Aging-related hearing deficiency is the most prevalent sensory disturbance in the aging population and is associated with the mitochondrial (mt)DNA4977 deletion (corresponding to mtDNA4834 deletion in rats). Superoxide dismutase 2 (SOD2), a key factor involved in aging, is located in the mitochondrial matrix and is related to DNA mutation. The present study investigated the relationship between SOD2 and aging-related mtDNA4834 deletion in rats. SOD2 expression was evaluated in marginal cells (MCs) isolated from the inner ear of rats subjected to D-galactose-induced aging-related mtDNA4834 deletion. SOD2 was downregulated in mtDNA-deficient MCs, and the process was associated with methylation of the SOD2-encoding gene. In addition, overexpression of SOD2 in mtDNA4834-deleted MCs resulted in increased cell viability and reduced apoptosis, as demonstrated by the upregulation of anti-apoptotic proteins and the downregulation of pro-apoptotic proteins. SOD2 overexpression suppressed mtDNA4834 deletion mutation and increased the copy number of mtDNA4834, while SOD2 silencing resulted in the opposite effect. The findings indicate that aging-related mtDNA4834 deletion is associated with SOD2 deficiency, implicating that SOD2 is a potential candidate for the treatment of mtDNA4977 deletion-related presbycusis.