Adenovirus-mediated SIRT1 protects cochlear strial marginal cells in a D-gal-induced senescent model in vitro.

Abstract

A primary obstacle in age-related hearing loss (ARHL) study is the lack of accelerated senescent models in vitro that explore the precise underlying mechanism in different types of ARHL. The damage to strial marginal cells (SMCs) is a subset of strial presbycusis-associated pathological changes. We aimed to establish a D-galactose (D-gal)-induced SMCs senescent model and study the effect of deacetylase sirtuin 1 (SIRT1) on presbycusis in vitro. SMCs from C57BL/6J neonatal mice were cultured and treated with D-gal to establish accelerated senescent models. And then D-gal-induced SMCs were transfected with adenovirus (Ad)-SIRT1-GFP or Ad-GFP. Oxidative stress and mitochondrial DNA (mtDNA) damage were determined by histological analysis or RT-PCR. Western blotting (WB) and RT-PCR were used to evaluate protein and mRNA levels of superoxide dismutase 2 (SOD2) and SIRT1, respectively. Additionally, apoptosis was investigated by WB and TUNEL staining. D-gal-induced SMCs exhibited several characteristics of senescence, including increased the level of 8-hydroxy-2'-deoxyguanosine, which is a marker of DNA oxidative damage, and elevated the amount of mtDNA 3860-bp deletion, which is a common type of mtDNA damage in the auditory system of mice. SIRT1 overexpression effectively inhibited these changes by upregulating the level of SOD2, thereby inhibiting cytochrome c translocation from mitochondria to cytoplasm, inhibiting cell apoptosis, and ultimately delaying aging in the D-gal-induced senescent SMCs. Altogether, the evidence suggests that the D-gal-induced SMCs accelerated aging model is successfully established, and SIRT1 overexpression protects SMCs against oxidative stress by enhancing SOD2 expression in ARHL.