葛根素对肝癌细胞增殖、凋亡及对铂类敏感性的影响  

Effects of puerarin on proliferation,apoptosis and sensitivity to platinum in hepatocarcinoma cells

作　　者：胡亚丽[1] 范玉宏[1] 杨杰[1] 庞茜茜 HU Yali;FAN Yuhong;YANG Jie;PANG Qianqian(Pharmacy Department,the First Affiliated Hospital of Hebei North University,Zhangjiakou 075000,China)

机构地区：[1]河北北方学院附属第一医院药学部,河北张家口075000

出　　处：《山东医药》2021年第22期27-31,共5页Shandong Medical Journal

基　　金：河北省卫计委2020医学科学重点课题(20200563)。

摘　　要：目的探讨葛根素对肝癌细胞增殖、凋亡及对铂类敏感性的影响。方法将对数生长期人肝癌HepG2细胞随机分为6组:空白对照组、葛根素10μg/mL组、葛根素20μg/mL组和葛根素40μg/mL组、顺铂(40μg/mL)组和联合(葛根素20μg/mL+顺铂20μg/mL)组。将各组细胞继续培养24 h后,将葛根素针无菌剂溶于无菌蒸馏水中配制成浓度为80μg/mL的葛根素母液,并用无菌蒸馏水稀释为10、20、40μg/mL的稀释液,在层流仓的超净台中取无菌顺铂溶于无菌蒸馏水中,配制浓度为40μg/mL的顺铂母液,并稀释为40、20μg/mL溶液。于每孔中加入不同实验浓度药物300μL,使最终药物浓度为葛根素组分别为10、20和40μg/mL,顺铂40μg/mL,葛根素20μg/mL+顺铂20μg/mL。采用MTT法测定细胞增殖抑制率,采用流式细胞术检测细胞周期,观察细胞凋亡并计算细胞凋亡率,采用Westernblotting技术检测BCL2-AssociatedX的蛋白质(Bax)、B细胞淋巴瘤/白血病-2(Bcl-2)、激活型半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)蛋白表达。结果联合组增殖抑制率最高,其次为葛根素40μg/mL组;葛根素40μg/mL组凋亡率最高,其次为联合组,差异有统计学意义(P均<0.05);顺铂40μg/mL组、葛根素20μg/mL组、葛根素40μg/mL组和联合组G0/G1期所占比例均高于空白对照组;空白对照组S期所占比例最高,其次为葛根素10μg/mL组;空白对照组G2/M期所占比例最高,其次为葛根素10μg/mL组;联合组G0/G1期所占比例最高,S期和G2/M期所占比例最低,差异有统计学意义(P均<0.05)。其余5组的Bax、Bax/Bcl-2和激活型Caspase-3蛋白表达均高于空白对照组,空白对照组Bcl-2蛋白表达最高,其次为葛根素10μg/mL组;葛根素40μg/mL组Bax/Bcl-2和激活型Caspase-3蛋白表达最高,其次为联合组,差异有统计学意义(P均<0.05)。结论40μg/mL葛根素能提高肝癌细胞的增殖抑制率和凋亡率,与顺铂联用临床效果更佳。此外,葛根素具有提高人肝癌细Objective To investigate the effects of puerarin on the proliferation,apoptosis and sensitivity to plati‐num in hepatocarcinoma cells.Methods Liver cancer HepG2 cells in the logarithmic growth phase were divided into 6 groups:the blank control group,puerarin groups(10μg/mL group,20μg/mL group,and 40μg/mL group),40μg/mL cisplatin group,and combination(puerarin 20μg/mL+cisplatin 20μg/mL)group.After the cells in each group were cultured for 24 h,we took the puerarin injection sterile and dissolved it in sterile distilled water to prepare the solution into a puerarin mother liquor with a concentration of 80μg/mL,and then diluted it with sterile distilled water to 10μg/mL,20μg/mL and 40μg/mL diluent,after that,we took sterile cisplatin in the ultra-clean table of the laminar flow chamber and dissolved it in sterile distilled water to prepare the solution as a cisplatin mother solution with a concentration of 40μg/mL,and then diluted to 40μg/mL and 20μg/mL solutions.We added 300μL of experimental drugs of different groups to each well,so that the final drug concentration was 10μg/mL,20μg/mL and 40μg/mL in the puerarin groups,was 40μg/mL in the cisplatin group,and 20μg/mL+20μg/mL in the puerarin+cisplatin group.Cell proliferation inhibi‐tion rate was determined by MTT colorimetric assay.Cell cycle was detected by flow cytometry.Apoptosis was observed and apoptosis rate was calculated.Western blotting was used to detect the expression of Bcl-2 associated X protein(Bax),B-cellymphoma-2(Bcl-2),and activated Caspase-3 protein.Results The combination group had the highest prolifera‐tion inhibition rate,followed by the 40μg/mL puerarin group;the 40μg/mL puerarin group had the highest apoptosis rate,followed by the combination group,and the difference was statistically significant(P<0.05).The percentages of cells in the G0/G1 phase of the 40μg/mL cisplatin group,the 20μg/mL puerarin group,the 40μg/mL puerarin group and the combination group were higher than that of the blank control group;the bla

关 键 词：肝癌 葛根素 半胱氨酸天冬氨酸蛋白酶-3 细胞增殖 细胞凋亡 细胞周期 顺铂 

分 类 号：R735.7[医药卫生—肿瘤]