脱氢表雄酮诱导CD4+T细胞系Jurkat细胞凋亡的初步探索

为研究脱氢表雄酮(DHEA)对Jurkat细胞增殖活性和凋亡的影响,用不同浓度的DHEA处理Jurkat细胞12 h、24 h后,用MTT法测定细胞活性;Hoechst染色观测细胞形态的改变;碘化丙啶(PI)染色检测细胞凋亡;Annexin V/PI双染法检测细胞膜变化;DNA阶梯状电泳分析细胞DNA断裂;荧光显微镜分析细胞线粒体膜电位变化;并用RT-PCR和流式检测凋亡时细胞中Fas和FasL的mRNA和蛋白表达的情况。结果显示,DHEA在10~(-7)mol/L时对Jurkat细胞有显著的增殖抑制作用(P<0.05)和诱导凋亡效果,细胞凋亡率比对照组分别升高2.48%和2.52%;Annexin V/PI双染法检测,处理组凋亡细胞比率比对照组也明显增加;且随着DHEA剂量的增加细胞线粒体膜电位倒塌明显增强,Fas和FasL的mRNA和蛋白水平也随之增加。以上结果表明,DHEA在高浓度时对Jurkat细胞有一定的抑制增殖并诱导其凋亡的作用,Fas/FasL通路和膜电位的改变在此机制中发挥了相应的作用。

Dehydroepiandrosterone induces apoptosis in Jurkat cells

The aim of this study is to investigate effects of DHEA on the viability and apoptosis of Jurkat cells,in which the effect of DHEA on apoptosis in Jurkat cells was determined by DNA ladder,Hoechst33258,PI staining and Annexin-V- FITC/PI double staining.RT-PCR method was used to analyze Fas and FasL mRNA expression and the Fas/FasL proteins were evaluated by flow cytometry.Changes in mitochondrial membrane potential were detected with JC-1 fluorescence.It was demonstrated viability of Jurkat cells was down regulated,and numbers of Annexin V-positive and propidium iodide-negative cells were increased significantly after DHEA treatment.The sub-diploid peak of cells with fragmented DNA were also in creased vs control.Agarose gel electrophoresis of DHEA-treated Jurkat cells showed DNA characteristic fragmentation of ap- optotic cells.JC-1 staining showed cells with green fluorescence increased significantly vs control.Fas and FasL mRNA and protein expression of Jurkat cells was also increased in DHEA treated classes than that of controls.The results revealed that DHEA could induce apoptosis in Jurkat cells in vitro and the Fas/FasL pathway and mitochondrion membrane potential muta- tion play roles in these regulations.