miR-199a-5p通过调节肠道菌群促进胆总管结扎诱导的大鼠肝纤维化  

miR-199a-5p promotes bile duct ligation-induced liver fibrosis of rats by regulating intestinal microbiota

作　　者：崔笑妍 于笑涵 周静 王璐瑶 张荣花 王梅梅 熊亚南 刘志勇 章广玲 Cui Xiaoyan;Yu Xiaohan;Zhou Jing;Wang Luyao;Zhang Ronghua;Wang Meimei;Xiong Yanan;Liu Zhiyong;Zhang Guangling(Hebei Provincial Key Laboratory for Chronic Diseases,School of Basic Medical Sciences,North China University of Science and Technology,Tangshan 063210;Hebei Key Laboratory of Integrated Precision Medicine,Clinical School of Medicine,North China University of Science and Technology,Tangshan 063000;Health Science Center of North China University of Science and Technology,Tangshan 063210)

机构地区：[1]华北理工大学基础医学院,河北省慢性疾病重点实验室,唐山063210 [2]华北理工大学临床医学院,河北省医工融合精准医疗重点实验室,唐山063000 [3]华北理工大学医学部,唐山063210

出　　处：《安徽医科大学学报》2024年第10期1794-1801,共8页Acta Universitatis Medicinalis Anhui

基　　金：河北省自然科学基金项目(编号:H2023209047、H2021209026);河北省教育厅在读研究生创新能力培养资助项目(编号:CXZZSS2024056);河北省引进留学人员项目(编号:C20210340);河北省重点研发计划项目(编号:213777115D);河北省政府资助临床医学优秀人才培养项目(编号:冀财预复[2020]397号)。

摘　　要：目的研究miR-199a-5p通过调节肠道菌群对胆总管结扎(BDL)诱导的大鼠肝纤维化的影响。方法25只SD大鼠随机分为Sham组、BDL组、空载腺病毒(NC adv)组、miR-199a-5p adv组和miR-199a-5p sponge adv组,每组5只。HE、Masson和Sirius Red染色观察大鼠肝组织病理变化及肝纤维化程度;全自动生化分析仪检测大鼠血清中天冬氨酸氨基转移酶(AST)、丙氨酸氨基转移酶(ALT)、总胆红素(TBIL)和直接胆红素(DBIL)含量;qRT-PCR检测大鼠肝组织中miR-199a-5p的表达水平;双重免疫荧光和Western blot实验检测大鼠肝组织中α-平滑肌肌动蛋白(α-SMA)和I型胶原α1链(COL1A1)的蛋白表达水平;收集大鼠粪便进行16S rRNA高通量测序。结果miR-199a-5p在BDL大鼠肝组织中表达上调(P<0.01)。与NC adv组相比,miR-199a-5p adv组大鼠肝损伤及胶原沉积现象更为严重,血清中AST、ALT、TBIL和DBIL含量以及肝组织中α-SMA和COL1A1表达水平显著升高(均P<0.05)。然而,miR-199a-5p sponge adv干预的结果与之相反(均P<0.05)。16S rRNA测序结果显示,miR-199a-5p adv干预可增加BDL大鼠肠道菌群的多样性和丰富度,改变肠道菌群组成,而miR-199a-5p sponge adv干预后菌群的结果与之相反(均P<0.05)。结论miR-199a-5p促进BDL大鼠肝纤维化,其机制可能与调节肠道菌群多样性和丰度相关。Objective To investigate the effect of miR-199a-5p on common bile duct ligation(BDL)-induced liver fibrosis in rats by regulating intestinal flora.Methods The 25 SD rats were randomly divided into five groups:the Sham group,the BDL group,the negative control adenovirus(NC adv)group,the miR-199a-5p adv group and the miR-199a-5p sponge adv group.The pathological changes of liver tissue and the degree of liver fibrosis were observed by HE,Masson and Sirius Red staining.The levels of aspartate aminotransferase(AST),alanine aminotransferase(ALT),total bilirubin(TBIL)and direct bilirubin(DBIL)in serum of rats were determined by a fully automatic biochemical analyzer.The mRNA expression level of miR-199a-5p in liver tissue of rats was detected by qRT-PCR.The protein expression levels ofα-smooth muscle actin(α-SMA)and collagen type 1 alpha 1(COL1A1)in liver tissue of rats were detected by double immunofluorescence staining and Western blot experiment.Rat feces were collected for 16S rRNA high-throughput sequencing.Results The expression of miR-199a-5p was up-regulated in the liver tissue of BDL rats(P<0.01).Compared with the NC adv group,the degree of liver injury and collagen deposition were relatively serious,the levels of AST,ALT,TBIL and DBIL in serum and the expression levels ofα-SMA and COL1A1 in liver tissue increased in the miR-199a-5p adv group(all P<0.05).However,the results of miR-199a-5p sponge adv intervention were opposite(all P<0.05).The 16S rRNA sequencing results showed that rats treated with miR-199a-5p adv were characterized by increased diversity and richness of intestinal microbiota,changed composition of intestinal microbiota,while the results of miR-199a-5p sponge adv interfering with the bacterial community were opposite(all P<0.05).Conclusion miR-199a-5p promotes liver fibrosis of BDL rats,and its mechanism may be related to regulating the diversity and abundance of intestinal microbiota.

关 键 词：miR-199a-5p 肠道菌群 胆总管结扎 肝纤维化 炎症 

分 类 号：R575.2[医药卫生—消化系统]