青藤碱对胃癌细胞SGC-7901生物学行为的影响及机制  

Effects of sinomenine on biological behaviors of gastric cancer cell SGC-7901 and its mechanism

作　　者：青泓屹 魏寿江[1] 李勋[1] 周程继[1] 张肖 祝晓娟 刘作良[1] Qing Hongyi;Wei Shoujiang;Li Xun(The First Dept of Gastroenterology,Affiliated Hospital of North Sichuan Medical College,Nanchong 637000)

机构地区：[1]川北医学院附属医院胃肠外一科,南充637000

出　　处：《安徽医科大学学报》2020年第9期1350-1356,共7页Acta Universitatis Medicinalis Anhui

基　　金：四川卫生和计划生育委员会科研课题项目(编号:17PJ104)。

摘　　要：目的探究青藤碱(SIN)对胃癌细胞SGC-7901增殖、凋亡、侵袭和迁移的影响及其机制。方法正常培养人胃正常黏膜上皮细胞GES-1、人胃癌细胞SGC-7901作对照组(Control),分别用0.5、1.0、2.5 mmol/L SIN处理细胞48 h,并设立阳性对照(50μmol/L塞来昔布),采用CCK-8法检测细胞生长,流式细胞仪检测细胞凋亡,Transwell小室检测细胞侵袭,划痕实验检测细胞迁移,Western blot检测Bax、Bcl-2、cleaved caspase-3、E-cadherin、N-cadherin和Vimentin的表达;经在线预测软件分析miR-33a-5p和乳酸脱氢酶A(LDHA)靶向关系,并通过双荧光素酶报告确认;将miR-33a-5p mimics转染SGC-7901细胞,采用RT-qPCR检测miR-33a-5p和LDHA mRNA的表达;重组构建过表达LDHA载体转染SGC-7901细胞,用0.5、1、2.5 mmol/L SIN处理细胞48 h,检测细胞增殖、凋亡、侵袭和迁移情况。结果与Control组相比,SIN可提高SGC-7901细胞凋亡率、Bax、cleaved caspase-3、E-cadherin及miR-33a-5p的表达,并降低其存活率、侵袭数、迁移率、Bcl-2、N-cadherin、Vimentin及LDHA mRNA的表达(P<0.05)。LDHA是miR-33a-5p的预测靶点。SIN可上调miR-33a-5p的表达,并下调LDHA mRNA的表达(P<0.05);与Contorl组相比,过表达LDHA后,细胞SGC-7901的凋亡率下降,其LDHA蛋白表达、存活率、侵袭数和迁移率上升(P<0.05);SIN处理后,细胞SGC-7901的凋亡率上升,其LDHA蛋白表达、存活率、侵袭数和迁移率下降(P<0.05);与pcDNA-LDHA组相比,干预SIN后,细胞SGC-7901的凋亡率上升,其存活率、侵袭数和迁移率下降(P<0.05)。结论SIN可靶向miR-33a-5p下调LDHA水平,介导细胞增殖、凋亡、侵袭和迁移等过程抑制胃癌细胞SGC-7901。Objective To explore the effects of sinomenine(SIN)on proliferation,apoptosis,invasion and migra-tion ability of gastric cancer cell line SGC-7901 and its mechanism.Methods Normal human gastric mucosal epi-thelial cells GES-1 and human gastric cancer cells SGC-7901 were normally cultured,and they were enrolled as blank control group(Control).They were treated with 0.5,1.0 and 2.5 mmol/L SIN for 48 h.The positive con-trol(50μmol/L celecoxib)was set up.Cell growth was detected by CCK-8 method.The apoptosis was detected by flow cytometry.Cell invasion was detected by Transwell chamber.Cell migration was detected by scratch test.Western blot was applied to detect the expression of Bax,Bcl-2,Cleaved caspase3,E-cadherin,N-cadherin and Vimentin.The targeted relationship between miR-33a-5p and lactate dehydrogenase A(LDHA)was analyzed by online prediction software,which was confirmed by double luciferase report.The miR-33a-5p mimics was transfect-ed into SGC-7901 cells.The expression of miR-33a-5p and LDHA mRNA in cells was detected by RT-qPCR.The LDHA overexpression vector was recombined and constructed to transfected into SGC-7901 cells.The cells were treated with 0.5,1.0 and 2.5 mmol/L SIN for 48 h.The cell proliferation,apoptosis,invasion and migration were detected.Results Compared with Control group,SIN could increase apoptosis rate of SGC-7901 cells,the expres-sion of Bax,Cleaved caspase-3,E-cadherin and miR-33a-5p,while decreased survival rate,invasion number,mi-gration rate,the expression of Bcl-2,N-cadherin,Vimentin and LDHA mRNA(P<0.05).LDHA was the pre-dicted target spot of miR-33a-5p.SIN could up-regulate the expression of miR-33a-5p,and down-regulate the ex-pression of LDHA mRNA(P<0.05).Compared with Control group,after LDHA overexpression,apoptosis rate of SGC-7901 decreased,while the expression of LDHA protein,survival rate,invasion number and migration rate in-creased(P<0.05).After SIN treatment,apoptosis rate of SGC-7901 increased,while the expression of LDHA protein,survival rate,invasion number and m

关 键 词：青藤碱 miR-33a-5p 乳酸脱氢酶A SGC-7901 

分 类 号：R965.1[医药卫生—药理学]