miR-27b-3p调控SMAD1对骨肉瘤细胞增殖、迁移和侵袭作用的影响

目的 寻找并验证miR-27b-3p和SMAD1的关系,探讨miR-27b-3p对骨肉瘤细胞增殖、侵袭、迁移作用的影响,为骨肉瘤靶向治疗提供理论依据。 方法 采用生物信息学方法预测候选靶基因;应用双荧光素酶报告实验确定miR-27b-3p和SMAD1靶向关系;采用qRT-PCR法选择SAOS-2骨肉瘤细胞系。采用miR-27b-3p转染SAOS-2细胞后,分为miR-27b-3p inhibitor组、空白对照组和阴性对照组。采用MTT、Transwell迁移和侵袭实验检测miR-27b-3p对SAOS-2细胞的影响;采用Western blotting法检测沉默miR-27b-3p后SMAD1的表达量。 结果 生物信息学检测显示,miR-27b-3p与SMAD1-UTR存在结合位点;双荧光素酶报告实验显示,miR-27b-3p inhibitor组SMAD1的表达量高于阴性对照组,差异有统计学意义(P<0.05),SMAD1是miR-27b-3p的靶基因。SAOS-2细胞MTT、Transwell迁移实验和侵袭实验结果均显示,miR-27b-3p inhibitor组与空白对照组和阴性对照组细胞数相比降低,差异有统计学意义(P<0.05),miR-27b-3p inhibitor组细胞的增殖、迁移和侵袭能力降低。miR-27b-3p inhibitor组与阴性对照组SMAD1蛋白表达量相比明显降低(P<0.05)。 结论 miR-27b-3p可以通过调控SMAD1的表达促进骨肉瘤细胞增殖、迁移和侵袭作用。miR-27b-3p可能在骨肉瘤细胞中起着促癌基因作用。

Effect of miR-27b-3p regulating SMAD1 on osteosarcoma cell proliferation,migration and invasion

Objective To assess the relationship between miR-27b-3p and SMAD1 and to explore the effects of MiR-27b-3p on osteosarcoma cell proliferation, invasion and migration, so as to provide theoretical basis for targeted therapy of osteosarcoma. Methods Candidate target genes were predicted with bioinformatics methods. The targeting relationship between miR-27b-3p and SMAD1 was determined with dual luciferase report experiments. The SAOS-2 osteosarcoma cell line was selected with qRT-PCR. After SAOS-2 cells were transfected with miR-27b-3p, they were divided into 3 groups: miR-27b-3p inhibitor group, blank control group and negative control group. The effects of miR-27b-3p on SAOS-2 cells were detected with MTT, Transwell migration and invasion assay. The expression of SMAD1 after miR-27b-3p silencing was determined with Western blotting. Results Bioinformatics analysis showed there were binding sites of miR-27b-3p and SMAD1-UTR. Dual luciferase experiments showed the miR-27b-3p inhibitor group had higher expression of SMAD1 than the negative control group(P<0.05), and SMAD1 was the target gene of miR-27b-3p. MTT, Transwell migration and invasion assay showed the miR-27b-3p inhibitor group had reduced cell proliferation, migration and migration than the blank control group and negative control group(P<0.05). The miR-27b-3p inhibitor group had significantly lower protein expression of SMAD1 than negative control group(P<0.05). Conclusion miR-27b-3p can promote the proliferation, migration and invasion of osteosarcoma cells by regulating the expression of SMAD1, and may play a role as an oncogene in osteosarcoma cells.