Tumor Necrosis Factor Alpha Ototoxicity Is Associated with Upregulation of Smac/Diablo Gene Expression in Rat Organ of Corti Explants

Objective: 1) Review the role of tumor necrosis factor alpha (TNFa) in programmed cell death of auditory hair cells (AHCs) in organ of Corti (OC) explants in vitro. 2) Understand effects of TNFa on Smac/Diablo gene expression in TNFa-induced apoptosis of OC explants in vitro. Method: OC explants were dissected from 3-day-old rat pups and cultured in (1) no treatment – control and (2) TNFa (2ug/mL) for 96 hours in vitro for hair cell count studies and 24 and 48 hours in vitro for gene expression studies for GAPDH and Smac/Diablo genes. Results: There are statistically significant decreases in the numbers of viable inner and outer hair cells in TNFa-treated OC explants, when compared with the control group. Gene expression of pro-apoptotic Smac/Diablo is significantly increased at 48 hours in vitro (but not at 24 hours), when compared with controls. On-going studies are evaluating the expression of Smac/Diablo gene at 72 and 96 hours in vitro. Conclusion: The role of Smac/Diablo gene expression has not previously been studied in TNFa-challenged OC explants. These results suggest late up regulation of Smac/Diablo expression in TNFa ototoxicity, which may be important for the propagation of the apoptosis pathway in OC explants in vitro. (NIDCD RO1-05575)