摘要
目的考察丹参酮IIa(tanshinone IIa,Tan IIa)诱导人胃癌细胞的凋亡作用,以及对端粒酶活性的影响。方法不同浓度TanⅡa处理人胃癌细胞株MKN-45后,显微镜下观察胃癌细胞形态变化,应用Annexin V/PI双重染色方法检测细胞凋亡比例,并用凋亡相关抗体检测通路情况,用RT-PCR以及TRAP-PCR法分别对胃癌细胞端粒酶基因hTert和端粒酶活性进行检测。结果TanⅡa处理胃癌细胞后,DAPI染色显示,细胞核内染色质聚缩,细胞核碎裂并有凋亡小体产生,提示TanⅡa诱导胃癌MKN-45细胞凋亡。Annexin V/PI双染结果显示,凋亡比例随化合物浓度提高而增加,40μM下晚期凋亡比例达到(45.02±6.48)%。凋亡相关蛋白的检测显示,PARP和Caspase-3活化,细胞色素C水平增加,而Caspase-8则无显著变化。同时TRAP-PCR结果显示,Tan IIa在诱导细胞凋亡的同时伴随端粒酶活性下调,并在40μM下显著抑制hTert基因的表达。结论 TanⅡa对人胃癌细胞具有明显诱导细胞凋亡作用,且可能通过激活内源性途径引起细胞凋亡。TanⅡa还可抑制端粒酶活性和hTERT基因表达,这些可能是其发挥抗癌作用的重要机制之一。
Objective To investigate the induced apoptosis effect of tanshinone IIa(Tan IIa)on human gastric cancer cells,as well as telomerase activity. Methods After treatment with different concentration of Tan Ⅱ,gastric cancer cell line MKN-45 cells were stained with DAPI and morphology changes were observed under a microscope,Annexin V /PI double staining was carried to detect the proportion of apoptosis cells,furthermore apoptosis pathway relates were also detected by western blot. Then RT-PCR and TRAP-PCR methods were used to detect the expression of telomerase gene hTERT and telomerase activity respectively. Results After Tan IIa treatment of gastric cancer cells,DAPI staining showed that the nuclear chromatin condensation and polymerization,nuclear fragmentation and apoptotic bodies produced,all of which indicated Tan IIa could induce the apoptosis of gastric cancer cell line MKN-45. Annexin V /PI double staining showed that the proportion of apoptotic increased with the increased concentration of Tan IIa,and ratio of later apoptotic cells could reached(45. 02 ± 6. 48)% at 40 μM. Detection of apoptosis-related proteins showed that,PARP and Caspase-3 activation,Cyto C levels increased,and Caspase-8 showed no significant changes. Meanwhile,TRAP-PCR results showed that,in company with Tan IIa-induced apoptosis,telomerase activity and the expression of hTERT genes were significantly inhibited under 40 μM. Conclusion Tan IIa induce the apoptosis of human gastric cancer cells through activation of endogenous pathways,futhermore,the inhibition of telomerase activity and hTERT gene expression,which may be one of the important mechanisms by these pharmacologic activity.
出处
《中国生化药物杂志》
CAS
北大核心
2014年第5期43-47,50,共5页
Chinese Journal of Biochemical Pharmaceutics
基金
邯郸市科学技术与发展计划项目(1223108085-4)
