蟾蜍灵对K562细胞凋亡诱导作用及bcl-2基因表达的影响  

Effect of Bufalin on Apoptosis of Myeloid Leukemia Cells and Expression of bcl-2 Gene

作　　者：贾彩云[1] 石贞玉[2] 呼文亮[3] 马远方[1] 

机构地区：[1]河南大学免疫学研究所,河南开封475001 [2]河南大学护理学院,河南开封475001 [3]武警医学院生物化学教研室,天津300612

出　　处：《河南职工医学院学报》2013年第5期537-540,共4页Journal of Henan Medical College For Staff and Workers

基　　金：天津市自然科学基金资助项目(013804311)

摘　　要：目的探讨中药蟾酥有效成分蟾蜍灵（bufalin）对人白血病细胞系K562细胞增殖和凋亡及bcl-2基因表达的影响。方法MTT实验测定K562细胞对蟾蜍灵的敏感性；原位缺口末端标记法（TUNEL）观察细胞凋亡率；琼脂糖凝胶电泳分析细胞凋亡的DNA断裂情况；半定量RT—PCR技术检测bcl-2mRNA表达。结果蟾蜍灵对K562细胞生长具有明显的抑制作用，MTT实验显示，蟾蜍灵对K562细胞的IC50值为0．013μmol·L^-1，统计学分析表明差异具有显著性（P〈0．01）；蟾蜍灵可明显诱导K562细胞调亡（P〈0．01），药物作用6h、12h和24hK562细胞的凋亡率分别为20．36％、34．35％和48．16％；经琼脂糖电泳，蟾蜍灵作用细胞可见到DNA梯形条带；bcl-2mRNA表达3h开始下降，6～12h持续下调，与对照组相比具有显著差异（P〈0．01）。结论蟾蜍灵对K562的增殖抑制作用可能是因为它的凋亡诱导作用，而K562细胞凋亡可能是由于bcl-2基因表达改变。Objective To explore the molecule mechanism and the effects of bufalin- inhibited growth and bufalininduced apoptosis in myeloid leukemia cell line K562. Methods The morphology and structure of K562 cells were observed by light microscopy and fluorescent microscopy. MTT assay were performed to evaluate the sensibility of K562 cells to bufalin. Apoptosis was detected by TUNEL method and DNA gel electrophoresis. The expression of bcl - 2 gene was detected by reverse - transcrip- tion polymerase chain reaction（RT- PC R）. Results Apparently morphological changes of cells treated by bufalin were detected by light microscopy and fluorescent microscopy. Compare with control, bufalin - treated K562 cells growth was remarkably reduced （P 〈 0.05）. By MTT assay, the IC50 value of bu- falin for K562 cells was 0. 013 μmol . L^-1 , the difference was significant by stastistic analysis （P 〈 0.01 ）. The apoptotic rate of K562 after treated with bufalin 6 h,12 h and 24 h were 20.36% ,34.35% and 48.16%, respectively. Agarose gel electrophoresis of genomic DNA from bufalintreated K562 cells showed typical DNA ladder. Bufalin could efficiently induce apoptosis of leukemia cells in a dose - and time -dependent manner. Campared with control, the down -regulation expression of bcl -2 mRNA in K562 occurred from 3 hour and decreased continuously from 6 hour to 12 hour, there was a significant difference （P 〈 0.05）. Conclusion Bufalin could efficiently induce apoptosis of leukemia K562 cells in a dose - and time - dependent manner; RT - PCR analysis suggest that bufalin induces apoptosis in human leukemia cells by altering the expression of bcl -2 gene.

关 键 词：蟾蜍灵 白血病 K562细胞 凋亡 基因表达 BCL-2 

分 类 号：R552[医药卫生—血液循环系统疾病]