三物白散含药血清通过TGF-β/Smad通路抑制TGF-β_(1)诱导的人胃癌SGC-7901细胞上皮间质转化  

Sanwubai San Inhibits TGF-β/_(1) Induced Epithelial Mesenchymal Transition of Human Gastric Cancer SGC-7901 Cells Through TGF-β/Smad Pathway

作　　者：王莉兰 徐中驰 凌云[1] 冯慧 周西彬 周春祥[1,4] WANG Lilan;XU Zhongchi;LING Yun;FENG Hui;ZHOU Xibin;ZHOU Chunxiang(Chinese Medicine Fever Laboratory,School of Chinese Medicine and School of Integrated Chinese and Western Medicine,Nanjing University of Chinese Medicine,Nanjing 210023,China;The First School of Clinical Medicine of Nanjing University of Chinese Medicine,Nanjing 210023,China;Second Military Medical University of the Chinese People's Liberation Army,Shanghai 200433,China;Nanjing BenQ Medical Center,Nanjing 210023,China)

机构地区：[1]南京中医药大学中医学院·中西医结合学院,中医热病实验室,南京210023 [2]南京中医药大学第一临床医学院,南京210023 [3]中国人民解放军第二军医大学,上海200433 [4]南京明基医院,南京210023

出　　处：《中国实验方剂学杂志》2022年第21期66-73,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81774021,82074504);江苏高校优势学科建设工程(中西医结合)项目。

摘　　要：目的:基于转化生长因子-β/Smad(TGF-β/Smad)信号通路,体外研究三物白散含药血清对TGF-β_(1)诱导的人胃癌SGC-7901细胞上皮间质转化(EMT)的影响及其机制。方法:SPF级3月龄雄性SD大鼠28只,随机分为空白组及三物白散低、中、高剂量组,每组7只,三物白散低、中、高剂量组分别按0.031 25、0.062 5、0.125 g·kg^(-1)·d^(-1)的剂量灌胃,空白组以等体积超纯水灌胃。每日1次,连续7 d。末次给药后45 min经腹主动脉取含药血清;细胞增殖与活性检测(CCK-8)法检测三物白散高剂量组含药血清对SGC-7901细胞活性的影响;镜下观察经TGF-β_(1)和三物白散含药血清处理后的细胞形态变化;细胞划痕实验和Transwell实验分别检测经TGF-β_(1)诱导和三物白散低、中、高剂量组含药血清处理后SGC-7901细胞的迁移率和侵袭率;蛋白免疫印迹法(Western blot)检测细胞E-钙黏蛋白(E-cadherin)、Snail、TGF-β_(1)、Smad3、磷酸化(p)-Smad3、Smad7蛋白的表达。结果:与空白组比较,10%、15%、20%三物白散高剂量组含药血清可呈浓度-时间依赖性抑制SGC-7901细胞活性;与空白组比较,模型组细胞失去梭形形态,多数细胞变圆、变长;与模型组比较,各药物组细胞伪足减少,细胞变小且形态恢复正常;与空白组比较,模型组细胞迁移和侵袭能力增强(P<0.01);与模型组比较,三物白散中、高剂量组处理SGC-7901细胞24 h后能显著抑制其迁移能力(P<0.01);三物白散低、中、高剂量组处理SGC-7901细胞48 h后均能显著抑制其迁移能力(P<0.01);与模型组比较,三物白散低、中、高组剂量组均能显著抑制其侵袭能力(P<0.01);与空白组比较,模型组E-cadherin及Smad7蛋白的表达水平显著降低(P<0.01),Snail、p-Smad3、TGF-β_(1)蛋白的表达水平显著升高(P<0.01),Smad3总蛋白水平不变;与模型组比较,三物白散高剂量组E-cadherin蛋白显著增加(P<0.01)、三物白散中剂量组有明显上升(P<0.05);Smad7蛋白Objective:To study the effect of serum containing Sanwubai San on TGF-β_(1) induced epithelial mesenchymal transition(EMT)of human gastric cancer SGC-7901 cells and its mechanism in vitro based on transforming growth factor-β/Smad(TGF-β/Smad)signaling pathway.Method:Twenty-eight male SDrats(SPF grade,three months)were randomly divided into blank group and Sanwubai low(0.031 25 g·kg^(-1)·d^(-1),ig),medium(0.062 5 g·kg^(-1)·d^(-1),ig)and high(0.125 g·kg^(-1)·d^(-1),ig)dose groups,seven in each group.The blank group was given the same volume of ultrapure water(ig).The gavage was performed once a day for seven consecutive days.The serum containing the drug was taken from the abdominal aorta 45 min after the last administration.Cell counting kit-8(CCK-8)method was used to detect the effect of serum in Sanwubai San high dose group on the activity of SGC-7901 cells.Changes of cell morphology after treatment with TGF-β_(1) and serum containing Sanwubai San were observed by microscopy,and the migration rate and invasion rate of the SGC-7901 cells were detected by cell scratch assay and transwell assay,respectively.Western blot was used to detect the expression of E-cadherin,snail,TGF-β_(1),Smad3,p-Smad3 and Smad7 proteins.Result:Compared with the blank group,10%,15%and 20%high-dose Sanwubai San inhibited the activity of SGC-7901 cells in a concentration and time dependent manner.Compared with the conditions in the blank group,the cells in the model group lost spindle shape,and most cells became round and long.Compared with the model group,the Sanwubai San groups had decreased pseudopodia and small cells with the morphology returning to normal.Compared with the conditions in the blank group,enhanced ability of cell migration and invasion(P<0.01),lowered expression of E-cadherin and Smad7(P<0.01),and increased expression of Snail,p-Smad3 and TGF-β_(1) (P<0.01)were found in the model group,with the total protein level of Smad3 remaining unchanged.Compared with the conditions in the model group,the cell migration

关 键 词：胃癌 上皮间质转化 三物白散 转化生长因子-β/Smad信号通路 中医药 

分 类 号：R22[医药卫生—中医基础理论] R242[医药卫生—中医学] R2-031R285.5