Effect of microRNA-129 expression on proliferation,apoptosis and cell cycle of esophageal squamous cell cancer and its possible molecular mechanism / 中国药理学与毒理学杂志

ABSTRACT

OBJECTIVE To investigate the effect of microRNA-129(miR-129)expression on malignant phenotypes of esophageal squamous cell cancer(ESCC) cells and its possible molecular mechanisms. METHODS The constructed miR-129-overexpressed vector (pGCMV/EGFP/miR-129) and negative control vector (pGCMV/EGFP/miR-NC) were stably transfected into ESCC cell lines (Eca109 and EC9706),respectively. Quantitative real-time PCR(qRT-PCR)was performed to detect the expression of miR-129. MTT and flow cytometry(FCM)assays were performed to analyze the effects of miR-129 on proliferation, cell cycle and apoptosis of ESCC cells. Furthermore,a luciferase reporter vector with the putative B-cell lymphoma-2(Bcl-2)3′-untranslated region(pLUC/Bcl-2-3′-UTR-wt and pLUC/Bcl-2-3′-UTR-mut)was constructed to explore whether Bcl-2 was a direct target gene of miR-129 by detecting luciferase activity. Next,Western blotting was performed to detect the expression of Bcl-2, cleaved caspase 3 and total caspase 3 proteins. RESULTS Overexpression of miR-129 significantly inhibited proliferation(P<0.01),induced cell arrest in G0/G1 phase(P<0.05)and enhanced apoptosis (P<0.05)in ESCC cells. Luciferase reporter assay indicated that Bcl-2 was identified as a direct target gene of miR-129. Results of Western blotting showed that overexpression of miR-129 significantly reduced the expression of Bcl-2 protein and increased the expression of cleaved caspase 3 protein,but induced no changes in total caspase 3 protein in ESCC cells. CONCLUSION miR-129 functions as a tumor suppressor in ESCC cells by targeting Bcl-2 gene. Therefore,miR-129 will be a potential molecular target for the treatment of human ESCC.