miR-335-3p通过靶向FOXD1发挥对骨肉瘤细胞肿瘤生物学行为的抑制作用

MiR-335-3p inhibits the biological behavior of osteosarcoma cells by targeting FOXD1

目的探讨miR-335-3p通过靶向调控FOXD1基因抑制骨肉瘤(osteosarcoma,OS)细胞的增殖和迁移的作用及机制。方法使用RT-PCR法评估miR-335-3p在OS组织和临近组织,正常人成骨细胞系(hFOB1.19f)和OS细胞系(SAOS2、U2OS、MG63)中的表达;使用细胞增殖实验测定研究miR-335-3p过表达对U2OS、MG63的活力的影响;EDU染色和DAPI染色评估miR-335-3p过表达对U2OS、MG63迁移和侵袭的影响;荧光素酶报告基因测定法确定miR-335-3p靶向调控FOXD1编码基因发挥作用;蛋白质免疫印迹法检测OS细胞中FOXD1蛋白表达;pcDNA-FOXD1转染OS细胞,并评估细胞增殖、迁移和侵袭的水平。结果miR-335-3p在OS癌组织和细胞中低表达;利用miR-335-33p mimic转染细胞后,可在体外抑制肿瘤细胞增殖、迁移和侵袭;miR-335-3p通过直接结合FOXD1 mRNA的3'-UTR发挥抑制作用,从而负面调节FOXD1的表达;miR-335-3p通过靶向调节FOXD1的表达,进而抑制OS癌细胞的增殖、迁移和侵袭。结论miR-335-3p可通过靶定FOXD1来抑制OS细胞的增殖、迁移和侵袭,并且可能是抗癌治疗中有前景的治疗方向。

Objective This paper aims to investigate the effect and mechanism of miR-335-3p on the proliferation and migration of osteosarcoma cells by targeting FOXD1.Methods The expression of miR-335-3p in osteosarcoma(OS)and adjacent tissues,normal human osteoblast line(hFOB1.19f)and OS cell line(SAOS2,U2OS,MG63)was evaluated by RT-PCR.The effect of miR-335-3p overexpression on the activity of U2OS and MG63 was determined by cell proliferation assay.The effects of miR-335-3p overexpression on migration and invasion of U2OS and MG63 were evaluated by EDU and DAPI.The luciferase reporter assay confirmed the role of miR-335-3p in the targeted regulation of FOXD1.pcDNA-FOXD1 was transfected into OS cells and the levels of cell proliferation,migration,and invasion were assessed.Results miR-335-3p was under-expressed in osteosarcoma carcinoma tissues and cells.After transfection with miR-335-3p mimic,the proliferation,migration and invasion of tumor cells could be inhibited in vitro.Furthermore,miR-335-3p negatively regulates FOXD1 expression by directly binding to the 3'-UTR of FOXD1mRNA.MiR-335-3p can inhibit the proliferation,migration and invasion of osteosarcoma cancer cells by targeting the expression of FOXD1.Conclusion miR-335-3p inhibit the proliferation,migration and invasion of osteosarcoma cells by targeting FOXD1,and may be a promising therapeutic direction in anticancer therapy.