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Stereotactic radiosurgery for lateral skull base tumors can cause hearing loss when the cochleae are exposed to high doses of single-fraction radiation. Currently, there are no known nondosimetric preventative treatments for radiation-induced ototoxicity.Intratympanic (IT) dexamethasone (DXM), a synthetic steroid, protects against radiation-induced auditory hair cell (HC) and hearing losses in rats in vivo.Seven rats received radiation (12 Gy) to both cochleae. In irradiated rats and six nonirradiated rats, IT DXM was randomized to one ear, while tympanic puncture without DXM was performed on the contralateral ear. Baseline and 4-week postradiation auditory-evoked potential tests were performed. The cochleae were processed for HC viability.Cochleae exposed to radiation demonstrated more outer HC (OHC) loss in all turns than nonirradiated ears (p 0.05). OHCs were more susceptible to radiation injury than inner HCs in the middle and basal turns (p 0.05). In irradiated cochleae, there was a nonsignificant trend for less OHC loss with IT DXM in the basal turn when compared with placebo. IT DXM did not improve radiation-induced hearing threshold shifts; however, a high rate of tympanic membrane perforations occurred with irradiated ears which may contribute to this finding.Radiation induced loss of OHCs in all turns of the cochlea. IT DXM reduced OHC loss in the basal turn of irradiated ears; however, this finding did not achieve statistical significance. Although IT DXM did not affect radiation-induced hearing threshold shifts in adult rats in vivo, this may be due to a high rate of tympanic membrane perforations.

Hypothesis: Dexamethasone (DXM) protects against radiation-induced loss of auditory hair cells (HCs) in rat organ of Corti (OC) explants by reducing levels of oxidative stress and apoptosis. Background: Radiation-induced sensorineural hearing loss (HL) is progressive, dose-dependent, and irreversible. Currently, there are no preventative therapeutic modalities for radiation-induced HL. DXM is a synthetic steroid that can potentially target many of the pathways involved in radiation-induced ototoxicity. Methods: Whole OC explants were dissected from 3-day-old rat cochleae exposed to specific dosages of single-fraction radiation (0, 2, 5, 10, or 20 Gy), were either untreated or treated with DXM (75, 150, 300 μg/mL), and then cultured for 48 or 96 hours. Confocal microscopy for oxidative stress (CellRox, 48 h) and apoptosis (TUNEL assay, 96 h) and fluorescent microscopy for viable HC counts (fluorescein isothiocyanate-phalloidin, 96 h) were performed. Analysis of variance and Tukey post hoc testing were used for statistical analysis. Results: Radiation exposure initiated dose-dependent losses of inner and outer HCs, predominantly in the basal turns of the OC explants. DXM protected against radiation-induced HC losses in a dose-dependent manner. DXM significantly reduced levels of oxidative stress and apoptosis in radiation-injured OC explants (p Conclusions: Radiation-initiated HC losses were dose-dependent in OC explants. DXM treatment protected explant HCs against radiation-initiated losses by decreasing the levels of oxidative stress and apoptosis. DXM may potentially be a therapeutic modality for preventing radiation-induced HL; further in vivo studies are necessary.

HYPOTHESIS Dexamethasone (DXM) protects against cisplatin-induced auditory hair cell (HC) loss in rat organ of Corti (OC) explants in vitro by reducing levels of oxidative stress and NADPH-Oxidase-3 (NOX-3). BACKGROUND Intratympanic DXM has demonstrated protective effects against cisplatin-induced hearing loss in a few animal studies and one clinical trial. However, levels of protection with intratympanic DXM vary significantly between studies, which may not be a result of the intrinsic properties of DXM but rather reflect the diffusion of DXM into the cochlea. The molecular mechanisms and degree of DXM protection against cisplatin ototoxicity are currently unknown. METHODS OC explants from 3-day-old rats were cultured with no treatment or various concentrations of cisplatin (2, 5, or 10 μM) and DXM (75, 150, or 300 μg/mL) in vitro. HC viability and TUNEL assay were performed after 72 hours in vitro and levels of oxidative stress and NOX-3 were evaluated with confocal microscopy after 48 hours in vitro. Analysis of variance with Tukey's post hoc testing was performed. RESULTS Cisplatin initiated dose-dependent losses of outer HCs (OHCs) in the basal turns of exposed explants (p

