CAJ | 학술논문

目的探讨雷公藤甲素(TP)诱导慢性粒细胞白血病(CML) K562/G01细胞凋亡的线粒体可能机制.方法用不同浓度的TP作用于K562/G01细胞,用四甲基偶氮唑蓝(MTT)法检测各实验组的细胞毒性作用,用流式细胞术检测各实验组细胞的凋亡率、线粒体膜电位及Caspase-9活性,用实时荧光定量聚合酶链反应(PCR)法检测Caspase-9及细胞色素C的mRNA水平,用Western blot检测细胞色素C的蛋白水平.结果 TP对K562/G01细胞的生长增殖均有抑制作用,呈时间-剂量依赖性(均P＜ 0.001).同时TP可以使细胞的线粒体膜电位消退、使Caspase-9的活性增高(均P＜ 0.001),TP上调Caspase-9和细胞色素C的转录水平(均P＜0.001)及细胞色素C的蛋白表达水平(0、10、20、40、80 nmol/LTP作用48 h后,细胞色素C对应条带灰度值分别为10.54±0.75、21.54±0.59、39.63±0.58、53.29±1.47、75.68±1.87,均P＜ 0.001),且上述作用均呈明显的剂量依赖性.结论雷公藤甲素可以有效抑制K562/G01细胞生长增殖,而线粒体凋亡途径很可能是TP促进CML细胞凋亡的多途径之一.
목적 탐토뢰공등갑소(TP)유도만성립세포백혈병(CML) K562/G01세포조망적선립체가능궤제.방법 용불동농도적TP작용우K562/G01세포,용사갑기우담서람(MTT)법검측각실험조적세포독성작용,용류식세포술검측각실험조세포적조망솔、선립체막전위급Caspase-9활성,용실시형광정량취합매련반응(PCR)법검측Caspase-9급세포색소C적mRNA수평,용Western blot검측세포색소C적단백수평.결과 TP대K562/G01세포적생장증식균유억제작용,정시간-제량의뢰성(균P＜ 0.001).동시TP가이사세포적선립체막전위소퇴、사Caspase-9적활성증고(균P＜ 0.001),TP상조Caspase-9화세포색소C적전록수평(균P＜0.001)급세포색소C적단백표체수평(0、10、20、40、80 nmol/LTP작용48 h후,세포색소C대응조대회도치분별위10.54±0.75、21.54±0.59、39.63±0.58、53.29±1.47、75.68±1.87,균P＜ 0.001),차상술작용균정명현적제량의뢰성.결론 뢰공등갑소가이유효억제K562/G01세포생장증식,이선립체조망도경흔가능시TP촉진CML세포조망적다도경지일.
Objective To investigate the possible mechanism of mitochondrial in chronic myeloid leukemia cells K562/G01 cells apoptosis induced by triptolide.Methods K562/G01 cells were treated with different concentrations of triptolide.MTT assay was used to assess cytotoxic effect.FCM was used to determine apoptosis rate,mitochondrial membrane potential and the activity of Caspase-9 of each experimental group.Real-time quantitative PCR assay was used to quantify mRNA levels of Caspase-9 and cytochrome C and Western blot assay was used to determine protein levels of cytochrome C.Results Triptolide inhibited the growth and proliferation of K562/G01 cells in a time-and dose-dependent manner (both P ＜ 0.001).Meantime,triptolide could make the mitochondria membrane potential fade away and enhance the activity of Caspase-9 (F =566.431,2 555.485,P ＜ 0.001).In addition,triptolide could dose-dependently up-regulated the transcription of Caspase-9 and cytochrome C (F =61 007.702,452 121.760,P ＜ 0.001),and the protein expression of cytochrome C,whose gray value in each experimental group was 21.54±0.59,39.63±0.58,53.29± 1.47 and 75.68±1.87 (F =5 677.928,P ＜ 0.001) respectively.Conclusion Triptolide could potently inhibit the growth and proliferation of K562/G01 cells,and the mitochondria apoptosis pathway might be one of the important apoptosis mechanisms in chronic myeloid leukemia cells induced by triptolide.