miR-143对宫颈癌HeLa细胞增殖和凋亡的影响以及对Bcl-2表达的调节

摘要

Aim To explore the effect of miR-143 on the poliferation and apoptosis of human cervical carcinoma, and investigate the role of miR-143 in the expression of Bc1-2 in HeLa cells.Methods Over-expression plasmids with miR-143, anti-miR-143 and control miRNA precursor were inserted into the HeLa cells using lipofectamine 2000.Methyl thiazolyl tetrazolium( MTT )was used to analyze the proliferation of the HeLa cells , and cell apoptosis was quantitated by Histone/DNA ELISA.The level of Bc1-2 protein and mRNA was measured by using Western blot and RT-PCR respectively.Dual-luciferase reporter gene assay was used to detect the target sites of miR-143.Results miR-143 inhibited significantly HeLa cell proliferation and promoted apoptosis, while anti-miR-143 promoted the cell proliferation, and suppressed the apoptosis.Sustained over-expression of miR-143 resulted in repression of Bcl-2 protein, and anti-miR-143 increased Bc1-2.whereas the levels of Bc1-2 mRNA were so minor.MiR-143 repressed the activity of a luciferase reporter carrying the 3 '-untranslated region of Bc1-2 , mutated the predicted binding site between miR-143 and Bc1-2.and the activity of luciferase was reversed.Conclusion miR-143 can inhibit proliferation and promote apoptosis in HeLa cells.at least in part by targeting Bc1-2.%目的 研究miR-143对宫颈癌HeLa细胞增殖和凋亡的影响,探讨miR-143对宫颈癌HeLa细胞系Bcl-2蛋白及mRNA表达的调节作用.方法 将miR-143,anti-miR-143以及microRNA空对照高表达质粒用脂质体转染进宫颈癌HeLa细胞中,MTT法检测宫颈癌HeLa细胞增殖的变化,Histone/DNA ELISA检测细胞凋亡;Western blot和RT-PCR检测Bcl-2蛋白和mRNA的表达情况;双荧光素酶报告基因法检测miR-143的靶位点.结果 MiR-143抑制HeLa细胞的增殖,促进细胞凋亡,而anti-miR-143促进HeLa细胞增殖,抑制细胞凋亡;高表达的miR-143抑制Bcl-2蛋白的表达,anti-miR-143增加Bcl-2蛋白的表达量,而Bcl-2 mRNA的变化很小;miR-143抑制Bcl-2 3′端的荧光素酶的活性,突变miR-143与Bcl-2的预测结合位点后,荧光素酶的活性恢复.结论 MiR-143至少部分通过打靶Bcl-2抑制宫颈癌HeLa细胞增殖,并且促进HeLa细胞凋亡.