siRNA沉默 B7-H4 表达对人肺腺癌A549细胞增殖、侵袭和迁移的影响

目的:探讨靶向B7-H4基因的小干扰RNA(small interference RNA,siRNA)对人肺腺癌细胞A549增殖、侵袭和迁移的影响。方法:体外化学合成B7-H4特异性siRNA(B7-H4-siRNA),转染A549细胞,MTT法、流式细胞术、Western blotting分别检测A549细胞的增殖、细胞周期,以及B7-H4和Cyclin D1蛋白的水平;Transwell实验检测A549细胞体外侵袭和迁移能力。结果:B7-H4-siRNA成功转染入A549细胞,Western blotting结果显示,B7-H4-siRNA转染抑制A549细胞中B7-H4和Cy-clin D1蛋白的表达。B7-H4-siRNA转染后A549细胞增殖活性明显下降,B7-H4-siRNA组A549细胞的倍增时间明显长于未转染组、Ctrl-siRNA组和转染试剂对照组[(33.78±0.26)h vs(28.69±0.18)、(27.32±0.13)、(26.93±0.19)h,P<0.05]。B7-H4-siRNA转染使A549细胞阻滞在G1期。B7-H4-siRNA组A549细胞的侵袭细胞数明显少于未转染组[(89.80±0.99)个vs(186.20±1.33)个,P<0.05]。B7-H4-siRNA组A549细胞的迁移细胞数明显少于未转染组、Ctrl-siRNA组和转染试剂对照组[(60.20±0.37)个vs(102.57±0.52)、(100.72±0.31)、(98.65±0.21)个,P<0.05〗。结论:B7-H4-siRNA能沉默A549细胞中B7-H4的表达,抑制A549细胞的增殖、侵袭和迁移,B7-H4可能成为肺癌基因治疗的候选靶点。

Effects of silencing B7-H4 expression by small interference RNA on proliferation, invasion and migration of human lung adenocarcinoma A549 cells

Objective:To investigate the effects of silencing B7-H4 expression by small interference RNA(siRNA) on proliferation,invasion and migration of human lung adenocarcinoma A549 cells.Methods: Chemically synthesized siRNA targeting B7-H4(B7-H4-siRNA) was transfected into A549 cells.The proliferation of A549 cells was determined by MTT assay,the cell cycle was detected by flow cytometry,the levels of B7-H4 and Cyclin D1 were verified by Western blotting,and the invasion and migration ability was detected by Transwell assay.Results: B7-H4-siRNA was successfully transfected into A549 cells.Western blotting results showed that B7-H4-siRNA transfection inhibited the expressions of B7-H4 and Cyclin D1 in A549 cells.In the B7-H4-siRNA group,the proliferation of A549 cells was significantly down-regulated,the doubling time was longer than that in the untransfected group,the Ctrl-siRNA group and the empty vector group([33.78±0.26] h vs [28.69±0.18],[27.32±0.13],[26.93±0.19] h,P<0.05),and the cell cycles were arrested in G1 phase.The invasion of A549 cells in the B7-H4-siRNA group was inhibited significantly as compared with the untransfected group([89.80±0.99] vs [186.20±1.33],P<0.05),and the migration of A549 cells in the B7-H4-siRNA group was suppressed significantly as compared with the untransfected group,the Ctrl-siRNA group and the empty vector group([60.20±0.37] vs [102.57±0.52],[100.72±0.31],[98.65±0.21],P<0.05).Conclusion: B7-H4-siRNA can silence B7-H4 expression in A549 cells,and inhibit the proliferation,invasion and migration of A549 cells effectively.B7-H4 can be regarded as a candidate gene for lung cancer gene therapy.