预知子醇提物联合雷公藤红素对SMMC7721细胞增殖和凋亡的影响  

Effects of Ethanol Extract of Akebia Fructus Combined with Celastrol on Proliferation and Apoptosis of SMMC7721 Cells

作　　者：吴晓 卢文丽[1] 张夜航 方肇勤[1] WU Xiao;LU Wenli;ZHANG Yehang;FANG Zhaoqin(College of Basic Medical Science,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)

机构地区：[1]上海中医药大学基础医学院,上海201203

出　　处：《中国中医药信息杂志》2021年第3期69-75,共7页Chinese Journal of Information on Traditional Chinese Medicine

基　　金：国家自然科学基金青年基金(81503482);上海市进一步加快中医药事业发展三年行动计划2018年-2020年[ZY(2018-2020)-CCCX-2001-01];上海中医药大学中医基础理论学科能力提升项目(A1-Z20-302-0107)。

摘　　要：目的观察预知子醇提物联合雷公藤红素对SMMC7721细胞增殖和凋亡的影响,探讨其可能的作用机制。方法将不同浓度预知子醇提物和雷公藤红素分别作用于SMMC7721细胞72 h,MTT法获得细胞存活率、72 h IC50后,计算药物相互作用指数(CDI)。将SMMC7721细胞分为空白对照组、预知子醇提物组、雷公藤红素组和预知子醇提物+雷公藤红素组(联合组),光镜观察细胞形态,荧光显微镜观察细胞核形态,流式细胞术检测细胞凋亡率,Western blot检测线粒体凋亡途径、内质网应激途径、自噬相关和线粒体-内质网结构偶联(MAM)相关蛋白的表达。结果预知子醇提物和雷公藤红素72 h IC50剂量CDI值为0.45,存在显著协同效应。联合组细胞存活率明显降低(P<0.05),细胞形态发生改变,且部分细胞核呈碎块状,细胞凋亡率较其余3组显著升高(P<0.05)。与空白对照组比较,雷公藤红素组和联合组线粒体凋亡相关蛋白Caspase-3表达明显降低(P<0.05),联合组cleaved Caspase-3表达明显降低(P<0.05),预知子醇提物组内质网应激相关蛋白Grp78表达明显降低(P<0.05),雷公藤红素组和联合组Grp78蛋白表达明显升高(P<0.05);联合组Chop蛋白表达较其余3组均明显升高(P<0.05);与空白对照组比较,预知子醇提物组和联合组自噬相关蛋白p62、LC3BⅡ及LC3BⅡ/Ⅰ比值均显著升高(P<0.05),各给药组MAM标志物Mfn2表达均升高但差异无统计学意义(P>0.05)。结论预知子醇提物联合雷公藤红素可抑制SMMC7721细胞增殖、诱导细胞凋亡,预知子醇提物和雷公藤红素作用靶点不尽相同;二者的一致性效应机制方面,除Caspase-3和cleaved Caspase-3、Chop和Mfn2等分子外,尚需进一步扩大关注面。Objective To observe the effects of ethanol extract of Akebia Fructus and Celastrol on proliferation and apoptosis of SMMC7721 cells;To discuss its possible mechanism of action.Methods Cells were treated by different concentrations of ethanol extract of Akebia Fructus or Celastrol for 72 hours,and MTT method was used to get cell viability data and IC50 of 72 h.Coefficient of drug interaction(CDI)was calculated.SMMC7721 cells were divided into blank control group,ethanol extract of Akebia Fructus group,Celastrol group,and ethanol extract of Akebia Fructus+Celastrol group(combined group).Cell morphology was observed under light microscope and nuclear morphology observed by fluorescence microscope.Apoptosis rate was inspected by Flow cytometry.Western blot was used to detect the expression of mitochondrial apoptosis pathway,endoplasmic reticulum stress pathway,autophagy related proteins and mitochondria-associated endoplasmic reticulum membrance(MAM)related proteins.Results The CDI value of ethanol extract of Akebia Fructus and Celastrol was 0.45 at 72 h IC50,which showed a good synergistic effect.The cell survival rate of the combined group was significantly reduced(P<0.05);the cell morphology was changed;some nuclei were fragmented;the apoptosis rate was significantly higher than that of the other three groups(P<0.05).Compared with the blank control group,the expression of mitochondrial apoptosis-related protein Caspase-3 in the Celastrol group and the combined group was significantly reduced(P<0.05);the expression of cleaved Caspase-3 in the combined group was significantly reduced(P<0.05);the expression of the endoplasmic reticulum stress-related protein Grp78 in the ethanol extract of Akebia Fructus group was significantly reduced(P<0.05).The expression of Grp78 protein in the Celastrol group and the combined group significantly increased(P<0.05);the expression of Chop protein in the combined group was significantly higher than that of the other three groups(P<0.05).Compared with the blank control group,the exp

关 键 词：预知子 雷公藤红素 肝癌 增殖 凋亡 机制 

分 类 号：R285.5[医药卫生—中药学]