Plant-derived extracellular vesicles improve endothelial viability and reduce cytotoxicity under oxidative stress

Objective: Endothelial dysfunction is a central feature of cardiovascular and metabolic diseases and is closely linked to oxidative stress–induced cellular injury. Excessive reactive oxygen species production disrupts endothelial viability, membrane integrity, and redox balance. Plant-derived extracellular vesicles (PD-EVs) have emerged as biocompatible nanovesicles with antioxidant potential; however, their effects on endothelial cells under oxidative stress remain insufficiently characterized. This study aimed to evaluate the protective effects of PD-EVs on endothelial cells exposed to hydrogen peroxide–induced oxidative stress. Methods: Extracellular vesicles were isolated from pomegranate, grape, and beetroot using differential ultracentrifugation and characterized by protein quantification, dynamic light scattering, and electron microscopy. Human umbilical vein endothelial cells (HUVECs) were exposed to hydrogen peroxide (H₂O₂) and subsequently treated with individual or combined PD-EVs. Cell viability and cytotoxicity were assessed using CCK-8 and LDH assays. Antioxidant responses were evaluated by quantitative real-time PCR analysis of GPX1 gene expression. Results: PD-EVs displayed nanoscale size and morphology consistent with extracellular vesicles. H₂O₂ exposure significantly reduced HUVEC viability and increased LDH release. PD-EV treatment significantly improved cell viability and reduced cytotoxicity under oxidative stress, with beetroot-derived and combined EV formulations showing the strongest effects. GPX1 expression was suppressed by H₂O₂ but significantly restored following PD-EV treatment, indicating activation of an adaptive antioxidant response. Conclusion: Plant-derived extracellular vesicles attenuate oxidative endothelial injury by improving cell viability, preserving membrane integrity, and restoring GPX1-associated antioxidant defenses. These findings highlight PD-EVs as a promising biocompatible, cell-free strategy for protecting endothelial cells against oxidative stress.