二烯丙基二硫上调RORα抑制人胃癌细胞Wnt/β-catenin通路靶基因  

Upregulation of RORα suppresses target genes of Wnt/β-catenin pathway in human gastric MGC803 cells by DADS

作　　者：苏坚[1,2] 赵晓红[2,3] 向姝霖[2] 刘芳[2] 夏红[2] 苏波[2] 凌晖[2] 曾希[2] 苏琦[2] 

机构地区：[1]南华大学附属第二医院病理科,湖南衡阳421001 [2]南华大学肿瘤研究所湖南省胃癌研究中心湖南省高校肿瘤细胞与分子病理学重点实验室,湖南衡阳421001 [3]海南省妇幼保健院,海南海口570206

出　　处：《中国药理学通报》2017年第4期522-528,共7页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81374013;81102854);湖南省卫计委科研课题(No B2015-182)

摘　　要：目的观察DADS上调RORα对人胃癌MGC803细胞Wnt/β-catenin通路相关分子与靶基因表达的影响。方法RT-PCR、Western blot与免疫共沉淀检测Wnt/β-catenin通路相关分子与靶基因表达。荧光素酶报告基因检测c-Myc基因启动子活性。结果 RT-PCR与Western blot显示,DADS处理后各组RORαmRNA与蛋白表达上调(P<0.05),而沉默RORα较对照组明显下调(P<0.05)。DADS处理后各组Wnt1 mRNA与蛋白表达下调(P<0.05)。沉默RORα较对照组Wnt1 mRNA与蛋白明显上调(P<0.05)。DADS处理前后各组β-catenin mRNA与蛋白表达差异无显著性(P>0.05)。免疫共沉淀显示,DADS处理组RORα与β-catenin结合明显增加,而沉默组与β-catenin结合明显减少(P<0.05)。DADS可明显下调核内β-catenin与p-β-catenin(P<0.05),而沉默RORα可明显上调β-catenin与p-β-catenin(P<0.05)。同时,DADS可明显下调TCF-4表达(P<0.05),而沉默RORα可明显上调TCF-4(P<0.05)。DADS可明显下调Axin、c-Myc、c-Jun mRNA与蛋白表达(P<0.05),而沉默RORα作用相反(P<0.05)。荧光素酶报告基因检测显示,DADS处理后,各组c-Myc启动子活性明显低于处理前(P<0.05)。沉默组c-Myc启动子活性明显高于对照组(P<0.05)。结论 DADS可上调RORα抑制Wnt/β-catenin通路相关分子与靶基因表达。Aim In ground of prophase work,to investigate the upregulation of RORα suppresses target genes of Wnt/β-catenin pathway in human gastric MGC803 cells by DADS.Methods The expressions of correlated molecules and target genes in Wnt/β-catenin pathway were tested by RT-PCR,Western blot and coimmunoprecipitation.Luciferase assay was used to assess the promoter activity of c-Myc.Results Soft agar colony formation showed that the proliferation in silence group of RORα was significantly strengthened compared with in control group,and the proliferation was notably decreased in all groups by DADS（ P 〈 0.05）.RT-PCR and Western blot showed that the expressions of mRNA and protein of RORα and Wnt1 were respectively downregulated and up-regulated in silence group,and the expressions of RORα and Wnt1 were respectively upregulated and downregulated in all groups by DADS（ P 〈 0.05）.The expressions of β-catenin mRNA and protein in all groups in circa treated by DADS had no significant deviation（ P 〉 0.05）.Coimmunoprecipitation exhibited that RORα bind to β-catenin had significant increase after treated by DADS,but RORα bind to β-catenin showed significant decrease in silence group（ P 〈 0.05）.The expressions of intranuclear β-catenin and p-β-catenin were markedly downregulated by DADS,however,the expressions were evidently up-regulated in silence group（ P 〈0.05）.At the same time,the expression of TCF-4 was downregulated by DADS,yet the expression of TCF-4was up-regulated in silence group（ P 〈 0.05）.RTPCR and Western blot showed that DADS decreased the expressions of mRNA and proteins of target genes of Wnt/β-catenin pathway,such as Axin,c-Myc and cJun,and inhibited the promoter activity of c-Myc by DADS.Conclusion The upregulation of RORα could suppress the expression of target genes of Wnt/β-catenin pathway,and then inhibit the proliferation in MGC803 cells by DADS.

关 键 词：二烯丙基二硫 人胃癌细胞 RORα Wnt/β-catenin信号通路 相关分子 靶基因 启动子活性 

分 类 号：R329.25[医药卫生—人体解剖和组织胚胎学] R392.11[医药卫生—基础医学]