蟾毒灵调控PI3K/Akt通路对结肠癌HT-29细胞凋亡及周期的影响  

Effects of Bufalin Regulating PI3K/Akt Pathway on Apoptosis and Cell Cycle of Colon Cancer HT-29 Cells

作　　者：董倩倩[1] 张俊萍[2] DONG Qianqian;ZHANG Junping

机构地区：[1]周口市中心医院肛肠外科,河南周口466000 [2]河南省中医药研究院附属医院肿瘤科,河南郑州450004

出　　处：《新中医》2021年第23期149-154,共6页New Chinese Medicine

基　　金：河南省中医管理局青苗人才项目(豫中医科教[2018]16号)。

摘　　要：目的:研究蟾毒灵诱导人结肠癌HT-29细胞凋亡及周期阻滞的能力并对其分子机制进行探讨。方法:将人结肠癌HT-29细胞分为对照组(野生型细胞),蟾毒灵(Bufalin)处理组(0.25,0.5,1μmol/L)和顺铂(DDP)组(1μg/mL)。通过流式细胞术(FCM)检测蟾毒灵诱导人结肠癌HT-29细胞凋亡及周期阻滞的作用;使用蛋白质免疫印迹法检测经蟾毒灵处理HT-29细胞24 h后磷脂酰肌醇-3激酶(PI3K)、磷酸化磷脂酰肌醇-3激酶(p-PI3K)、蛋白激酶B(Akt)和磷酸化蛋白激酶B(p-Akt)蛋白表达的变化。通过实时荧光定量PCR法(RT-qPCR)检测蟾毒灵对HT-29细胞PI3K、Akt mRNA表达的影响。结果:FCM结果显示,与对照组比较,蟾毒灵(0.25,0.5,1μmol/L)组能诱导结肠癌HT-29细胞的凋亡(P<0.05),并呈剂量依赖性。与对照组比较,蟾毒灵(0.25,0.5,1μmol/L)组能够诱导结肠癌HT-29细胞的周期阻滞于G2/M期,并且蟾毒灵浓度越大,周期阻滞效果越显著(P<0.05)。蛋白质印迹实验结果表明,与对照组比较,蟾毒灵能够下调PI3K、p-PI3K、Akt和p-Akt的表达水平,并呈一定的剂量依赖(P<0.05)。RT-qPCR结果显示,与对照组比较,蟾毒灵能够显著下调PI3K和Akt mRNA的表达。结论:蟾毒灵能够诱导结肠癌HT-29细胞凋亡及周期阻滞,其机制可能与抑制PI3K/Akt信号通路有关。Objective:To study the ability of bufalin to induce apoptosis and cycle arrest of human colon cancer HT-29 cell,and to explore its molecular mechanism.Methods:Human colon cancer HT-29 cells was divided into control group(wild-type cells)and bufalin treatment group(0.25,0.5,1μmol/L)and cisplatin(DDP)group(1μg/mL).The apoptosis and cycle arrest of human colon cancer HT-29 cell induced by bufalin were detected by flow cytometry(FCM).Western blotting was used to detect the changes of protein expression of phosphatidylinositol-3 kinase(PI3K),phosphorylated phosphatidylinositol-3 kinase(p-PI3K),protein kinase B(Akt)and phosphorylated protein kinase B(p-Akt)in HT-29 cells treated with bufalin for 24 hours.The effect of bufalin on the mRNA expression of PI3K and Akt in HT-29 cells was detected by real-time fluorescence quantitative PCR(RT-qPCR).Results:The results of FCM showed that compared with the control group,bufalin(0.25,0.5,1μmol/L)group could induce apoptosis of colon cancer HT-29 cell in a dose-dependent manner(P<0.05).Compared with the control group,bufalin(0.25,0.5,1μmol/L)group could induce the cycle arrest of colon cancer cell line HT-29 in G2/M phase,and the greater the concentration of bufalin,the more significant the cycle arrest effect(P<0.05).Western blot results showed that compared with the control group,bufalin could down regulate the expression levels of PI3K,p-PI3K,Akt and p-Akt in a dose-dependent manner(P<0.05).RT-qPCR results showed that compared with the control group,bufalin could significantly down regulate the expression of PI3K and Akt mRNA.Conclusion:Bufalin can induce apoptosis and cycle arrest of colon cancer HT-29 cell,and its mechanism may be related to the inhibition of PI3K/Akt signal pathway.

关 键 词：蟾毒灵 结肠癌 凋亡 周期阻滞 

分 类 号：R285[医药卫生—中药学]