The effect and mechanism of microRNA-83a targeting on differentiation of bone marrow mesenchymal stem cells in vitro

Abstract

Objective To study the anti-proliferation and inducing apoptosis effects of microRNA (miRNA, miR)-83a on bone marrow mesenchymal stem cells. Methods The experiment was divided into 3 groups: group A (PEF miR-83a and PSV2neo co-transfection group), group B (non-transfection group), and group C (PEF and PSV2neo co-transfection group). Group A served as the experimental group, and groups B and C group as the control groups. Transfection according to instructions, effects of application of expression detected miR-83a transcription of bone marrow mesenchymal stem cells lines in vitro antiproliferative effect of and biological function; by detecting the expression level of miR-83a immunohistochemical method and real-time quantitative polymerase chain reaction (Real-time PCR) technology in cells, Real-time PCR was measured after transfection, bone marrow mesenchymal stem cells lines Ras, Raf, protein kinase B (Akt) and extracellular signal-regulated kinase (ERK): mRNA expression changes, and through the methyl thiazol tetrazolium (MTT) method, the scratch test, hematoxylin and eosin (HE) staining, microscope observation and comparison of miR-83a transfection on the migration ability of cell proliferation, apoptosis and morphological changes. To analyse by using SPSS 14.0 statistical software. Results Real-time PCR and immunohistochemistry confirmed in miR-83a transfected cells with stable expression; Assingland staining results: after seventh days culture, the cells of the induction differentiation group were full of brown specific staining granules, and no specific staining granules were found in the undifferentiated differentiation group. Real-time PCR results: The expression of Ras, Raf and Akt mRNA in group A were 0.97±0.27, 0.54±0.18 and 0.99±0.31, in group B were 1.68±0.30, 1.58±0.25 and 2.06±0.42, in group C were 1.68±0.30, 1.58±0.25 and 1.89±0.31. Compared with group B and C, the levels of Ras, Raf and Akt mRNA in group A were significantly lower (t=1.531 and 1.838, P=0.016 and 0.011). MTT assay showed that the apoptotic rates of the cells were (23.49±4.64)%, (45.73±8.72)% and (59.25±13.16)% after 24, 48 and 72 h, respectively. There was significant difference between the two groups (χ2= 5.203 and 2.146, P =0.013 and 0.045). The scratch test showed that miR-83a transfected cell migration ability is lower than the normal cells. The effect of miR-83a on lung cancer cell 24 h, morphological observation of lung cancer cell apoptosis or necrosis. Conclusion Transfection of miR-83a can inhibit proliferation and promote apoptosis effect on pituitary adenoma cells in vitro cells, which is likely to play a role through inhibition of epidermal growth factor receptor (EGFR) and EGFRvⅢ signal transduction pathway in the RAS-RAF-mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3 kinase (PI3K)-Akt of two important signaling pathways activated. Key words: Bone marrow mesenchymal stem cells; MicroRNA-83a; Proliferation; Apoptosis; Biological function