Response to OKN-007 and NAC in a Patient with Unilateral Hearing Loss and Chronic Tinnitus from Vestibular Schwannoma.

BACKGROUND: Tinnitus is the perception of sound in the absence of external acoustic stimulation. Being one of the most common diseases of the ear, it has a global prevalence ranging from 4.1 to 37.2%. To date, it has been difficult to treat tinnitus as its pathophysiology is poorly understood and there are limited treatment options. OBJECTIVE: To investigate the effect of OKN-007 (also known as HPN-07), a nitrone-based investigational drug, in combination with oral N-acetylcycsteine (NAC), for the treatment of hearing loss and chronic tinnitus under an individual expanded access protocol. PATIENT CASE: We report the case of a patient who presented with left-sided ear fullness, mild tinnitus, and mild high frequency sensorineural hearing loss with 100% word recognition. A large enhancing mass seen on MRI revealed a vestibular schwannoma. He underwent subtotal resection of the tumor resulting in a moderate-to-profound sensorineural hearing loss and catastrophic tinnitus. The patient was treated with intravenous OKN-007 at 60 mg/kg dosed three times per week and oral NAC 2500 mg twice daily. RESULTS: Post-treatment audiometric testing revealed an average of 16.66 dB in hearing threshold improvement in three frequencies (125, 250 and 500 Hz) with residual hearing in the affected left ear. His tinnitus loudness matching improved from 90 dB to 19 dB post-treatment. His Tinnitus Handicap Inventory improved from 86/100 (Catastrophic) to 40/100 (Moderate). He also experienced improvements in sleep, concentration, hearing, and emotional well-being, and reported significantly decreased levels of tinnitusrelated distress. CONCLUSIONS: This case report highlights the feasibility and therapeutic potential of the combination of OKN-007 and NAC in treating hearing loss and tinnitus that warrants further investigation.

Regeneration of Cochlear Hair Cells and Hearing Recovery through Hes1 Modulation with siRNA Nanoparticles in Adult Guinea Pigs.

Deafness is commonly caused by the irreversible loss of mammalian cochlear hair cells (HCs) due to noise trauma, toxins, or infections. We previously demonstrated that small interfering RNAs (siRNAs) directed against the Notch pathway gene, hairy and enhancer of split 1 (Hes1), encapsulated within biocompatible poly(lactic-co-glycolic acid) nanoparticles (PLGA NPs) could regenerate HCs within ototoxin-ablated murine organotypic cultures. In the present study, we delivered this sustained-release formulation of Hes1 siRNA (siHes1) into the cochleae of noise-injured adult guinea pigs. Auditory functional recovery was measured by serial auditory brainstem responses over a nine-week follow-up period, and HC regeneration was evaluated by immunohistological evaluations and scanning electron microscopy. Significant HC restoration and hearing recovery were observed across a broad tonotopic range in ears treated with siHes1 NPs, beginning at three weeks and extending out to nine weeks post-treatment. Moreover, both ectopic and immature HCs were uniquely observed in noise-injured cochleae treated with siHes1 NPs, consistent with de novo HC production. Our results indicate that durable cochlear HCs were regenerated and promoted significant hearing recovery in adult guinea pigs through reversible modulation of Hes1 expression. Therefore, PLGA-NP-mediated delivery of siHes1 to the cochlea represents a promising pharmacologic approach to regenerate functional and sustainable mammalian HCs in vivo.

Enhanced sucrose-mediated cryoprotection of siRNA-loaded poly (lactic-co-glycolic acid) nanoparticles.

