Nitration of cochlear proteins indicates oxidative posttranslational modification that can lead to catastrophic consequences, depending upon the functional activity of the nitrated proteins. We have reported evidence for cisplatin-induced nitration of two prominent cochlear bands. A functional role for these proteins is suggested by sterocilia and neuronal staining with anti-nitrotyrosine and by the ability of an otoprotective drug to block nitration. The long term aim of this research is to understand the role of protein nitration in ototoxicity and discover better interventional drugs that prevent ototoxicity.
The specific aims of this proposal are 1) to determine cisplatin-induced nitration of cochlear proteins and its correlation with hearing loss 2) to establish the identity the major nitrated cochlear protein(s) and 3) to manipulate the nitrated proteins using inhibitor and correlate with prevention of ototoxicity. Auditory brain stem response and distortion product otoacoustic emissions will be used to evaluate the hearing function, while cochleograms will be used to assess morphological damage. Quantification and localization of nitrated proteins in the cochlea will be done by immunoblotting and immunocytochemistry, respectively, with antibodies against nitrotyrosine. Nitrated proteins will be identified by gel electrophoresis followed by MALDI-TOF mass spectrometry. Proteomic responses associated with nitration will be investigated using antibody microarray and analyzed with pathway analysis software. The outcome of this study will reveal the role of cochlear protein nitration in cisplatin mediated ototxicity and will facilitate the identification of effective targets for therapeutic intervention.
Ototoxicity is one among the major dose-limiting factors of the widely used antineoplastic drug cisplatin. Nitration of proteins is considered as an important oxidative posttranslational modification that can lead to serious alterations in protein function. This study will reveal the correlation between cisplatin induced nitration of cochlear proteins and associated loss of hearing function. Identification of the cochlear proteins nitrated by cisplatin treatment and analysis of accompanying cochlear proteomic responses, with and without inhibition of nitration, will add more clarity to the molecular mechanism underlying cisplatin ototoxicity. These in turn will facililitate the identification and development of highly effective therapeutic targets to prevent cisplatin mediated ototoxicity.
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