表没食子儿茶素没食子酸酯对人宫颈癌细胞系HeLa细胞凋亡及bcl-2/bax表达的影响

目的 探讨表没食子儿茶素没食子酸酯(EGCG)对宫颈癌细胞系HeLa细胞凋亡及bcl-2/bax表达的影响。方法 体外培养HeLa细胞,分为对照组和实验组,实验组用不同浓度的EGCG处理24、48、72 h,对照组加入等体积的培养基。采用MTT法测定EGCG对HeLa细胞增殖的影响;倒置显微镜下观察细胞的形态学变化;EGCG(浓度分别为20、40、80μg/mL)处理HeLa细胞48 h后,分别采用流式细胞仪检测细胞凋亡和周期、分光光度计检测半胱氨酸天冬氨酸蛋白酶(Casepase-3)相对活性、Western blot法检测bcl-2/bax蛋白表达情况。结果 EGCG(浓度10~100μg/mL)能抑制HeLa细胞的增殖,呈剂量和时间依赖性;倒置显微镜下可观察到典型的细胞凋亡形态;浓度为20、40、80μg/mL的EGCG作用HeLa细胞48 h后,细胞凋亡率逐渐上升,分别为12.4%、23.4%、35.4%,明显高于对照组(3.3%);细胞周期结果显示EGCG能抑制细胞的G1期行进和G1/S转换,降低S期细胞比例;实验组HeLa细胞Caspase-3相对活性分别为(1.36±0.07)、(1.85±0.06)、(2.45±0.07),均高于对照组(0.93±0.06),差异均有统计学意义(P<0.05);bax蛋白表达量升高,而bcl-2蛋白表达量降低,呈剂量依赖性。结论EGCG能抑制HeLa细胞的增殖并诱导其凋亡,其作用机制可能与增加细胞Casepase-3活性、下调bcl-2基因的表达及上调bax基因有关。EGCG是一种前景广阔的抗肿瘤药物。

Effects of EGCG on cell apoptosis in cervical cancer HeLa cells and ex-pression of bcl-2/bax

Objective To explore the effect of EGCG on cell apoptosis in cervical cancer line HeLa cells and expres-sion of bcl-2/bax. Methods HeLa cells were cultured in v itro, and they were divided into two groups, including control group and experimental group. The control group was received only the culture medium, and the experimental group was treated by EGCG at different concentrations for 24、48、72 h respectively. The cell survival of the two groups were determined by the MTT method; the changes of cell morphology were observed by inverted microscope; apoptosis and cell cycle were analyzed by flow cytometry; the relative activity of caspase-3 was monitored by spectrophotometer; the changes of protein expression of bcl-2 and bax were detected by Western blot. Results From the data of MTT, the cell proliferation of human cervical cancer HeLa cells was inhibited by EGCG （10-100 μg/mL） in a dose-dependent and time-dependent manner. Typical apoptosis morphology of HeLa cells was observed by inverted microscope. Flow cy-tometry assays showed that EGCG significantly induced apoptosis in HeLa cells. 48 h after treated with EGCG （20, 40, 80μg/mL）, the apoptosis rate of the experimental group were increased gradually, they were 12.4%, 23.4%, 35.4% re-spectively, higher than that of the control group （3.3%） obviously. Flow cytometry assays demonstrated that EGCG blocked the cell cycle at the G1 to S transition and decreased the percentage of S-phase cells. The relative activity of Caspase-3 of experimental group were （1.36±0.07）, （1.85±0.06） and （2.45±0.07） respectively, higher than that of the control group （0.93±0.06）, the differences were statistically significant （P〈0.05）. The data of Western blot showed that EGCG down-regulated bcl-2 and up-regulated bax in a dose-dependent manner. Conclusion EGCG can inhibit the proliferation of HeLa cells and promote apoptosis, and the anticancer effect of EGCG may be associated with the down regulation of bcl-2 expression and up regulation of