The present study aimed to determine the effects of sucrose on the physical stability, cellular entry pathways and functional efficacy of poly(lactic-co-glycolic acid) nanoparticles (PLGA-NPs). PLGA-NPs were synthesized in the absence or presence of 10 % sucrose, using HEI-101, an unmodified small interfering RNA (siRNA), as a drug model. The newly synthesized HEI-101-loaded PLGA-NPs (HEI-101-NPs) were exposed to repeated freeze-thaw cycles and iteratively tested over a six-month evaluation period. The effect of sucrose stabilization on HEI-101-NPs was independently tested in vitro for biocompatibility and cellular uptake in IMO-2B1 cells. Data analyses suggest that, without sucrose, freeze-thaw cycles of HEI-101-NPs resulted in increased particle diameter, increased polydispersity index, and reduced zeta potential. In contrast, a substantial improvement in the physical stability of HEI-101-NPs was observed in the presence of 10 % sucrose. The data revealed that the release of HEI-101 from the PLGA-NPs was governed by polymer erosion and drug diffusion. Data from cellular uptake study in IMO-2B1 cells demonstrated that, 10 % sucrose significantly reduced the inhibitory effect of nocodazole on the microtubule-dependent uptake of PLGA-NPs. In addition, the presence of 10 % sucrose seemed to lessen the inhibitory effect of sodium azide on the energy-dependent uptake of PLGA-NPs. Overall, the current data suggest that the cellular internalization of PLGA-NPs occurred through the polymerization of actin filaments under the control of the microtubules. Our findings reveal cryoprotective effect of 10 % sucrose on HEI-101-NPs that confers marked improvements in the stability, cellular uptake and efficiency for the delivery of biomolecules to inner ear cells.

Electrophysiological assessment and pharmacological treatment of blast-induced tinnitus.

Tinnitus, the phantom perception of sound, often occurs as a clinical sequela of auditory traumas. In an effort to develop an objective test and therapeutic approach for tinnitus, the present study was performed in blast-exposed rats and focused on measurements of auditory brainstem responses (ABRs), prepulse inhibition of the acoustic startle response, and presynaptic ribbon densities on cochlear inner hair cells (IHCs). Although the exact mechanism is unknown, the "central gain theory" posits that tinnitus is a perceptual indicator of abnormal increases in the gain (or neural amplification) of the central auditory system to compensate for peripheral loss of sensory input from the cochlea. Our data from vehicle-treated rats supports this rationale; namely, blast-induced cochlear synaptopathy correlated with imbalanced elevations in the ratio of centrally-derived ABR wave V amplitudes to peripherally-derived wave I amplitudes, resulting in behavioral evidence of tinnitus. Logistic regression modeling demonstrated that the ABR wave V/I amplitude ratio served as a reliable metric for objectively identifying tinnitus. Furthermore, histopathological examinations in blast-exposed rats revealed tinnitus-related changes in the expression patterns of key plasticity factors in the central auditory pathway, including chronic loss of Arc/Arg3.1 mobilization. Using a formulation of N-acetylcysteine (NAC) and disodium 2,4-disulfophenyl-N-tert-butylnitrone (HPN-07) as a therapeutic for addressing blast-induced neurodegeneration, we measured a significant treatment effect on preservation or restoration of IHC ribbon synapses, normalization of ABR wave V/I amplitude ratios, and reduced behavioral evidence of tinnitus in blast-exposed rats, all of which accorded with mitigated histopathological evidence of tinnitus-related neuropathy and maladaptive neuroplasticity.

Dosing study on the effectiveness of salicylate/N-acetylcysteine for prevention of noise-induced hearing loss.

The efficacy of three different doses of sodium salicylate (SAL) in combination with one dose of N-acetylcysteine (NAC) to prevent noise-induced hearing loss was studied in chinchillas. After obtaining baseline-hearing thresholds, the chinchillas were randomly assigned to one of four treatment groups: three sets were injected intraperitoneally with 325 mg/kg NAC combined with 25, 50, or 75 mg/kg SAL, and a separate control group was injected with an equal volume of saline. Animals were injected twice daily for 2 days prior to and 1 hour before the noise exposure (6 hours to a 105-dB Standard Pressure Level octave band noise centered at 4 kHz). Immediate post-noise hearing thresholds were obtained followed by post-noise treatments at 1 hour then twice-daily for 2 days. Hearing tests continued at 1, 2, and 3 weeks post-noise, and immediately after the last hearing test, animals' cochleae were stained for hair cell counts. All the groups showed hearing improvement until week 2. However, at week 3, saline treated animals demonstrated a 17-33 dB SPL permanent threshold shift (PTS) across the test frequencies. Hearing loss was lowest in the 50 SAL/325 NAC mg/kg group (all frequencies, P < 0.001), and although PTS was reduced in the 25 and 75 mg/kg SAL dosage groups compared to the saline group, only the 75 mg/kg SAL group was significantly different at all but 2 kHz frequency. Coupled with the hearing loss, outer hair cell (OHC) loss was maximal in the 4-8 kHz cochlear region of saline treated animals. However, there was a substantial reduction in the mean OHC loss of the NAC plus 50 or 75 mg/kg (but not the 25 mg/kg) SAL groups. These findings suggest that SAL in combination with NAC is effective in reducing noise damage to the cochlea, but SAL has a relatively narrow therapeutic dosing window.

