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  • miR‑148a‑3p regulates alcoholic liver fibrosis through targeting ERBB3.

miR‑148a‑3p regulates alcoholic liver fibrosis through targeting ERBB3.

International journal of molecular medicine (2020-06-26)
Jie Xiong, Jianbo Ni, Congying Chen, Kezhou Wang
ABSTRACT

Alcoholic liver disease greatly affects human health. Previous studies have identified that microRNAs (miRNAs) are associated with the pathogenesis of alcoholic liver fibrosis (ALF). Therefore, the present study explored the regulatory mechanism of miR‑148a‑3p in ALF. An ALF model was established in rats by alcohol gavage, followed by treatment with miR‑148a‑3p. Reverse transcription‑quantitative (RT‑q) PCR was performed to detect miR‑148a‑3p expression in the rat liver tissues. The levels of lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine transaminase (ALT) and alkaline phosphatase (ALP) were determined by enzyme‑labeled colorimetry. Liver damage was evaluated by liver indices and histology. The direct target gene of miR‑148a‑3p was predicted by a dual luciferase reporter assay. The effects of miR‑148a‑3p and miR‑148a‑3p in combination with receptor tyrosine‑protein kinase erbB‑3 (ERBB3) on HSC‑T6 cell viability and apoptosis were detected by MTT and flow cytometry assays, respectively. Western blotting and RT‑qPCR assays were performed to detect the expression levels of proteins and mRNA associated with fibrosis and apoptosis. The data showed that miR‑148a‑3p mimics inhibited the expression levels of AST, ALT, ALP, LDH, α‑SMA and type I collagen in the model, decreased the liver indices, and improved the liver damage caused by alcohol. ERBB3, which was predicted as the direct target gene of miR‑148a‑3p, reversed the effects of ERBB3 on promoting cell viability and inhibiting apoptosis. Concomitantly, miR‑148a‑3p reversed the increased expression of Bcl‑2 and inhibited the expression levels of Bax and c‑cleaved‑3 caused by ERBB3. These data suggested that miR‑148a‑3p regulated ALF and the viability and apoptosis of hepatic stellate cells through targeting ERBB3.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
SYBR® Green I nucleic acid gel stain, 10,000 × in DMSO
Sigma-Aldrich
Sphingomyelin Quantification Colorimetric Assay Kit, sufficient for 100 colorimetric tests
Sigma-Aldrich
ALT Activity Assay, sufficient for 100 colorimetric or fluorometric tests