocytes. Moreover, the expression of SMAD7 induced by MEG3 or si-MEG3 was markedly reversed by the introduction of miR-16 or anti-miR-16. Furthermore, MEG3 exerted its anti-proliferation and pro-apoptosis by regulating miR-16 and SMAD7.

Conclusion

MEG3 was down-regulated and miR-16 was up-regulated in cartilage tissues of rat OA model. MEG3 knockdown might lead to the progression of OA through miR-16/SMAD7 axis.

     

Targeting of HER3 with Functional Cooperative miRNAs Enhances Therapeutic Activity in HER2-Overexpressing Breast Cancer Cells

Background

The HER3 receptor functions as a major cause of drug resistance in cancer treatment. It is believed that therapeutic targeting of HER3 is required to improve patient outcomes. It is not clear whether a novel strategy with two functional cooperative miRNAs would effectively inhibit erbB3 expression and potentiate the anti-proliferative/anti-survival effects of a HER2-targeted therapy (trastuzumab) and chemotherapy (paclitaxel) on HER2-overexpressing breast cancer cells.

Results

Combination of miR-125a and miR-205, as compared to either miRNA alone, potently inhibited expression of HER3 in HER2-overexpressing breast cancer BT474 cells. Co-expression of the two miRNAs not only reduced the levels of phosphorylated erbB3 (P-erbB3), Akt (P-Akt), and Src (P-Src), it also inhibited cell proliferation and increased cells at G1 phase. A multi-miRNA lentiviral vector - the cluster of miR-125a and miR-205 - was constructed to simultaneously express the two miRNAs in HER2-overexpressing breast cancer cells. Concurrent expression of miR-125a and miR-205 via the miRNA cluster transfection significantly enhanced trastuzumab-mediated growth inhibition and cell cycle G1 arrest in BT474 cells and markedly increased paclitaxel-induced apoptosis in another HER2-overexpressing breast cancer cell line HCC1954.

Conclusions

Here, we showed that functional cooperative miRNAs effectively suppressed erbB3 expression. This novel approach targeting of HER3 was able to enhance the therapeutic efficacy of trastuzumab and paclitaxel against HER2-overexpressing breast cancer.

     

Evaluation of miR-21 and miR-150 expression in immune thrombocytopenic purpura pathogenesis: a case-control study

Background

Immune thrombocytopenic purpura (ITP) is a common autoimmune disorder diagnosed with thrombocytopenia and bleeding symptoms due to production of autoantibodies (Abs) against platelets. Nowadays, microRNAs are known as novel biomarkers for diagnosis of diseases. The aim of this study was to investigate the expression levels of miR-21 and miR-150 in ITP patients and determine the role of these miRNAs in ITP pathogenesis.

Materials and Methods

Thirty newly diagnosed patients with acute ITP and 30 healthy subjects( age and sex matched) as controls were enrolled in this study. The expression level of miR-21 and miR-150 was investigated using Real-time-PCR. Comparison of demographic characteristics of the cases was done using independent t-test and chi-square test. Comparison of the expression level of miR-21 and miR-150 with the related parameters was done using independent t-test or Mann–Whitney and Kruskal–Wallis test. Spearman rho correlation coefficient was used to investigate the relationship between the expression of miR-21 and miR-150 with demographic characteristics.

Results

The expression of miR-21, 150 in the patients was not different compared with the control group in general. A significant relationship between the expression of miR-21 with hemoglobin, hematocrit and red blood cell hemoglobin concentration was observed.

Discussion

Expression of miR-21 and miR-150 is not associated with pathogenesis of acute ITP and can involve the synergistic role of other miRNAs. Investigation of miR-21 and miR-150 expression along with other miRNAs and cytokines can be helpful in diagnosis and pathogenesis of ITP.

     

miR-34a attenuates glioma cells progression and chemoresistance via targeting PD-L1

Objective

To investigate the roles of miR-34a in progression and chemoresistance of glioma cells.

Results

Quantitative real-time PCR analysis showed that miR-34a level was lower, but PD-L1 expression level was higher in glioma tissue specimens compared with normal brain tissues and their expression levels were negatively correlated. Ectopic expression of miR-34a inhibited glioma cell proliferation, promoted cell cycle arrest in G1/S phase and cell apoptosis. Additionally, miR-34a/PD-L1 axis was again confirmed and co-expression of PD-L1 with miR-34a mimics attenuated the effects of miR-34a on cell proliferation and apoptosis in glioma cells. Importantly, PD-L1 overexpression resulted in chemoresistance in glioma cells, this effect was attenuated by miR-34a overexpression.

Conclusions

miR-34a inhibits glioma cells progression and chemoresistance via targeting PD-L1.

     

MiRNA-145 suppresses lung adenocarcinoma cell invasion and migration by targeting N-cadherin

Objective

To investigate the roles of miR-145 in lung adenocarcinoma (LAC) and to clarify the regulation of N-cadherin by miR-145.

Results

In 57 paired clinical LAC tissues, diminished miR-145 was significantly correlated with the lymph node metastasis and was negatively correlated with N-cadherin mRNA level expression. Wound healing and transwell assays revealed a reduced capability of tumor metastasis induced by miR-145 in LAC. miR-145 negatively regulated the invasion of cell lines through targeting N-cadherin by directly binding to its 3′-untranslated region. Silencing of N-cadherin inhibited invasion and migration of LAC cell lines similar to miR-145 overexpression.