Gene therapy for hair cell regeneration: Review and new data.

Hair cells (HCs) in the cochlea are responsible for transducing mechanical sound energy into neural impulses which lead to the perception of sound. Loss of these sensory cells is the most common cause of sensorineural hearing loss, and spontaneous HC regeneration does not occur in mature mammals. Among the future potential treatment modalities is gene therapy, which is defined as the administration of either DNAs or RNAs as active pharmaceutical ingredients for inducing a clinically-beneficial response. Gene therapy is being envisioned and evaluated as a potential tool for addressing a number of human inner ear disorders. This paper is a hybrid Review and Research Paper, including unpublished data and a review of HC regeneration studies in live animal models. Current gene therapeutic approaches for replacing lost HC populations have been aimed at converting supporting cells surviving within the neuro-epithelium to new HCs by inducing upregulation of bHLH transcription factors such as Atoh1 or reciprocal silencing of Notch signaling with siRNAs, to tip the balance of transcriptional regulation toward a HC fate. Development of one or more of these techniques may yield a path to effective restoration of inner ear form and function. This review also describes other approaches and molecular targets that may prove efficacious and provides perspectives on future clinical challenges and opportunities for gene therapy to become a valuable weapon for the long-anticipated realization of this regenerative treatment.

Detecting incipient inner-ear damage from impulse noise with otoacoustic emissions.

Audiometric thresholds and otoacoustic emissions (OAEs) were measured in 285 U.S. Marine Corps recruits before and three weeks after exposure to impulse-noise sources from weapons' fire and simulated artillery, and in 32 non-noise-exposed controls. At pre-test, audiometric thresholds for all ears were

Effectiveness of 4-hydroxy phenyl N-tert-butylnitrone (4-OHPBN) alone and in combination with other antioxidant drugs in the treatment of acute acoustic trauma in chinchilla.

Acute acoustic trauma (AAT) results in oxidative stress to the cochlea through overproduction of cellular reactive oxygen, nitrogen, and other free radical species appearing from 1 h to 10 days after noise exposure. It has been shown that N-acetyl-L-cysteine (NAC), a glutathione prodrug, and acetyl-L-carnitine (ALCAR), a mitochondrial biogenesis agent, are effective in reducing noise-induced hearing loss. Phenyl N-tert-butylnitrone (PBN), a nitrone-based free radical trap, appears to suppress oxidative stress in a variety of disorders and several biological models. In this study, we tested whether 4-hydroxy PBN (4-OHPBN), a major metabolite of PBN, administered 4 h after noise exposure is effective in treating noise-induced hearing loss and whether a combination of antioxidant drugs (4-OHPBN plus NAC and 4-OHPBN plus NAC plus ALCAR) provides greater efficacy in attenuating AAT since each agent addresses different injury mechanisms. Chinchilla were exposed to a 105 dB octave-band noise centered at 4 kHz for 6 h. 4-OHPBN and combinations of antioxidant drugs were intraperitoneally administered beginning 4 h after noise exposure. Hearing threshold shifts in auditory brainstem responses and missing outer hair cell counts were obtained. 4-OHPBN reduced threshold shifts in a dose-dependent manner while both drug combinations showed greater effects. These results demonstrate that 4-OHPBN and combinations of antioxidants can effectively treat acute acoustic trauma and drug combinations may increase the effectiveness of treatment and decrease the required individual medication dose.

