CAJ | 학술논문

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(VCM)检测紫草素处理JEG-3细胞后发生凋亡的形态变化;Wstern 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(IC_(50))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.
배경여목적:연구자초소(shikonin)유도인융모막암JEG-3세포적조망작용급기궤제.재료여방법:채용사갑기우담서염(MTT)법측정자초소대JEG-3세포생장적억제작용;Hoechst33258형광염색화류식세포술(VCM)검측자초소처리JEG-3세포후발생조망적형태변화;Wstern blot검측조망상관단백적활화.결과:MTT분석표명,자초소억제JEG-3세포증식,병정시간화제량의뢰성(P<0.01),기반수유효억제농도(IC_(50))위(6.3±0.6)μmol/L;자초소처리JEG-3세포후,Hoechst33258염색출현전형적조망특정,차FCM검측출현명현적아이배체봉,Annexin V/PI쌍염출현조기조망세포;Western blot검측결과현시경자초소처리후JEG-3세포적Caspase-3、ERK、JNK단백균피격활,이PARP단백피전절성cleaved PARP(85 KD). 결론:자초소가능경Caspase-3도경유도인융모막암세포JEG-3조망,병차MAPK통로적활화、억제대세포적조망유일정적영향.
BACKGROUND AND AIM: To investigate proliferation-inhibiting effects and mechanisms of shikonin on human choriocarcinoma JEG-3 cells. MATERIALS AND METHODS: 3-(4, 5- dimethylthiazol-2-yl) -2, 5- diphemyltetra-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 IC_(50) of a 24 h time course for JEG-3 cells was 6.3 ±0.6 μmol/ 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.