Effect of miRNA-147 regulates proliferation, apoptosis and migration via targeting VEGF in human retinal pigment epithelial cells

Abstract

AIM: To investigate the effect of miRNA-147 targeted regulation of vascular endothelial growth factor(VEGF)on the proliferation, apoptosis and migration of human retinal pigment epithelial cells, and to explore its molecular mechanism. METHODS: Human retinal pigment epithelial(ARPE-19)cells were selected and divided into 7 groups: blank control group(untreated), nonsense miRNA group(transfected with mimic NC), miRNA-147 simulant group(transfected with miRNA-147 mimic), inhibitor negative control group(transfected with shRNA NC), VEGF inhibitor group(transfected with shRNA VEGF), miRNA-147 simulant+empty viral vector group(transfected with miRNA-147 mimic and pcDNA3.1)and miRNA-147 simulant+VEGF overexpression group(transfected with miRNA-147 mimic and pcDNA3.1 VEGF). RT-qPCR was used to detect the expression of miRNA-147 and VEGF mRNA. Dual luciferase experiments were used to verify the targeting relationship between miRNA-147 and VEGF. Western blot was used to detect the expression of VEGF protein. MTT method was used to detect the proliferation. Flow cytometry to detect the apoptosis level and cell cycle changes. Cell scratch test to detect the level of cell migration. RESULTS: Compared with the blank control group and the nonsense miRNA group, the expression level of miRNA-147 in miRNA-147 simulant group was significantly increased, while the expression levels of VEGF mRNA and protein were significantly reduced(P<0.05). Compared with the inhibitor negative control group, the expression levels of VEGF mRNA and protein in the VEGF inhibitor group were significantly reduced(P<0.05). Compared with the miRNA-147 simulant+empty viral vector group, the expression level of VEGF mRNA in the miRNA-147 simulant+VEGF overexpression group was significantly increased(P<0.05). The dual luciferase report shows that VEGF is the target gene of miRNA-147. Transfection of miRNA-147 mimic and shRNA VEGF can reduce the proliferation and migration of ARPE-19 cells and promote apoptosis can reduce the proliferation and migration of ARPE-19 cells and promote apoptosis(P<0.05). Transfection VEGF overexpression reverses the effect of miRNA-147 mimics on proliferation, migration and apoptosis of ARPE-19 cells(P<0.05). CONCLUSION: miRNA-147 can inhibit ARPE-19 cell proliferation, migration and promote cell apoptosis by targeting VEGF.

Keywords

• mirna-147
• vascular endothelial growth factor(vegf)
• human retinal pigment epithelial cells
• proliferation
• apoptosis
• migration