miR-195表达在乳腺癌细胞对阿霉素药物敏感性中的意义

Significance of microRNA-195 expression in breast cancer cells to adriamycin drug sensitivity

目的调控miR-195在乳腺癌细胞中的表达对阿霉素药物(ADR)敏感性的影响及其作用机制进行研究。方法利用脂质体将成熟miR-195模拟物(miR-195mimic)及阴性对照转染至乳腺癌细胞MCF-7,采用实时定量聚合酶链式反应(RT-PCR)检测细胞中miR-195的表达变化;采用MTT比色法及流式细胞术(FCM)检测转染miR-195模拟物及其对照经ADR作用后细胞的增殖及凋亡;Western-blot检测细胞中Raf-1、P-gp、Bcl-2蛋白的表达。结果转染miR-195mimic后,明显上调了细胞中miR-195的表达水平(P<0.01);在miR-195mimic与ADR联合作用后,明显抑制了细胞的增殖,其敏感性与对照组相比,提高近2.5倍(P<0.05),同时促进了细胞的凋亡(P<0.05);Western-blot检测发现在转染miR-195mimic后,细胞中Raf-1蛋白及BCL-2、P-gp蛋白的表达显著降低(P<0.01)。结论在乳腺癌细胞中,miR-195作为抑癌基因,能够有效促进乳腺癌细胞对ADR所介导的化疗敏感性,其作用机制至少部分是通过miR-195负性调控Raf-1蛋白的表达来实现。

Objective Study on the possibility of regulating Micro RNA （miRNA）195 expression to increase the sensitivity of the breast cancer cell （MCF-7） to adriamycin （ADR） and its mechanism .Methods Transfect through liposome the mature Micro RNA（miRNA）195 mimic and its negative contrast into MCF-7 and detect chan-ges of Micro RNA（miRNA）195 in the cells by Real-time PCR ;detect Micro RNA（miRNA）195 mimic and its contrast by MTT colorimetric method and FCM ,and detect cells′reproduction and apoptosis after the effect of ADR ;detect by Western-blot the protein expression of Raf-1 ,P-gp and Bcl-2 in the cells .Results The Micro RNA（miRNA） 195 expression in the cells increases significantly （P〈0 .01） after the transfection of Micro RNA （miRNA）195 mimic;the combined effect of Micro RNA （miRNA）195 mimic and ADR significantly suppresses the cell reproduction ,its sensitivity increases almost 2 .5 times（P〈0 .05） in comparison with the control group ,and the apoptosis rate of the cells increases ;it is observed that ,by Western-blot detection ,the protein expressions of Raf-1 ,Bcl-2 and P-gp in the cells decrease significantly（P〈0 .01） after the transfection of Micro RNA （miRNA）195 mimic .Conclusion Micro RNA（miRNA）195 ,as the tumor suppressor gene in breast cancer cells ,can effectively increase the chemotherapy sensitivity of breast cancer cells to ADR mediation .Its mechanism is realized at least partly by Micro RNA （miRNA） 195′s negative regulation on Raf-1 protein .