Astragalin from Morus alba L. ameliorates high-fat diet-induced obesity-related metabolic disturbances in mice and is associated with hepatic AMPK/PI3K/AKT signaling.

Morus alba L. (M. alba) also known as white mulberry is recognized in traditional Chinese medicine (TCM) and Ayurveda as a functional food and nutraceutical supplement for treating conditions such as hypertension, diabetes, and obesity. In TCM, mulberry leaves are specifically described as enhancing hepatic lipid metabolism, promoting lipid excretion to support obesity treatment. This study aimed to investigate the anti-obesity potential of M. alba leaf extract and its flavonoid component, astragalin (ASG), using a high-fat diet (HFD)-induced obese mouse model, and to elucidate the underlying molecular mechanisms. HFD-fed mice were treated with M. alba aqueous extract or ASG using a curative treatment approach. Body weight, glucose tolerance, insulin sensitivity, serum lipid profiles, hepatic steatosis, and inflammatory cytokines were assessed. Systemic and hepatic inflammation was measured by enzyme-linked immunosorbent assay (ELISA), while glucose transporter (GLUT1 and GLUT2) transcripts and protein levels were measured by RT-PCR and Western blotting, respectively. Molecular interactions were evaluated using cellular thermal shift assay (CETSA), drug affinity responsive target stability (DARTS), and molecular docking analyses, while AMP-activated protein kinase (AMPK), phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT), calcium/calmodulin-dependent protein kinase kinase 2 (CaMKK2) and liver kinase B1 (LKB1) phosphorylation was also assessed by Western blotting. Pharmacologic inhibition with Compound C was performed to confirm AMPK involvement. Treatment with M. alba extract significantly reduced body weight gain, improved serum lipid profiles, and attenuated hepatic lipid accumulation in obese mice. Phytochemical analysis identified ASG as a major bioactive constituent. ASG recapitulated several beneficial effects of the extract, including improvements in body weight gain, glucose tolerance, insulin sensitivity, hepatic lipid burden, and inflammatory status. ASG reduced systemic and hepatic levels of pro-inflammatory cytokines and modulated hepatic and intestinal GLUT1/2 transcripts and protein levels. Mechanistically, ASG showed evidence of AMPK engagement in CETSA, DARTS, and docking analyses, and was associated with increased hepatic AMPK and PI3K/AKT phosphorylation. Co-administration of Compound C attenuated several metabolic benefits of ASG, supporting AMPK involvement. ASG ameliorates obesity-related metabolic abnormalities in HFD-fed mice and is associated with AMPK engagement and increased hepatic AMPK/PI3K/AKT phosphorylation. These findings support ASG as a promising natural lead compound for obesity-related metabolic dysfunction, although further studies are needed to define its direct target interaction and downstream signaling more conclusively.