Metformin's Effect on Non-Alcoholic Fatty Liver Disease through the miR-200a-5p and AMPK/SERCA2b Pathway

Objectives : Metformin, a primary treatment for type 2 diabetes due to its insulin-sensitizing capabilities, has been widely recognized for its efficacy in improving or mitigating non-alcoholic fatty liver disease (NAFLD). Recent investigations have highlighted the potential involvement of miRNAs in the pathogenesis of NAFLD, yet the precise mechanisms remain elusive. This study aims to explore the role of miR-200a-5p in regulating AMPK/SERCA2b to diminish liver fat accumulation and endoplasmic reticulum (ER) stress in metformin-treated NAFLD, thereby shedding light on novel therapeutic strategies. Methods : The PA cell model, induced by mixing oleic and palmitic acid solutions in a 2:1 ratio, was utilized to simulate lipid accumulation, which was assessed using oil red O and Nile red staining. RT-PCR measured the mRNA levels of miR-200a-5p and key lipid metabolism genes (Acc1, Fasn, Cpt1, Scd1, Cd36, and Serbp1). Western blotting determined the protein expression of AMPK, phosphorylated AMPK (p-AMPK) and SERCA2b. The interaction between miR-200a-5p and AMPK was investigated via a luciferase reporter assay, and the effect of miR-200a-5p overexpression on AMPK and SERCA2b proteins was also assessed. A high-fat diet-induced NAFLD mouse model was developed to evaluate the impact of metformin on liver steatosis and lipid deposition through HE and oil red O staining. Additionally, liver lipid profiles (TG, TC, LDL-C, and HDL-C) were quantified, and RT-PCR analyzed the mRNA levels of miR-200a-5p and lipid metabolism genes. The expression of AMPK, p-AMPK, and SERCA2b proteins were examined, alongside the mRNA levels of miR-200a-5p in the serum of NAFLD patients. Results : Elevated mRNA levels of miR-200a-5p and lipid metabolism genes were observed in the PA cell model, while AMPK and SERCA2b had lower protein expression levels. Luciferase reporter assays confirmed miR-200a-5p's interaction with AMPK. The miR-200a-5p mimic group exhibited decreased AMPK and SERCA2b expression levels. Animal studies demonstrated metformin's capacity to lessen liver steatosis and lipid deposition, improving lipid profiles (TG, TC, LDL-C, and HDL-C). Metformin treatment adjusted the mRNA levels of miR-200a-5p and lipid metabolism genes back to normal, as well as the protein expression levels of AMPK, p-AMPK, and SERCA2b. Elevated miR-200a-5p mRNA levels were detected in the serum of NAFLD patients. Conclusion : Our findings indicate that metformin may alleviate lipid deposition and ER stress in the livers of NAFLD mice by modulating the AMPK/SERCA2b pathway through miR-200a-5p. This study introduces a novel perspective on the mechanism of metformin's therapeutic effects in NAFLD treatment.