Background/Objectives: Nobiletin, one of the main components of citrus peel, exhibits potent antioxidant, anti-inflammatory, and metabolic regulatory properties. However, its effect on obesity-associated vasculopathay remains unknown. We aim to investigate the effect of nobiletin in ameliorating oxidative stress and endothelial dysfunction induced by a high-fat diet (HFD). Methods: Male C57BL/6J mice were fed a HFD (60 kcal% fat) or normal chow for four months and orally administered with vehicle or nobiletin (50 mg/kg/day) for 8 weeks. Vasoreactivity in aortas was measured on a wire myograph. Primary rat aortic endothelial cells (RAECs) were isolated from Sprague-Dawley rats for in vitro study. Protein expressions were detected by Western blot. Superoxide production was determined by fluorescence imaging. Results: Exposure to high glucose increased the phosphorylation of JNK (Tyr185) and decreased the protein expressions of Nrf2 and HO-1, as well as downregulated the phosphorylation of AMPK and eNOS (Ser1177) in RAECs. This led to reduced nitric oxide (NO) generation and elevation of oxidative stress. High glucose induction also impaired the endothelium-dependent relaxations (EDRs) in murine aortas. These high glucose-induced impairments were restored by co-treatment of nobiletin (1 μM or 10 μM) whereas effects of nobiletin were abolished by AMPK inhibitor Compound C. The DIO-induced diabetic animal model showed increased body weight and blood pressure, imbalance of glucolipid metabolism, impaired EDRs, and elevated oxidative stress in aortas. AMPK/eNOS and Nrf2/HO-1 pathways were downregulated in aortas from DIO mice. Oral administration of nobiletin could at least partially reverse the above damage. Conclusions: Nobiletin ameliorates endothelial dysfunction by reducing oxidative stress and enhancing NO bioavailability upon activation of AMPK/eNOS and Nrf2/HO-1 pathways in obese diabetic mice.

Prolonged high‐fat diet causes insulin resistance and metabolic dysfunction‐associated steatotic liver disease (MASLD). 3,3′,4,5′‐Tetramethoxy‐trans‐stilbene (3,3′,4,5′‐TMS), a methoxy derivative of resveratrol, has been demonstrated to ameliorate oxidative stress and insulin resistance in vitro. However, it remains unclear whether 3,3′,4,5′‐TMS exhibits protective effects on insulin resistance‐related MASLD, which is investigated in the current study. Male C56BL/6 J mice were fed a high‐fat diet (60 kcal% fat) for 12 weeks to establish a model of diet‐induced obesity (DIO) with insulin resistance and associated MASLD. 3,3′,4,5′‐TMS (10 mg/kg/d) was intragastrically administered for 4 weeks. Hematoxylin and eosin, Oil‐red O, and periodic acid‐Schiff staining were performed to observe pathological changes of the liver. Protein markers for antioxidant and insulin signaling pathways were assessed by Western blot. 3,3′,4,5′‐TMS treatment ameliorated hyperglycemia, hyperlipidemia, and fatty liver in insulin‐resistant and MASLD mice. 3,3′,4,5′‐TMS regulated blood glucose through the activation of the insulin signaling pathway: increasing phosphorylation of IRS1 (Ser307), Akt (Ser473), and GSK3β (Ser9), upregulating protein expression of IRS2 and PI3K, and decreasing phosphorylation of GS (Ser641) in DIO mouse liver. Furthermore, 3,3′,4,5′‐TMS reduced oxidative stress in livers by upregulating Nrf2/HO‐1 signaling pathway. These findings indicate that 3,3′,4,5′‐TMS ameliorates insulin resistance and MASLD, possibly through the activation of IRS/PI3K/Akt/GSK3β/GS and Nrf2/HO‐1 pathways.