Chaihu Shugan powder influences nonalcoholic fatty liver disease in rats in remodeling microRNAome and decreasing fatty acid synthesis

Abstract

Ethnopharmacological relevance: Chaihu Shugan powder (CSP) plays an important role in the prevention and treatment of nonalcoholic fatty liver disease (NAFLD) through a variety of biological mechanisms. However, whether the mechanism involves microRNA (miRNA) regulation remains unknown.

Aim of the study: To investigate the effects of CSP on the miRNA expression profile of rats with NAFLD induced by high-fat diet (HFD), and to explore the mechanism of CSP in the treatment of NAFLD.

Methods: NAFLD rat models were established by an 8-week HFD. The therapeutic effects of CSP on NAFLD were evaluated by physiological, biochemical and pathological analysis and hepatic surface microcirculation perfusion test. MicroRNA sequencing was used to study the effect of CSP on the miRNA expression profile of NAFLD rats, and the target genes of differentially expressed (DE) miRNAs were predicted for further function enrichment analysis. Next, targets of CSP and NAFLD were collected by a network pharmacological approach, and Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analysis were performed for the common target genes of CSP, NAFLD and DE miRNAs, and the expression levels of key genes and proteins were verified by quantitative Real-time PCR and Western blot. Finally, a network among formula-herb-compound-miRNA-target-biological processes-disease was established to explained the complex regulation mechanism of CSP on NAFLD.

Results: The results showed that CSP significantly improved liver lipid accumulation, serum lipid and transaminase levels and liver surface microcirculation disturbance in HFD-induced NAFLD rats. The intervention of CSP reversed the high expression of 15 miRNAs in liver tissues induced by HFD, including miR-34a-5p, miR-146a-5p, miR-20b-5p and miR-142-3p. The results of pathway and functional enrichment analysis showed that, CSP might play an anti-NAFLD role via regulating DE miRNAs related to fatty acid metabolic process. Combined with the network pharmacological analysis, it was found that the DE miRNAs might affected the fatty acid biosynthesis pathway in the treatment of NAFLD by CSP. Molecular biology experiments have conformed the decreased the gene and protein levels of acetyl-CoA carboxylase alpha (ACACA), fatty acid synthase (FASN) and other fatty acid biosynthesis related enzymes on NAFLD rats after intervention of CSP.

Conclusions: CSP can significantly reduce hepatic lipid accumulation of NAFLD rat model induced by HFD, and its mechanism may be through the action of 15 miRNAs such as miR-34a-5p, miR-146a-5p, miR-20b-5p and miR-142-3p. Reduce the gene and protein expression levels of ACACA, FASN and other fatty acid biosynthesis related enzymes, thus reducing fatty acid biosynthesis. Based on an epigenetic perspective, this study explains the key anti-NAFLD mechanism of CSP via combination of microRNA sequencing and network pharmacological analysis, providing a new reference for the modernization of traditional Chinese medicine.

Keywords: Chaihu Shugan powder; Fatty acid biosynthesis; Network pharmacology; Nonalcoholic fatty liver disease; microRNA.