miR-199a在2,4,6-三硝基苯磺酸/乙醇溃疡性结肠炎大鼠中的表达及雷公藤多苷的作用研究  

Expression of miR-199a in Ulcerative Colitis Rats Induced by 2,4,6-Trinitrobenzene Sulfonic Acid/Ethanol and Study on the Effect of Tripteryginum wilfordii Polyglycoside

作　　者：钦丹萍[1,2] 杨新艳[2] 周毅骏 张春丽[3] 杨雪静[4] 李艳平[4] 孙佩娜[2] 代群[4] 

机构地区：[1]浙江中医药大学附属第一医院消化内科,杭州310006 [2]浙江中医药大学第一临床医学院,杭州310053 [3]浙江中医药大学附属第一医院病理科,杭州310006 [4]浙江中医药大学附属第一医院胃肠研究实验室,杭州310006

出　　处：《中国药学杂志》2016年第22期1934-1940,共7页Chinese Pharmaceutical Journal

基　　金：国家自然科学基金资助项目(81273903)

摘　　要：目的探讨miR-199a在2,4,6-三硝基苯磺酸(TNBS)/乙醇诱导的溃疡性结肠炎(UC)大鼠中的表达情况及雷公藤多苷(TWP)对其的作用。方法采用2,4,6-三硝基苯磺酸(TNBS)/乙醇灌肠法建立溃疡性结肠炎动物模型。对大鼠结肠组织进行大体形态损伤评分和镜下结肠损伤评分;采用芯片分析、Real-time PCR技术评估各组结肠黏膜组织中miR-199a的表达水平;针对milwalk数据库中预测的下游靶基因mRNA用表达谱进一步筛选分析靶基因mRNA,并用Real-time PCR技术验证靶基因mRNA在各组中的表达,最后在DAVID数据库进行pathway分析,以分析miR-199a参与UC炎症活动时所调节的靶基因mRNA。结果 TWP高剂量组结肠黏膜大体形态损伤评分、组织病理损伤评分与模型对照组间差异有统计学意义(P<0.01)。芯片分析显示,模型对照组中miR-199a较正常对照组上调(P<0.01),在AZA组表达较模型对照组下调(P<0.01,P<0.05)。miR-199a-3p在TWP中剂量组,miR-199a-5p在TWP高剂量组中较模型对照组均下调(P<0.05)。miR-199a的Real-time PCR结果显示,模型对照组miR-199a的表达水平较正常对照组明显上调(P<0.01);TWP的中、高剂量组和AZA组中miR-199a-3p的表达水平较模型对照组均下调(P<0.05);TWP中剂量组miR-199a-5p的表达水平较模型对照组下调(P<0.05)。表达谱分析显示,FASL为miR-199a的靶基因。模型对照组中靶基因FASL表达水平较正常对照组上调(P<0.05),TWP组、AZA组中靶基因FASL表达水平较模型对照组下调,其中以AZA组的下调为显著(P<0.01)。靶基因FASL的Real-time PCR结果显示,模型对照组中靶基因FASL表达水平较正常对照组明显上调(P<0.01);TWP中剂量组、高剂量组和AZA组中靶基因FASL表达水平较模型对照组下调(P<0.01)。结论 miR-199a在TNBS/乙醇UC大鼠中有上调表达,FASL是miR-199a的下游靶基因。TWP能改善UC的炎症活动,对UC活动时上调的miR-199a具有下调作用;FASL在UC活动时呈上调表达,TWP对OBJECTIVE To explore the expression of miR-199a in ulcerative colitis （UC） rats induced by 2,4,6-trinitrobenzene sulfonic （TNBS） / ethanol and the study on the effect of TWP on them. METHODS Through injecting TNBS/ethyl alcohol acid liq- uid into the anus of the rats to establish the UC rat model . The colitics eommom morphous damage and grade the histopathologieal score （CMDI） of colon mucosa injury were evaluated. Chip analysis and Real-time PCR were used to verify the expression of miR-199a in each colon mucosa tissue. Based on the expression profile, the downstream target genes mRNA in milwalk database was selected, then the expression of target genes mRNA by Real-time PCR in each group was veritied, at last the relevant signal pathway in the DA- VID database was analyzed. Doing these to analyse the target gene mRNA regulated by the miR-199a in the inflammatory activity of UC. RESULTS Compared with the model group, TWP high dose group was significantly lower on gross morphological damage score and histopathologieal injury score（ P 〈 0. 01 ）. Chip analysis showed that in model group, the expression of miR-199a was significantly higher than the normal group（ P 〈 0. 01 ） , and expression of the AZA group was significantly lower than the model group（ P 〈 0. 01 ,P 〈 0. 05 ）. The expression of miR-199a-3p in medium dose group and the expression of miR-199a-Sp in high dose group were significantlylower than the model group（ P 〈 0. 05 ）. The results of Real-time PCR showed that expression of miR-199a in the model group was sig- nificantly increased than that in the normal group（P 〈0. 01 ）. The expression of miR-199a-3p in TWP medium dose group, high dose group and AZA group were decreased than that in model group（P 〈 0. 05）. Meanwhile, the expression of miR-199a-5p in TWP medi- um dose group was decreased than that in model group（ P 〈 0. 05）. The gene expression profile showed that FASL was the target gene of miR-199a. In the model group, the expression of FASL

关 键 词：溃疡性结肠炎 雷公藤多苷 miR-199a 表达谱分析 FASL 

分 类 号：R965[医药卫生—药理学]