Nitrones as therapeutics.

Nitrones have the general chemical formula X-CH=NO-Y. They were first used to trap free radicals in chemical systems and then subsequently in biochemical systems. More recently several nitrones, including alpha-phenyl-tert-butylnitrone (PBN), have been shown to have potent biological activity in many experimental animal models. Many diseases of aging, including stroke, cancer development, Parkinson disease, and Alzheimer disease, are known to have enhanced levels of free radicals and oxidative stress. Some derivatives of PBN are significantly more potent than PBN and have undergone extensive commercial development for stroke. Recent research has shown that PBN-related nitrones also have anti-cancer activity in several experimental cancer models and have potential as therapeutics in some cancers. Also, in recent observations nitrones have been shown to act synergistically in combination with antioxidants in the prevention of acute acoustic-noise-induced hearing loss. The mechanistic basis of the potent biological activity of PBN-related nitrones is not known. Even though PBN-related nitrones do decrease oxidative stress and oxidative damage, their potent biological anti-inflammatory activity and their ability to alter cellular signaling processes cannot readily be explained by conventional notions of free radical trapping biochemistry. This review is focused on our studies and others in which the use of selected nitrones as novel therapeutics has been evaluated in experimental models in the context of free radical biochemical and cellular processes considered important in pathologic conditions and age-related diseases.

Influence of ultrahigh-frequency hearing thresholds on distortion-product otoacoustic emission levels at conventional frequencies.

This study examined the association between ultrahigh-frequency (UHF) hearing sensitivity and distortion-product otoacoustic emission (DPOAE) levels at conventional frequencies. Behavioral thresholds were measured from 2 through 16 kHz, and DPOAE levels were measured at discrete f2 frequencies between 2 through 8 kHz in 553 young normal-hearing adult male participants. A DPOAE frequency sweep was measured with primary stimulus levels of L1/L2 = 65/55 dB SPL and an f2/f1 of 1.2. Significant negative correlations, although weak, were found between UHF behavioral thresholds and DPOAE levels. As UHF behavioral thresholds worsened, DPOAE levels decreased at all frequencies. When the data were categorized into two groups, "better" and "worse" UHF behavioral thresholds, significant differences were apparent between the two groups for DPOAEs. Additionally, those with better UHF thresholds had better conventional thresholds compared to those in the worse UHF threshold group. The results of this age-restricted, large-sample-size study confirm and augment findings from earlier studies demonstrating that UHF hearing sensitivity has some influence on DPOAE measures at frequencies from 2 through 8 kHz with moderate stimulus levels. However, because those with better UHF thresholds also had better conventional thresholds and the significant correlations found were weak, this work supports the importance of UHF hearing testing in conjunction with otoacoustic emission measures to identify basal cochlear insults not evident from behavioral testing at conventional frequencies.

NAC for noise: from the bench top to the clinic.

Noise-induced hearing loss (NIHL) is an important etiology of deafness worldwide. Hearing conservation programs are in place and have reduced the prevalence of NIHL, but this disorder is still far too common. Occupational and recreational pursuits expose people to loud noise and ten million persons in the US have some degree of noise-induced hearing impairment. It is estimated that 50 million in the US and 600 million people worldwide are exposed to noise hazards occupationally. Noise deafness is still an important and frequent cause of battlefield injury in the US military. A mainstay of hearing conservation programs is personal mechanical hearing protection devices which are helpful but have inherent limitations. Research has shown that oxidative stress plays an important role in noise-induced cochlear injury resulting in the discovery that a number of antioxidant and cell death inhibiting compounds can ameliorate deafness associated with acoustic trauma. This article reviews one such compound, N-acetylcysteine (NAC), in terms of its efficacy in reducing hearing loss in a variety of animal models of acute acoustic trauma and hypothesizes what its therapeutic mechanisms of action might be based on the known actions of NAC. Early clinical trials with NAC are mentioned.

