Objective: To investigate the effects of geniposide (Geni) on hearing loss and Nrf2/HO-1 signaling pathway in rats with acute otitis media (AOM). Methods: AOM rat model was constructed and randomly separated into Control group, AOM group, low, medium, and high dose Geni groups (Geni-L, Geni-M, Geni-H groups), and high-dose Geni+pathway inhibitor ML385 group (Geni-H+ML385 group). Rats in each group were subjected to hearing tests. ELISA was applied to detect serum inflammatory factors and oxidative stress level. HE staining was applied to observe pathological damage of ear mucosal tissue. Immunoblotting was applied to detect the expression of Nrf2/HO-1 signaling pathway related proteins. Results: Compared with the Control group, the middle ear mucosal tissue structure in the AOM group was disordered, with obvious swelling and shedding of ear mucosal cells, significant infiltration of inflammatory cells and proliferation of fibrous tissue, and significant thickening of mucosal thickness, the ABR response threshold, levels of IL-1β, IL-6, TNF-α, MDA, and expression of KEAP1 were elevated, the activities of SOD and GSH, and the expression of Nrf2 and HO-1 were reduced (P<0.05). The pathological symptoms of the middle ear mucosal tissue in the Geni-L, Geni-M, and Geni-H groups were reduced compared to the AOM group, and the degree of mucosal swelling and inflammatory cell infiltration were reduced, the mucosal thickness became thinner, the ABR response threshold, levels of IL-1β, IL-6, TNF-α, MDA, and expression of KEAP1 were reduced, the activities of SOD and GSH, and the expression of Nrf2 and HO-1 were increased, with the most obvious changes observed in the Geni-H group (P<0.05). The pathological symptoms of the middle ear mucosal tissue in the Geni-H+ML385 group were aggravated compared to the Geni-H group, with obvious swelling and shedding of ear mucosal cells, increased inflammatory cell infiltration and fibrous tissue proliferation, and increased mucosal thickness, the ABR response threshold, levels of IL-1β, IL-6, TNF-α, MDA, and expression of KEAP1 were elevated, the activities of SOD and GSH, and the expression of Nrf2 and HO-1 were reduced (P<0.05). Conclusion: Geniposide can improve hearing loss in AOM rats, and its mechanism of action is related to the activation of the Nrf2/HO-1 signaling pathway.