miRNA-122在异烟肼肝损伤大鼠中的调控机制

Regulatory mechanism of miRNA-122 in isoniazid induced rat liver injuries

目的:探讨miRNA-122在异烟肼致大鼠肝损伤中的调控机制。方法:采用异烟肼55 mg·kg-1·d-1连续灌胃3,7,10,14,21,28 d建立肝损伤大鼠模型,对照组给予等容积纯化水灌胃。全自动生化分析仪检测血清丙氨酸氨基转移酶(ALT)、门冬氨酸转移酶(AST)活性;比色法检测肝组织中MDA含量和SOD酶活力,实时荧光定量PCR(RT-PCR)方法检测肝组织中miRNA-122、pri-miRNA-122、HNF4α、C/EBPα、Cycling G1、CAT-1表达水平;酶联免疫吸附法(ELISA)检测Cycling G1、CAT-1蛋白表达水平。结果:与对照组比较,肝组织病理变化在7 d表现为肝损伤;血浆ALT、AST的水平呈升高趋势(均P<0.01),且均在10 d发生明显上升(P<0.05);各用药组肝组织MDA含量逐渐升高,SOD活力却逐渐下降,miRNA-122表达水平整体呈下降趋势(P<0.01),其上游pri-miRNA-122及转录因子HNF4α、C/EBPαmRNA表达水平整体呈下降趋势(均P<0.01),其下游靶基因CyclingG1、CAT-1的mRNA及蛋白表达水平均呈上升趋势(均P<0.01)。结论:转录因子HNF-4α、C/EBPα调控miRNA-122低表达,miRNA-122低表达进一步调控其下游靶基因Cycling G1、CAT-1参与异烟肼致肝损伤的过程。

OBJECTIVE To explore regulatory mechanism of miRNA-122 in isoniazid induced rat liver injuries. METHODS Rat liver injury model was established by intragastric administration of isoniazid at 55 mg·kg-1 ·d- 1 for 3, 7, 10, 14, 21 and 28 days, respectively. Rats in control group were injected equal volume of distilled water. Serum alanine aminotransferase （ALT） and aspartate aminotransferase （AST） were measured by an automatic biochemical analyzer. MDA contents and SOD activities were detected by colorimetry method. Expression of miRNA-122, pri-miRNA-122, HNF4α, C/EBPα, Cycling G1 and CAT-1 were determined by real time fluorescent quantitative PCR （RT-PCR）. Protein contents of Cycling G1, CAT-1 were determined by enzyme-linked immunosorbant assay. RESULTS Compared with control group, pathology of liver tissues indicated that liver injury was observed at 7 days. Serum activities of ALT and AST showed a trend of increase and a sharp in- crease at 10 days （P〈0. 01）. Content of MDA was gradually increased and activity of SOD was decreased. Expression level of miRNA-122 was decreased （P〈0. 01）, expression levels of pri-miRNA-122 and transcription factor HNF4a, C/EBPa showed a trend of increase （P〈0. 01）. Expression levels of mRNA and protein of cycling G1 and CAT-1 showed an upward trend （P 〈0. 01）. （X）NCLUSION Transcription factors, HNF4a and C/EBPa can regulate low expression of miRNA-122 which further regulates its downstream target genes, cycling G1 and CAT-l, and takes part in process during which isoniazid induces rat liver injuries.