Objective: To study the effects and mechanisms of miR-181a-5p on the proliferation, cycle and migration of HOS osteosarcoma cells.
Methods: Real-time quantitative PCR was used to detect the expression of miR-181a-5p and HOXB4 in osteoblast hFOB1.19 cell line and osteosarcoma cell lines (HOS, U2OS, MG63). miR-181a-5p mimics and miR-181a-5p inhibitors were respectively transfected into HOS cells by Lipofectamine 2000, and miR NC group was set as control group. CCK-8 method was used to detect the change in cell proliferation. Flow cytometry was used to detect the changes in cell cycles. Wound healing experiments and Transwell migration experiments were used to detect the changes in cell migration ability. The target gene of miR-181a-5p was predicted by Targetscan website and validated by Dual-luciferase reporter gene system and Western blot.
Results: Compared with osteoblast hFOB1.19, miR-181a-5p was low expressed in osteosarcoma cells HOS, U2OS, and MG63(P<0.05), while HOXB4 was high expressed in osteosarcoma cells HOS, U2OS, and MG63(P<0.05). Compared with the miR NC group, over expression of miR-181a-5p inhibited the proliferation and migration of osteosarcoma HOS cells, and the number of cells in S phase decreased(P<0.05). However, knockdown miR-181a-5p promoted the proliferation and migration of osteosarcoma HOS cells, the cells in S phase increased(P<0.05). Bioinformatics prediction and Dual-luciferase reporter gene system validate HOXB4 as a downstream target gene of miR-181a-5p(P<0.05). Western blot showed that miR-181a-5p over expression or knockdown significantly down-regulated or up-regulated HOXB4 expressions in the HOS cells respectively(P<0.05).
Conclusion: miR-181a-5p is down expressed in osteosarcoma cells, and over-expression miR-181a-5p inhibits the proliferation, cell cycle and migration ability of osteosarcoma cells by targeting HOXB4.
Keywords: Cell cycle; Cell migration; Cell proliferation; Osteosarcoma.