Myocardial ischemia‒reperfusion injury is a severe cardiovascular disease, and its treatment and prevention are crucial for improving patient prognosis and reducing the economic burden. This study aimed to explore the impact of hydrogen (H 2 ) on hypoxia/reoxygenation (H/R) injury in H9c2 cells (derived from rat embryonic heart tissue) induced by hydrogen peroxide (H 2 O 2 ) and to elucidate its underlying mechanism. An H/R injury model was established in H9c2 cells via exposure to 15 μM H 2 O 2 for 3 hours, followed by incubation in a 5% CO 2 atmosphere at 37°C for 24 hours. Then, the cells were treated with H 2 (50%) for 6, 12 or 24 hours. The results demonstrated that H9c2 cells exposed to H 2 O 2 and subjected to H/R injury presented a marked decrease in the cell survival rate, accompanied by severe morphological alterations, such as curling and wrinkling, and elevated lactate dehydrogenase levels. Notably, H 2 mitigated H/R injury induced by H 2 O 2 in a time-dependent manner, improving the morphological damage observed in H9c2 cells and decreasing lactate dehydrogenase levels. Compared with the model group, treatment with H 2 increased the activities of antioxidant enzymes, including catalase, superoxide dismutase, and glutathione peroxidase, while concurrently reducing the level of malondialdehyde, an indicator of cellular damage. Furthermore, H 2 treatment downregulated the expression of inflammatory cytokines and inflammatory-related factors, specifically interleukin-6, high-mobility group box 1, tumor necrosis factor-alpha, and Toll-like receptor 4, in H9c2 cells post-H/R injury. Furthermore, H 2 treatment resulted in a marked decrease in the expression levels of proteins associated with the Wnt/C-X3-C-motif receptor 1 signaling pathway, such as β-catenin, glycogen synthase kinase-3 beta, adenomatous polyposis coli, and Wnt and C-X3-C-motif receptor 1. This observation suggests a potential mechanism for its protective effects against H/R injury. Therefore, H 2 exerts a protective effect against H/R injury in H9c2 cells induced by H 2 O 2 , potentially by inhibiting the activated Wnt/C-X3-C-motif receptor 1 signaling pathway. This inhibition, in turn, prevents the generation of oxidative stress, inflammatory cytokines, and inflammation-associated factors.