Deinococcus radiodurans is a robust bacterium that can withstand harsh environments that cause oxidative stress to macromolecules due to its cellular structure and physiological functions. In this study, we investigated the protective effects of extracellular vesicles derived from D. radiodurans (R1-EVs) against H₂O₂-induced oxidative stress in HaCaT cells. R1-EVs were isolated by ultracentrifugation with density gradient separation steps and characterized as spherical molecules with an average size of 322 nm by DLS, TEM and SEM. The antioxidative properties of R1-EVs were evaluated in H₂O₂-induced oxidative stress in HaCaT cells. Pretreatment with R1-EVs inhibited H₂O₂-induced apoptosis by preserving MMP and reducing ROS production. Additionally, R1-EVs increased the activities of antioxidant enzymes SOD and CAT, restored GSH homeostasis, and decreased MDA levels. The protective effect of R1-EVs against oxidative stress was mediated by downregulating MAPK phosphorylation and upregulating Nrf2/ARE pathway. Furthermore, the weak protective ability of R1-EVs from the SlpA protein-deletion ΔDR2577 mutant compared to wild-type R1-EVs confirmed our inferences that SlpA protein plays an important role in antioxidant effect. In conclusion, R1-EVs show significant protection against H₂O₂-induced oxidative stress in keratinocytes, suggesting its potential application in radiation-induced oxidative stress models.