紫草素诱导人绒毛膜癌JEG-3细胞凋亡机制的研究  

Mechanisms of Shikonin-induced Apoptosis in JEG-3 Cells

作　　者：黄巍巍[1] 孟松树[1] 潘芹[1] 吴建富[1] 胡茂志[1] 范健[1] 

机构地区：[1]扬州大学生物科学与技术学院,江苏扬州225009

出　　处：《癌变．畸变．突变》2009年第6期426-430,共5页Carcinogenesis,Teratogenesis & Mutagenesis

基　　金：江苏省自然科学基金(BK2006071);江苏省卫生厅项目(H200763)

摘　　要：背景与目的:研究紫草素(shikonin)诱导人绒毛膜癌JEG-3细胞的凋亡作用及其机制。材料与方法:采用四甲基偶氮唑盐(MTT)法测定紫草素对JEG-3细胞生长的抑制作用;Hoechst33258荧光染色和流式细胞术(FCM)检测紫草素处理JEG-3细胞后发生凋亡的形态变化;Western blot检测凋亡相关蛋白的活化。结果:MTT分析表明,紫草素抑制JEG-3细胞增殖,并呈时间和剂量依赖性(P<0.01),其半数有效抑制浓度(IC50)为(6.3±0.6)μmol/L;紫草素处理JEG-3细胞后,Hoechst33258染色出现典型的凋亡特征,且FCM检测出现明显的亚二倍体峰,Annexin V/PI双染出现早期凋亡细胞;Western blot检测结果显示经紫草素处理后JEG-3细胞的Caspase-3、ERK、JNK蛋白均被激活,而PARP蛋白被剪切成cleaved PARP(85 KD)。结论:紫草素可能经Caspase-3途径诱导人绒毛膜癌细胞JEG-3凋亡,并且MAPK通路的活化、抑制对细胞的凋亡有一定的影响。[ABSTRACTJ BACKGROUND AND AIM： To investigate proliferation-inhibiting effects and mechanisms of shikonin on human choriocarcinoma JEG-3 cells. MATERIALS AND METHODS： 3-（4, 5- dimethyhhiazol-2-yl） -2, 5- diphemyhetra-zolium Bromide （MTT） assay was used to determine the inhibitory rate of shikonin on the proliferation of JEG-3 cells. Apoptosis induced by shikonin was detected with Hoechst 33258 dye , flow cytometry （FCM） and Annexin V/PI assay. Western blot was used to evaluate the changes of pro-caspase-3, cleaved PARP, active MAPK in protein levels in JEG-3 cells. RESULTS： Shikonin induced JEG-3 apoptosis in a time- and dose-dependent manner. The IC50 of a 24 h time course for JEG-3 cells was 6.3 ±0.6 Vmol/ L. Typical morphological changes of apoptosis were observed in JEG-3 cells with Hoechst staining after induced by shikonin. The shikonin-treated JEG-3 cells with condensed and fragmented chromatin, apoptosis peak and hypo-diploid cells were revealed by proidum iodide （PI） staining. Furthermore, Annexin-V/PI staining showed the early apoptotic cells. To elucidate the apoptotic pathways induced by shikonin, we assessed the expression of active caspase-3, cleavages of poly（ADP-ribose） polymerase （PARP） and MAPK. Caspase-3 in JEG-3 cells was activated after 12 h treatment and PARP was cleaved subsequently. Phosphorylation of ERK and JNK was respectively blocked after 8 h and 12 h treatment. CONCLUSION： Shikonin could significantly inhibit the proliferation of JEG-3 cell and induce apoptosis. Activation and inhibition of MAPK pathway may affect apoptosis.

关 键 词：紫草素 人绒毛膜癌细胞 细胞凋亡 CASPASE-3 MAPK 

分 类 号：R730.5[医药卫生—肿瘤]