Metformin Plus Berberine Synergistically Reverse Fatty Liver via AMPK Pathway

Nonalcoholic fatty liver disease (NAFLD) affects hundreds of millions globally and spans a spectrum from benign steatosis to cirrhosis and hepatocellular carcinoma. Despite its prevalence and serious consequences, no approved pharmacological treatments exist, creating urgent demand for effective strategies. Both metformin—a widely used antidiabetic biguanide—and berberine—an isoquinoline alkaloid derived from Coptis chinensis—independently activate AMPK and improve metabolic parameters, but their potential synergy in NAFLD had not been fully characterized.

The study employed a dual experimental design. In vivo, C57BL/6 mice were fed a high-fat diet for 12 weeks and concurrently treated with Met, BBR, or their combination. In vitro, HepG2 human hepatocytes were exposed to oleic acid and palmitic acid (OA/PA) to simulate lipid overload, then treated with the same agents. Drug synergy was quantified using combination index (CI) analysis (CI < 1 confirming synergy), and an AMPK inhibitor was applied in cell experiments to verify mechanistic dependence on AMPK signaling.

Across both models, combination therapy outperformed monotherapy on virtually every measured endpoint. In mice, the Met+BBR group showed the greatest reductions in body weight gain, liver weight, hepatic-to-body weight ratio, and visceral fat mass. Fasting glucose, fasting insulin, and HOMA-IR were all significantly lowered. Serum lipids improved substantially: LDL-C, TG, and TC fell while HDL-C rose. Histological analysis (H&E and Oil Red O staining) confirmed reduced hepatic steatosis, inflammatory infiltration, and Kupffer cell activation, and the NAFLD Activity Score (NAS) dropped significantly only in the combination group. Serum liver enzymes ALT and AST, along with pro-inflammatory cytokines TNF-α, IL-1β, and IL-6, were markedly suppressed by combination treatment. Mechanistically, p-AMPK levels were highest in the combination group, while SREBP1 and FASN protein expression were most strongly downregulated. In HepG2 cells, these effects were partially reversed by AMPK inhibition, confirming that the pathway is necessary for the observed benefits.

These findings suggest that Met and BBR engage complementary or overlapping mechanisms within the AMPK–SREBP1–FASN axis, producing additive-to-synergistic suppression of de novo lipogenesis and inflammation. The combination represents a potentially accessible, low-cost therapeutic strategy given both drugs' established safety profiles and availability. The results provide a mechanistic rationale for clinical trials evaluating Met+BBR co-administration in NAFLD patients.