内质网应激-自噬肝癌SMMC-7721细胞模型的复制及金刚藤的干预  

Reproduction of an SMMC-7721 hepatocellular carcinoma cell model of endoplasmic reticulum stress induced autophagy:Impact on interventional effect of Smilax China L

作　　者：凌江红 文一惠 周芬敏 陈珺明 郭锦荣 Jing-Hong Ling;Yi-Hui Wen;Fen-Min Zhou;Jun-Ming Chen;Jin-Rong Guo(Department of Traditional Chinese Medicine,Zhoupu Hospital Affiliated to Shanghai University of Medicine&Health Sciences,Shanghai 200021,China;Department of Gastroenterology,Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai 200021,China;Department of Traditional Chinese Medicine,First Affiliated Hospital Guangxi Medical University,Nanning 530021,Guangxi Autonomous Region,China)

机构地区：[1]上海健康医学院附属周浦医院中医科,上海市200021 [2]上海中医药大学附属曙光医院消化科,上海市200021 [3]广西医科大学第一附属医院中医科,广西壮族自治区南宁市530021

出　　处：《世界华人消化杂志》2020年第20期1009-1015,共7页World Chinese Journal of Digestology

基　　金：上海市浦东新区卫生系统重点学科建设项目,No.PWZxk2017-02;国家自然科学基金资助项目,No.81973774.

摘　　要：背景内质网应激(endoplasmic reticulum stress,ERS)-自噬是肝癌细胞一种自我保护性机制.内质网(endoplasmic reticulum,ER)应激诱导剂二硫苏糖醇(dithiothreitol,DTT)能诱导肝癌细胞ERS-自噬.课题组前期研究证实金刚藤具有抑制肝癌细胞的作用,那么肝癌细胞ERS-自噬是否影响金刚藤抑制肝癌细胞的作用尚不清楚.目的探讨ERS-自噬肝癌SMMC-7721细胞模型的复制及金刚藤的干预.方法ERS-自噬肝癌SMMC-7721细胞模型的复制:采用不同剂量的DTT(50、200、500μmol/L)处理SMMC-7721细胞24 h,设DTT 0μmol/L组为control组,透射电镜观察各模型组细胞自噬小体,运用western blot法测定各组细胞微管相关蛋白1轻链3-Ⅱ(microtubuleassociated protein 1 light chain 3-Ⅱ,LC3-Ⅱ)蛋白的表达.金刚藤对ERS-自噬肝癌SMMC-7721细胞存活率的影响:将SMM C-7721细胞分为空白组(control组)、金刚藤组(JGT组)、DTT+JGT组,control、JGT组分别给予空白血清、金刚藤含药血清处理,DTT+JGT组予DTT构建的ERS模型后给予金刚藤含药血清处理,采用CCK-8法检测细胞的存活率.结果DTT(50、200、500μmol/L)组细胞的细胞质内均可见自噬小体,随着DTT浓度的增加,自噬小体数量逐渐增加,DTT 500μmol/L组可见大量自噬小体;各模型组细胞LC3-Ⅱ蛋白的表达:与control组比较,DTT(50、200、500μmol/L)组细胞中自噬相关蛋白LC3-Ⅱ的表达均显著升高(P均<0.05);与DTT 200μmol/L组比较,500μmol/L组LC3-Ⅱ蛋白的表达显著升高(P<0.05).金刚藤对ERS-自噬肝癌细胞模型存活率的影响:JGT、DTT+JGT组细胞存活率均显著低于control组(P均<0.05);JGT组细胞存活率显著低于DTT+JGT组(P<0.05).结论DTT可诱导肝癌SMMC-7721细胞ERS-自噬;金刚藤可抑制SMMC-7721细胞的存活率,DTT诱导的ERS-自噬可抵抗金刚藤抑制SMMC-7721细胞的存活率.BACKGROUND Endoplasmic reticulum stress induced autophagy is a self-protective mechanism of liver cancer cells.The endoplasmic reticulum stress inducer dithiothreitol(DTT)can induce autophagy in liver cancer cells.Previous studies have confirmed that Smilax China L has the effect of inhibiting liver cancer cells,but it is not clear whether the endoplasmic reticulum stress induced autophagy of liver cancer cells affects the role of Smilax China L in inhibiting liver cancer cells.AIM To reproduce a SMMC-7721 liver cancer cell model of endoplasmic reticulum stress induced autophagy and to explore the impact of endoplasmic reticulum stress induced autophagy on the interventional effect of Smilax China L.METHODS Different doses of dithiothreitol(DTT;0,50,200,and 500μmol/L)were used to treat SMMC-7721 cells for 24 h.Cells were then observed for autophagosomes by transmission electron microscopy.The expression of microtubule-associated protein 1 light chain 3-Ⅱ(LC3-Ⅱ)protein in each group was determined by Western blot.The effect of smilax China L on the survival rate of model SMMC-7721 cells was observed by divividing SMM C-7721 cells into the following groups:Control,smilax China L(JGT)alone,and DTT+JGT groups.The control and JGT groups were given blank serum and smilax China L containing serum,respectively.The DTT+JGT group was treated with DTT and smilax China L medicated serum.The CCK-8 method was used to detect the cell survival rate.RESULTS Autophagosomes can be seen in the cytoplasm of DTT(50,200,and 500μmol/L)treated cells.As the concentration of DTT increased,the number of autophagosomes gradually increased,with a large number of autophagosomes visible in the DTT 500μmol/L group.Compared with the control group,the expression of autophagy-related protein LC3-Ⅱin DTT(50,200,and 500μmol/L)treated cells was significantly increased(P<0.05 for all).Compared with the DTT 200μmol/L group,the expression of LC3-Ⅱprotein in the DTT 500μmol/L group was significantly increased(P<0.05).The cell survival rates

关 键 词：金刚藤 肝癌 内质网应激 自噬 细胞增殖 

分 类 号：R735.7[医药卫生—肿瘤]