<i>Eucommia ulmoides</i><scp>O</scp>liv. leaves flavonoids attenuate methylglyoxal‐induced endothelial cell apoptosis in vitro and in vivo by upregulating <scp>AKT</scp>‐<scp>N</scp>rf2 signaling and downregulating oxidative stress

Methylglyoxal (MGO) triggers oxidative stress responses in vascular endothelial cells, leading to apoptosis linked to diabetic vascular complications. Total flavonoids of <i>Eucommia ulmoides</i> leaves (TFEL) display antioxidant activity, yet its prevention of MGO-induced apoptosis and mechanisms are unclear. Our study used western blotting and ELISA to evaluate protein levels and enzyme activities. Cell viability and apoptosis were evaluated using CCK8 assay and PE Annexin V/7-AAD double staining. Reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP) were measured using fluorescence probes. Vascular pathological changes and apoptosis were analyzed through H&E and TUNEL staining. In vitro, MGO-stimulated human umbilical vein endothelial cells (HUVECs) were treated with varying TFEL concentrations. Our results demonstrated that TFEL significantly enhanced cell viability, reduced apoptosis, downregulated caspase-3 activity, and Bax/Bcl-2 ratio. Moreover, TFEL markedly suppressed MGO-induced ROS and malondialdehyde (MDA) production while restoring antioxidant enzyme activity and MMP. TFEL pretreatment promoted the expression of p-Akt, Nrf2, and HO-1 proteins. Pharmacological inhibition of p-Akt significantly suppressed the upregulation of Nrf2 and HO-1 protein levels mediated by TFEL. Consistently, pharmacological inhibition of Nrf2 or p-Akt partially abrogated the protective effects of TFEL against MGO-induced damage in HUVECs. In vivo studies revealed that TFEL (100 and 200 mg/kg) partially restored antioxidant capacity and reduced aortic thickness and apoptosis in MGO-injured mice. In conclusion, the findings indicate that TFEL mitigates MGO-induced apoptosis via activation of p-Akt/Nrf2/HO-1 and scavenging of oxidative stress, highlighting its potential in diabetic vascular complication management.