Abstract: Objective: this study aimed to investigate whether grape seed procyanidin extract (GSPE) can enhance autophagy through the nuclear factor erythroid 2-related factor 2(Nrf2)/heme oxygenase-1 (HO-1) signaling pathway and thereby inhibit glucolipotoxicity-induced ferroptosis in mouse islet β-cell lines (MIN6). Methods: MIN6 cells were treated with 25 mmol/L glucose and 200 μmol/L sodium palmitate (GP) for varying durations (0, 6, 12, 24, and 48 h) to establish an islet β-cell injury model. MIN6 cells were subsequently treated with the autophagy inducer rapamycin (RAPA, 0.1 μmol/L), the autophagy inhibitor chloroquine (CQ, 60 nmol/L), and different concentrations of GSPE (10, 20, and 30 mg/L). Nrf2 expression was silenced using small interfering RNA (siRNA). Cell viability was assessed using a cell counting Kit-8 (CCK-8). Mitochondrial membrane potential was evaluated with the JC-1 assay kit. Western blot (WB) was employed to analyze autophagy markers microtubule-associated protein 1 light chain 3 (LC3) and sequestosome-1 (P62), ferroptosis-associated key proteins acyl-CoA synthetase long-chain family member 4 (ACSL4) and glutathione peroxidase 4 (GPX4), and signaling proteins Nrf2 and HO-1. Results: Compared to the control group, GP treatment for 24 h significantly reduced cell viability and GPX4 expression, while increasing LC3II, P62, and ACSL4 expression (P<0.05). These findings suggest that GP induced ferroptosis and autophagic impairment in MIN6 cells. CQ-mediated autophagy inhibition further exacerbated ferroptosis, whereas RAPA-induced autophagy activation alleviated it (P<0.05). In addition, GSPE treatment enhanced mitochondrial membrane potential and upregulated GPX4, LC3II, Nrf2, and HO-1 expression, while downregulating P62 expression (P<0.05). Silencing of Nrf2 antagonized the effects of GSPE in enhancing autophagy and inhibiting ferroptosis (P<0.05). Conclusion: GSPE attenuates glucolipotoxicity-induced ferroptosis in pancreatic islets by enhancing autophagy through the activation of the Nrf2/HO-1 signaling pathway.