Distribution of PLGA nanoparticles in chinchilla cochleae.

OBJECTIVES: To study the distribution of polylactic/glycolic acid-encapsulated iron oxide nanoparticles (PLGA-NPs) in chinchilla cochleae after application on the round window membrane (RWM). STUDY DESIGN AND SETTING: Six chinchillas (12 ears) were equally divided into controls (no treatments) and experimentals (PLGA-NP with or without magnetic exposure). After 40 minutes of PLGA-NP placement on the RWM, perilymph was withdrawn from the scala tympani. The RWM and cochleae were fixed with 2.5% glutaraldehyde and processed for transmission electron microscopy. RESULTS: Nanoparticles were found in cochleae with or without exposure to magnet forces appearing in the RWM, perilymph, endolymph, and multiple locations in the organ of Corti. Electron energy loss spectroscopy confirmed iron elements in nanoparticles. CONCLUSION: The nanoparticles were distributed throughout the inner ear after application on the chinchilla RWM, with and without magnetic forces. SIGNIFICANCE: PLGA-NP applied to the RWM may have potential for sustained therapy to the inner ear.

Noise protection with N-acetyl-l-cysteine (NAC) using a variety of noise exposures, NAC doses, and routes of administration.

CONCLUSION: These studies extend previous work on N-acetyl-l-cysteine (NAC) and noise, showing protection with NAC against a high-kurtosis noise, showing protection with NAC at low doses, as well as protection by oral gavage. The studies further reveal the potential for the use of NAC in a clinical population exposed to noise. OBJECTIVE: To extend previous work on NAC protection from noise, the current study examined the effectiveness of NAC against a high-kurtosis noise that combined continuous and impact noise, tested the effectiveness of NAC at varying doses, and tested NAC when administered by gavage. MATERIALS AND METHODS: Chinchillas were tested for auditory brainstem responses (ABRs) at five frequencies before and at three time points after one of three noise exposures: high-kurtosis (2 h, 108 dB L(eq)), impulse (75 pairs of 155 dB pSPL impulses), or continuous (4 kHz octave band, 105 dB SPL for 6 h). Animals were treated with NAC or saline vehicle before and after noise. RESULTS: The NAC was protective against the high-kurtosis noise both at low doses and when given orally by gavage.

Risk factors for distortion product otoacoustic emissions in young men with normal hearing.

The purpose of this study was to evaluate the possible effects of risk factors on distortion product otoacoustic emissions (DPOAEs) in young adult men with normal hearing. Four hundred thirty-six United States Marine recruit men (mean age = 19.2 years +/- 1.8 years; age range = 17-29 years) participated in this study. Questionnaires were given to each recruit to obtain demographic data and history of noise exposure, solvent exposure, smoking history, and hearing-related histories. Otoscopy, tympanometry, pure-tone air-conduction audiometry (2.0-8.0 kHz) and DPOAEs (2.3-8.0 kHz) were measured. DPOAE levels were lower in Not Hispanic or Latino recruits, in heavy smokers, in recruits who reported loud live music exposure and ringing in their ears after noise exposure. These differences were not statistically significant at all frequencies. Recruits with multiple risk factors had the lowest DPOAEs as compared to recruits with fewer, or no, risk factors; these differences were not statistically significant. Obtaining risk factor data as part of an audiometric evaluation is important even though the individual may have normal hearing.

siRNA-loaded biodegradable nanocarriers for therapeutic MAPK1 silencing against cisplatin-induced ototoxicity.

