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

Objective: Determine the molecular mechanism(s) behind tumor necrosis factor-alpha (TNF alpha)-induced loss of auditory hair cells and the ability of dexamethasone base (DXMb) to protect against TNF alpha ototoxicity. Methods: Hair cell counts: Three-day-old rat organ of Corti explants were cultured under three different conditions: 1) untreated-control; 2) TNF alpha (2 mu g/ml); and 3) TNF alpha (2 mu g/ml)+DXMb (70 mu g/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 beta-actin (housekeeping gene), TNFR1, Bcl-2, Bax, and Bcl-xl. Results: DXMb protected explant hair cells from TNF alpha-induced loss. Bax gene expression was greater in TNF alpha-exposed explants compared with TNF alpha+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 TNF alpha-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 TNF alpha-exposed explants. Conclusions: TNF alpha'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 TNF alpha ototoxicity by up-regulating Bcl-2 and Bcl-xl expression and by inhibiting TNF alpha-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. (C) 2008 IBRO. Published by Elsevier Ltd. All rights reserved.