miR-205-5p介导四君子汤对胃癌细胞迁移、侵袭及上皮间质转化的作用机制研究  

Study on the Mechanism of miR-205-5p-Mediated Sijunzi Decoction on Migration,Invasion and Epithelial-Mesenchymal Transition of Gastric Cancer Cells

作　　者：王纪鸾 赵琳琳 李乡南[1,2] 边月 马天驰 周晶[1,2] 郭隽馥 WANG Jiluan;ZHAO Linlin;LI Xiangnan;BIAN Yue;MA Tianchi;ZHOU Jing;GUO Junfu(Liaoning University of Traditional Chinese Medicine,Shenyang 110847 Liaoning,China;Shenyang Key Laboratory for TCM Emotional Disorder,Shenyang 110847 Liaoning,China;Shenyang Medical College,Shenyang 110034 Liaoning,China;Traditional Chinese Medicine Multidisciplinary Innovation Team of Liaoning Province,Shenyang 110001 Liaoning,China)

机构地区：[1]辽宁中医药大学,辽宁沈阳110847 [2]沈阳市中医神志病学重点实验室,辽宁沈阳110847 [3]沈阳医学院,辽宁沈阳110034 [4]辽宁省中医药多学科交叉创新团队,辽宁沈阳110001

出　　处：《中药新药与临床药理》2025年第3期317-327,共11页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：国家自然科学基金项目(81803855);辽宁省科技计划联合计划应用基础研究项目(2023JH2/101700205);辽宁省“兴辽英才计划”项目(XLYC2203180);辽宁省教育厅科技创新团队项目(LJ222410162020)。

摘　　要：目的探讨四君子汤能否通过上调miR-205-5p表达进而抑制胃癌细胞的迁移力、侵袭力以及上皮间质转化(EMT)进程,并筛查可能参与上述过程的miR-205-5p下游靶基因。方法四君子汤含药血清制备:SD大鼠以四君子汤(6.9 g·kg-1)灌胃,每日1次,共7 d。(1)将胃癌细胞MGC-803、AGS作为研究对象,分组及干预:生理盐水组(10%空白血清),四君子汤组(10%四君子汤含药血清),四君子汤+inhibitor NC组(10%四君子汤含药血清+转染抑制剂阴性对照),四君子汤+miR-205-5p inhibitor组(10%四君子汤含药血清+转染miR-205-5p抑制剂),各组血清处理均为24 h。采用qPCR或Western Blot法检测各组细胞中miR-205-5p表达水平,Snail、MMP9、TIMP1 mRNA及蛋白表达水平;划痕实验检测细胞迁移能力;Transwell实验检测细胞侵袭能力。(2)以人胃癌MGC-803细胞为研究对象,分组及干预:阴性对照组(转染阴性对照小干扰RNA),miR-205-5p模拟物组(转染miR-205-5p模拟物)。通过转录组测序分析miR-205-5p模拟物组vs阴性对照组的差异表达基因;采用qPCR技术对miR-205-5p过表达后下调的3个基因(ENC1、FGF7、CSF1)的表达水平进行验证;对差异表达基因进行GO功能及KEGG通路富集分析。结果(1)与生理盐水组比较,四君子汤组MGC-803、AGS细胞的miR-205-5p表达水平均显著升高(P<0.01)。与四君子汤+inhibitor NC组比较,四君子汤+miR-205-5p inhibitor组MGC-803、AGS细胞的miR-205-5p表达均显著下调(P<0.01)。(2)与生理盐水组比较,四君子汤组MGC-803、AGS细胞的愈合率明显降低(P<0.05),穿过基质胶的细胞数目显著减少(P<0.01),细胞MMP9、Snail mRNA及蛋白表达显著下调(P<0.05,P<0.01),TIMP1 mRNA及蛋白表达显著上调(P<0.05,P<0.01)。与四君子汤+inhibitor NC组比较,四君子汤+miR-205-5p inhibitor组MGC-803、AGS细胞的划痕愈合率均明显升高(P<0.05,P<0.01),穿过基质胶的细胞数目显著增多(P<0.01),细胞MMP9、Snail mRNA及蛋白表达显著上调(P<0.Objective To investigate whether Sijunzi Decoction(SJZT)inhibits the migration,invasion and epithelialmesenchymal transition(EMT)process of gastric cancer cells by up-regulating the expression of miR-205-5p,and to screen the downstream target genes of miR-205-5p that might be involved in the above process.Methods Preparation of serum containing SJZT:SD rats were given SJZT(6.9 g·kg-1)by gavage once a day for 7 days.(1)Gastric cancer cells MGC-803 and AGS were used as research objects.Grouping and intervention:normal saline group(10%blank serum),SJZT group(10%SJZT containing serum),SJZT+inhibitor NC group(10%SJZT containing serum+transfection inhibitor negative control),SJZT+miR-205-5p inhibitor group(10%SJZT containing serum+transfection miR-205-5p inhibitor).The serum treatment of each group was 24 hours.The mRNA and protein expression levels of miR-205-5p,Snail,MMP9 and TIMP1 in each group were detected by qPCR and Western Blot.Scratch assay was used to detect cell migration ability;transwell assay was used to detect cell invasion ability.(2)Human gastric cancer MGC-803 cells were used as the research object,grouping and intervention:negative control group(transfected with negative control small interfering RNA),miR-205-5p mimic group(transfected with miR-205-5p mimic).The differentially expressed genes of miR-205-5p mimic group versus negative control group were analyzed by transcriptome sequencing.The expression levels of three genes(ENC1,FGF7,CSF1)down-regulated by miR-205-5p overexpression were verified by qPCR.GO function and KEGG pathway enrichment analysis were performed on differentially expressed genes.Results(1)Compared with the normal saline group,the expression levels of miR-205-5p in MGC-803 and AGS cells in the SJZT group were significantly increased(P<0.01).Compared with SJZT+inhibitor NC group,the expression of miR-205-5p in MGC-803 and AGS cells in SJZT+miR-205-5p inhibitor group was significantly down-regulated(P<0.01).(2)Compared with the normal saline group,the healing rate of MGC-803 and A

关 键 词：胃癌 四君子汤含药血清 miR-205-5p MGC-803细胞 AGS细胞 迁移 侵袭 上皮间质转化 大鼠血清 

分 类 号：R285.5[医药卫生—中药学]