Conclusions

MiR-145 could inhibit invasion and migration of lung adenocarcinoma cell lines by directly targeting N-cadherin.

     

KIAA0101, a target gene of miR-429, enhances migration and chemoresistance of epithelial ovarian cancer cells

Background

Ovarian cancer is a common type of gynecological malignancies, and is the fifth leading cause of cancer-related death in women in the United States. MiR-429 and KIAA0101 have been found to be involved in several human malignancies, respectively. However, the role of miR-429 and KIAA0101, and the correlation between them during development of epithelial ovarian cancer (EOC) remain to be investigated.

Methods

The expression of KIAA0101 in EOC tissues and cells was measured by Quantitative real-time PCR, western blot, and immunochemistry. Cell proliferation assay, colony formation assay, and transwell assay was performed to assess the role of miR-429 and KIAA0101 in regulation of proliferation, migration, and chemoresistance of EOC cells. Luciferase assay was used to test the Wnt/β-catenin signaling activity in response to depletion of KIAA0101 and overexpression of miR-429.

Results

We found that KIAA0101 was upregulated in metastatic EOC tissues, compared to primary EOC tissues, and KIAA0101 was required for the migration activity and chemoresistance of EOC cells by enhancing Wnt/β-catenin signaling. Furthermore, we revealed KIAA0101 is direct target of miR-429. Similar to knockdown of KIAA0101, overexpression of miR-429 reduced invasion and chemoresistance of EOC cells. Co-transfection of KIAA0101 partially abrogates the inhibitory effects on invasion and chemoresistance in EOC cells.

Conclusions

KIAA0101, a target gene of miR-429, was upregulated in the metastatic EOC tissues, and enhanced the migration activity and chemoresistance of EOC cells. Both miR-429 and KIAA0101 may represent the potential therapeutic targets of EOC.

     

Intramuscular injection of mechano growth factor E domain peptide regulated expression of memory-related <Emphasis Type="Italic">sod</Emphasis>, miR-134 and miR-125b-3p in rat hippocampus under simulated weightlessness

Objective

To investigate the expression of memory-related antioxidant genes and miRNAs under simulated weightlessness and the regulation of mechano growth factor (MGF) E domain, the peptide preventing nerve damage.

Results

Igf-iea and mgf mRNA levels, expression of antioxidant genes sod1 and sod2 and levels of miR-134 and miR-125b-3p increased in rat hippocampus after 14 days tail suspension to simulate weightlessness which was inhibited with intramuscular injection of E domain peptide. Therefore, administration of MGF E domain peptide could reverse increased expressions of memory-related igf-iea, mgf, sod1, sod2, miR-134 and miR-125b-3p in rat hippocampus under simulated weightlessness.

Conclusions

MGF may regulate the redox state and miRNA-targeted NR-CREB signaling, and intramuscular injection may be the alternative administration because of its safety, convenience and ability to pass through the blood brain barrier.

     

Anticancer bioactive peptide-3 inhibits human gastric cancer growth by targeting miR-338-5p

Background

Cancer incidence and mortality have been increasing in China, making cancer the leading cause of death since 2010 and a major public health concern in the country. Cancer stem cells have been studied in relation to the treatment of different malignancies, including gastric cancer. Anticancer bioactive peptide-3 (ACBP-3) can induce the apoptosis of gastric cancer stem cells (GCSCs) and reduce their tumorigenicity. In the present study, for the first time, we used a miRNA microarray and bioinformatics analysis to identify differentially expressed miRNAs in ACBP-3-treated GCSCs and GCSC-derived tumors in a xenograft model and functionally verified the identified miRNAs. miR-338-5p was selected based on its significant upregulation by ACBP-3 both in cultured GCSCs and in tumor tissues.

Results

miR-338-5p was downregulated in GCSCs compared with normal gastric epithelial cells, and the ectopic restoration of miR-338-5p expression in GCSCs inhibited cell proliferation and induced apoptosis, which correlated with the upregulation of the pro-apoptotic Bcl-2 proteins BAK and BIM. We also found that ACBP-3-treated GCSCs could respond to lower effective doses of cisplatin (DDP) or 5-fluorouracil (5-FU), possibly because ACBP-3 induced the expression of miR-338-5p and the BAK and BIM proteins and promoted GCSC apoptosis.

Conclusions

Our data indicate that miR-338-5p is part of an important pathway for the inhibition of human gastric cancer stem cell proliferation by ACBP-3 combined with chemotherapeutics. ACBP-3 could suppress GCSC proliferation and lower the required effective dose of cisplatin or 5-fluorouracil. Therefore, this study provides not only further evidence for the remarkable anti-tumor effect of ACBP-3 but also a possible new approach for the development of GCSC-targeting therapies.

     

MicroRNA-1284 inhibits proliferation and induces apoptosis in SGC-7901 human gastric cancer cells

Objectives

To explore the functional effects of miR-1284 on gastric cancer cells.

Results

Overexpression of miR-1284 significantly reduced SGC-7901 cell proliferation, but improved apoptosis. However, miR-1284 suppression displayed the inversed impacts. Furthermore, the protein levels of p27, Bax, procaspase-3 and active caspase-3 were up-regulated by miR-1284 overexpression, but were down-regulated by miR-1284 suppression. The level of Bcl-2 was down-regulated by miR-1284 overexpression, while it was up-regulated by miR-1284 suppression. The level of p21 was unaffected.

Conclusion

These results suggest that miR-1284 overexpression might be a suppressor for gastric cancer via controlling of cell proliferation and apoptosis.