Gentamicin is a widely used antibiotic, which causes hearing loss because of destruction of auditory hair cells. Mannitol has been shown to have cytoprotective properties in the cochlea both in vitro and in vivo. Mannitol has been shown to be safe in concentrations up to 100 mM in organ of Corti explants. It is proposed as an otoprotective agent against gentamicin ototoxicity.Organ of Corti were dissected from P-3 rat pups and cultured under the following conditions for 96 hours: 1) control, 2) gentamicin (10 μM for all hair cell count experiments), 3) gentamicin + mannitol 10 mM, 4) gentamicin + mannitol 50 mM, and 5) gentamicin + mannitol 100 mM. The tissues were then fixed and stained, and hair cells were counted for segments of the apex, middle, and basal turns. Quantitative RT-PCR (qRT-PCR) was performed on organ of Corti explant extracted RNA after 24 hours in vitro: 1) control; 2) gentamicin (100 μM for all gene expression and CellRox experiments); 3) gentamicin +mannitol 100 mM; and 4) mannitol 100 mM for tumor necrosis factor-alpha (TNF-α), TNF-α receptor (TNFR1A), interleukin-1 beta (IL-1β) and cyclooxygenase-2 (COX-2). In vitro examination of oxidative stress was performed for the same test groups at 24 hours using CellRox Deep Red assay.Gentamicin induced loss of both inner hair cells and outer hair cells with increasing severity from apex to middle to basal segments (Pearson r = -0.999 for inner hair cells and -0.972 for outer hair cells). Mannitol demonstrated dose-dependent otoprotection of IHCs and outer hair cells (p0.001 for mannitol at 100 mM). CellRox demonstrated increased oxidative stress induced by gentamicin exposure, and this effect was attenuated by treatment of gentamicin-exposed explants with mannitol (p0.05). TNF-α, IL-1β TNFR1A, and COX-2 mRNA levels were upregulated by gentamicin (p0.05). Mannitol treatment of gentamicin explants downregulated the gene expression of the proinflammatory cytokines, but this difference did not achieve significance. Interestingly, in gentamicin-challenged organ of Corti explants, Mannitol upregulated the expression of TNFR1A, but this increase did not achieve significance (p0.05).Gentamicin ototoxicity is increasingly severe from the apex to basal turn of the cochlea. Treatment with mannitol prevents gentamicin-induced hair cell loss in a dose-dependent manner, protecting both IHCs and outer hair cells. Mannitol appears to act as a free radical scavenger to reduce the cytotoxic effects of gentamicin by reducing the level of oxidative stress.

Mannitol has otoprotective effects against tumor necrosis factor (TNF) α-induced auditory hair cell (HC) loss.Mannitol has been demonstrated to possess cytoprotective effects in several organ systems. Its protective effect on postischemic hearing loss has also been shown. Mannitol's otoprotective mechanism and site of action are at present unknown.Organ of Corti (OC) explants were dissected from 3 day-old rat pups. The safety (nonototoxicity) of mannitol was assessed at 4 different concentrations (1-100 mM). Three experimental arms were designed including: a control group, TNFα group, and TNFα + mannitol group. Cell viability was determined by counts of fluorescein isothiocyanate (FITC) phalloidin stained HC. Immunofluorescence assay of phospho-c-Jun and the proapoptotic mediators, cleaved caspase-3, apoptosis inducing factor (AIF), and endonuclease G (Endo G) were performed.Analysis of HC density confirmed the safety of mannitol at concentration ranges of 1 to 100 mM. The ototoxic effect of TNFα was demonstrated (p0.05). The otoprotective effect of 100 mM mannitol in TNFα-challenged OC explants was also demonstrated (p0.001). Mannitol treatment reduced the high levels of phospho-c-Jun observed in the TNFα-challenged group. AIF cluster formation and EndoG translocation into the nuclei of HCs were also reduced by mannitol treatment.Mannitol significantly reduces the ototoxic effects of TNFα against auditory HC's potentially by inhibiting c-Jun N terminal kinase (JNK) activation pathway and AIF, EndoG nuclear translocation. This local otoprotective effect may have therapeutic implications in inner ear surgery, for example, cochlear implants, protection of residual hearing, as well as implications for postischemic inner ear insults.

The progressive loss of hearing that develops after electrode insertion trauma (EIT) can be attenuated by local dexamethasone (DXM) therapy.Hearing loss (HL) that develops after cochlear implant EIT occurs in two stages in laboratory animals, that is, an immediate loss followed by a progressive loss. Direct infusion of DXM into the guinea pig cochlea can attenuate both ototoxin- and noise-induced HL.Auditory-evoked brainstem responses (ABRs) of guinea pigs were measured for 4 frequencies (i.e., 0.5, 1, 4, and 16 kHz) before, immediately after, and more than 30 days post-EIT for experimental (EIT,EIT + artificial perilymph, and EIT + DXM) and for the contralateral unoperated cochleae of each group. An electrode analog of 0.14-mm diameter was inserted through a basal turn cochleostomy for a depth of 3 mm and withdrawn. DXM in artificial perilymph was delivered immediately post-EIT into the scala tympani via a miniosmotic pump for 8 days.The ABR thresholds of EIT animals increased progressively post-EIT. Contralateral unoperated cochleae had no significant changes in ABR thresholds. Immediately post-EIT, that is, Day 0, the DXM-treated animals exhibited a significant HL at 1, 4, and 16 kHz, but this HL was no longer significant by Day 30 compared with contralateral control ears.The results from immediate local treatment of the cochlea with DXM in an animal model of EIT-induced HL suggest a novel therapeutic strategy for hearing conservation by attenuating the progressive HL that can result from the process of electrode array insertion during cochlear implantation.