USE OF ORGANOTYPIC CULTURES OF CORTIS ORGAN TO STUDY THE PROTECTIVE EFFECTS OF ANTIOXIDANT MOLECULES ON CISPLATIN-INDUCED DAMAGE OF AUDITORY HAIR-CELLS

Hypothesis: Cisplatin causes the generation of reactive oxygen species (ROS), which interferes with the antioxidant defense system of Corti' s organ and results in damage to the hair cells. Background: Cisplatin is a widely used chemotherapeutic agent with the dose-limiting side e ffect of ototoxicity. Evidence is accumulating that cisplatin interfer es with the antioxidant defense system of Corti's organ. Methods: Orga notypic explants of P-3 rat organ of Corti were the in vitro model sys tem. Presence of intact auditory hair cells and stereocilia bundle int egrity was assayed by phalloidin-FITC staining. Fluorescent dye probes detected H2O2 and intracellular thiol [e.g., glutathione (GSH)]. Spec trophotometric analysis determined antioxidant enzyme levels. Results: There was a rapid dose-dependent cisplatin cytotoxicity in the explan ts after 48 h of exposure. An accumulation of H2O2 and a reduction of GSH levels were observed within cisplatin-exposed hair cells. L-buthio nine sulfoximine, an inhibitor of GSH formation, enhanced cisplatin ot otoxicity, whereas N-6-(2-phenylisopropyl) adenosine, an adenosine ago nist, elevated antioxidant enzyme levels and ameliorated cisplatin tox icity. The following molecules protected hair cells from cisplatin-ind uced damage: GSH; glutathione diethyl ester (GSHe); ebselen (EBS); 4-m ethylthiobenzoic acid (MTBA); and D-methionine (D-MET). EBS, MTBA, and D-MET in vitro protection correlates with in vivo protection in rats. Conclusions: Organotypic culture of Corti's organ has been validated as a model for studying cisplatin toxicity and for screening otoprotec tive molecules. Some of the events that contribute to cisplatin's abil ity to damage auditory hair cells are generation of ROS (e.g., H2O2), depletion of intracellular GSH, and interference with antioxidant enzy mes within the cochlea. Agents that bolster the cochlea's antioxidant system can prevent cisplatin destruction of auditory hair cells. Ident ified protective agents may prove to be clinically useful in limiting or completely protecting from cisplatin ototoxicity.