消痰散结方对结肠癌干细胞样细胞凋亡的影响及相关机制研究  

Study on the effect of Xiaotan Sanjie Recipe on apoptosis of colon cancer stem-like cells and its related mechanism

作　　者：周昱岐[1,2] 张映城[1] 魏品康[1] 叶敏[1] 

机构地区：[1]第二军医大学附属长征医院中医科,上海200003 [2]解放军第一〇一医院血液肿瘤科,江苏无锡214044

出　　处：《中国医药导报》2017年第24期17-20,共4页China Medical Herald

基　　金：上海市卫生和计划生育委员会中医药科研基金项目(2014JP012A);上海市卫生和计划生育委员会科研课题(20144Y0247)

摘　　要：目的探讨消痰散结方对结肠癌干细胞样细胞(CCSCs)凋亡的影响及相关机制。方法采用无血清培养法由HCT116结肠癌细胞诱导培养CCSCs,通过检测CCSCs标志物CD44进行鉴定。消痰散结方浸膏冻干粉末溶解在细胞培养液中,制备消痰散结方保存液。以不同浓度消痰散结方(0.125、0.25、0.5、1.0 mg/mL)作用CCSCs 72 h,Annexin-V/PI双染色结合流式细胞术检测各浓度消痰散结方对CCSCs的影响。Western blot检测药物干预后Bax、Bcl-2蛋白表达变化。结果 HCT116细胞在无血清培养下成球生长,CCSCs中CD44+细胞占(49.69±1.89)%。消痰散结方可诱导CCSCs,其凋亡率与对照组(0 mg/mL)比较,差异有统计学意义(P<0.05),其凋亡效应呈浓度依赖性。消痰散结方作用后,Bcl-2家族的促凋亡蛋白Bax表达增强,抑凋亡蛋白Bcl-2表达减弱,以0.5、1.0 mg/mL两组浓度效应更为显著。结论从HCT116结肠癌细胞成功诱导富集CCSCs,消痰散结方能剂量依赖性地诱导CCSCs,其机制可能与促凋亡蛋白Bax表达增强、抑凋亡蛋白Bcl-2表达减弱相关。Objective To investigate effect of Xiaotan Sanjie Recipe on apoptosis of colon cancer stem-like cells（CCSCs） and its related mechanism. Methods The CCSCs were induced from colon cancer cell line HCTl16 in serum-free medium. The expressions of CD44 were detected to identify CCSCs. The lyophilized powder of Xiaotan Sanjie Recipe Extract was dissolved in cell culture fluid, so as to make preserving fluid of Xiaotan Sanjie Recipe. CCSCs were treated with different concentrations of Xiaotan Sanjie Recipe（0.125, 0.25, 0.5, 1.0 mg/mL） for 72 h. The effects of different concentrations of Xiaotan Sanjie Recipe for CCSCs were detected by Annexin-V/PI staining combined with flow cytometry. The expression changes of Bax, Bcl-2 protein after drug intervention were detected by Western blot. Results HCT116 cells formed cancer stem cell spheres in serum-free medium. The proportion of CD44 cells in cell spheres was（49.69 ±1.89）%. Xiaotan Sanjie Recipe could induce the cell apoptosis, the apoptosis rate was statistically significant different compared with the control group（0 mg/mL）（P 0.05）, the apoptotic effects showed concentration-dependent.After application of Xiaotan Sanjie Recipe, the expression of pro-apoptotic protein Bax of Bcl-2 family was increased,meanwhile the expression of anti-apoptotic protein Bcl-2 was decreased. The effect was more pronounced in 0.5 mg/mL and 1.0 mg/mL concentration groups. Conclusion CCSCs are successfully induced from HCT116 colon cancer cell line. Xiaotan Sanjie Recipe can induce CCSCs in a dose-dependent manner, the mechanism may be related to the increased expression of pro-apoptotic protein Bax and decreased expression of anti-apoptotic protein Bcl-2.

关 键 词：消痰散结方 结肠癌干细胞样细胞 凋亡 BAX蛋白 BCL-2蛋白 

分 类 号：R735.35[医药卫生—肿瘤]