1. Downregulation of microRNA-145 may contribute to liver fibrosis in biliary atresia by targeting ADD3
- Author
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Lei Liu, Zimin Chen, Zhihan Li, Dahao Zhang, Lihui Zhang, Qi Feng, Jian-yao Wang, Bin Wang, Xiaoshuo Ye, Zhouguang Wu, Yongqin Ye, and Wei Gao
- Subjects
0301 basic medicine ,Liver Cirrhosis ,Male ,Cirrhosis ,lcsh:Medicine ,Gene Expression ,Gastroenterology ,Biochemistry ,Liver disease ,Binding Analysis ,Gene expression ,Medicine and Health Sciences ,lcsh:Science ,Cells, Cultured ,Regulation of gene expression ,Multidisciplinary ,Liver Diseases ,Luciferase Assay ,Enzymes ,Nucleic acids ,Bioassays and Physiological Analysis ,Liver Fibrosis ,Female ,Oxidoreductases ,Luciferase ,Cell Binding Assay ,Research Article ,medicine.medical_specialty ,Down-Regulation ,Gastroenterology and Hepatology ,Biology ,Transfection ,Research and Analysis Methods ,03 medical and health sciences ,Extraction techniques ,Downregulation and upregulation ,Biliary atresia ,Biliary Atresia ,Internal medicine ,medicine ,Genetics ,Humans ,Molecular Biology Techniques ,Non-coding RNA ,Molecular Biology ,Chemical Characterization ,Enzyme Assays ,Retrospective Studies ,Three prime untranslated region ,lcsh:R ,Biology and Life Sciences ,Proteins ,Infant ,medicine.disease ,Molecular biology ,RNA extraction ,Gene regulation ,MicroRNAs ,030104 developmental biology ,Gene Expression Regulation ,Case-Control Studies ,Hepatic stellate cell ,Enzymology ,RNA ,lcsh:Q ,Calmodulin-Binding Proteins ,Biochemical Analysis - Abstract
Background and objectives Biliary atresia (BA) is a pediatric liver disease characterized by fibro-obliteration and obstruction of the extrahepatic biliary system, that invariably leads to cirrhosis and even death, if left untreated for extended time. However, its pathology and etiology still remained unknown. In this study, we tested the expression of adducin 3 (ADD3), the gene identified as a susceptibility gene in BA by GWAS, and uncovered its upstream regulatory microRNA in the pathogenesis of BA. Methods In this study, 14 infants with BA and 14 infants with choledochal cyst (CC) were enrolled as experimental group and control group, respectively. ADD3 and microRNA-145 (miR-145) expression profiles in liver tissues of BA and CC were determined using qPCR. Luciferase reporter assay was performed to verify the direct interaction between miR-145-5p and ADD3 3’ Untranslated Regions (3’UTR). The Lentiviral vectors containing miR-145, miR-145-3p inhibitor, miR-145-5p inhibitor, empty vector were transfected into human hepatic stellate cell line (LX-2) to determine the functional effect of miR-145 on ADD3 expression at both mRNA and protein level. Results MiR-145 was shown to be down-regulated in liver tissues of infants with BA compared to CC (p = 0.0267). ADD3, verified as a target of miR-145-5p, was shown to be overexpressed in infants with BA at the mRNA level (p = 0.0118). Transfection of lentiviruses containing miR-145 into LX-2 cells decreased the expression of ADD3 at both mRNA and protein level compared to negative control group, and suppressed the expression of p-Akt at protein level. Conclusions Our study has shown that overexpressed ADD3 and downregulated miR-145 were detected in BA liver tissues. MiR-145-5p was confirmed to target ADD3 by luciferase reporter assay. The downregulation of miR-145 may contribute to liver fibrosis in BA by upregulating the expression of ADD3.
- Published
- 2017