Dexamethasone protects organ of corti explants against tumor necrosis factor-alpha–induced loss of auditory hair cells and alters the expression levels of apoptosis-related genes

Abstract

Objective: Determine the molecular mechanism(s) behind tumor necrosis factor-alpha (TNFα)–induced loss of auditory hair cells and the ability of dexamethasone base (DXMb) to protect against TNFα ototoxicity. Methods: Hair cell counts: Three-day-old rat organ of Corti explants were cultured under three different conditions: 1) untreated-control; 2) TNFα (2 μg/ml); and 3) TNFα (2 μg/ml)+DXMb (70 μg/ml) for 4 days, fixed, and stained with FITC–phalloidin. Hair cells were counted in the basal and middle turns. Gene expression: total RNA was extracted from the three different groups of explants at 0, 12, 24 and 48 h. Using quantitative real-time RT-PCR, mRNAs were transcribed into cDNAs and amplification was performed using primers for rat ß-actin (housekeeping gene), TNFR1, Bcl-2, Bax, and Bcl-xl. Results: DXMb protected explant hair cells from TNFα-induced loss. Bax gene expression was greater in TNFα-exposed explants compared with TNFα+DXMb-treated explants at 48 h ( P=0.023), confirmed by the increase in the Bax/Bcl-2 ratio at 48 h ( P<0.001). These results correlated with increased TNFR1 expression at 24 h ( P=0.038). DXMb otoprotection in TNFα-exposed cultures was accompanied by an up-regulation of Bcl-xl at both the 24 ( P<0.001) and 48 h time points ( P=0.030) and up-regulation of Bcl-2 expression at 24 h ( P=0.018). DXMb treatment also prevented increases in the expression levels of Bax, TNFR1, and the Bax/Bcl-2 ratio that occurred in untreated TNFα-exposed explants. Conclusions: TNFα's ototoxicity may be mediated through an up-regulation of Bax and TNFR1 expression as well as an increase in the Bax/Bcl-2 ratio. DXMb protects the organ of Corti against TNFα ototoxicity by up-regulating Bcl-2 and Bcl-xl expression and by inhibiting TNFα-induced increases in Bax, TNFR1, and the Bax/Bcl-2 ratio. These results support the use of local dexamethasone treatment to conserve hearing following a trauma.