Ototoxicity represents a major adverse side-effect of cis-diamminedichloroplatinum-II (cisplatin, CDDP). The mitogen-activated protein kinase (MAPK) pathway is thought to play a central role in potentiating the apoptotic effect of CDDP within the cochlea. We hypothesized that prophylactic inhibition of MAPK signaling, using small interfering RNA (siRNA), might confer a protective effect against CDDP-induced apoptosis within the auditory sensory epithelia. To enhance the therapeutic utility of this approach, we synthesized biocompatible siMAPK1-loaded nanoparticles (NPs) and performed physicochemical characterizations for size, morphology, drug loading and release kinetics, using dynamic light scattering, electron microscopy and spectrophotometric analyses, respectively. Our findings show 183.88±6.26 nm-sized spherical siMAPK1-loaded NPs with -27.12±6.65mV zeta potential and 112.78±0.24pmol/mg of siMAPK1 loading that exhibit a sustained release profile for prolonged therapeutic efficacy. Synthesized NPs were validated for biocompatibility and prophylactically protected against CDDP-induced cytotoxicity in HEI-OC1 cells and hair cell loss in murine organotypic cochlear explants. Our study confirms a pivotal role for MAPK1 signaling as a potentiating factor for CDDP-induced apoptosis and cochlear hair cell loss, and highlights siMAPK1 NP treatment as a therapeutic strategy for limiting the ototoxic side-effects associated with systemic CDDP administration.

HPN-07, a free radical spin trapping agent, protects against functional, cellular and electrophysiological changes in the cochlea induced by acute acoustic trauma.

Oxidative stress is considered a major cause of the structural and functional changes associated with auditory pathologies induced by exposure to acute acoustic trauma AAT). In the present study, we examined the otoprotective effects of 2,4-disulfophenyl-N-tert-butylnitrone (HPN-07), a nitrone-based free radical trap, on the physiological and cellular changes in the auditory system of chinchilla following a six-hour exposure to 4 kHz octave band noise at 105 dB SPL. HPN-07 has been shown to suppress oxidative stress in biological models of a variety of disorders. Our results show that administration of HPN-07 beginning four hours after acoustic trauma accelerated and enhanced auditory/cochlear functional recovery, as measured by auditory brainstem responses (ABR), distortion product otoacoustic emissions (DPOAE), compound action potentials (CAP), and cochlear microphonics (CM). The normally tight correlation between the endocochlear potential (EP) and evoked potentials of CAP and CM were persistently disrupted after noise trauma in untreated animals but returned to homeostatic conditions in HPN-07 treated animals. Histological analyses revealed several therapeutic advantages associated with HPN-07 treatment following AAT, including reductions in inner and outer hair cell loss; reductions in AAT-induced loss of calretinin-positive afferent nerve fibers in the spiral lamina; and reductions in fibrocyte loss within the spiral ligament. These findings support the conclusion that early intervention with HPN-07 following an AAT efficiently blocks the propagative ototoxic effects of oxidative stress, thereby preserving the homeostatic and functional integrity of the cochlea.

Antioxidants reduce neurodegeneration and accumulation of pathologic Tau proteins in the auditory system after blast exposure.

Cochlear neurodegeneration commonly accompanies hair cell loss resulting from aging, ototoxicity, or exposures to intense noise or blast overpressures. However, the precise pathophysiological mechanisms that drive this degenerative response have not been fully elucidated. Our laboratory previously demonstrated that non-transgenic rats exposed to blast overpressures exhibited marked somatic accumulation of neurotoxic variants of the microtubule-associated protein, Tau, in the hippocampus. In the present study, we extended these analyses to examine neurodegeneration and pathologic Tau accumulation in the auditory system in response to blast exposure and evaluated the potential therapeutic efficacy of antioxidants on short-circuiting this pathological process. Blast injury induced ribbon synapse loss and retrograde neurodegeneration in the cochlea in untreated animals. An accompanying perikaryal accumulation of neurofilament light chain and pathologic Tau oligomers were observed in neurons from both the peripheral and central auditory system, spanning from the spiral ganglion to the auditory cortex. Due to its coincident accumulation pattern and well-documented neurotoxicity, our results suggest that the accumulation of pathologic Tau oligomers may actively contribute to blast-induced cochlear neurodegeneration. Therapeutic intervention with a combinatorial regimen of 2,4-disulfonyl α-phenyl tertiary butyl nitrone (HPN-07) and N-acetylcysteine (NAC) significantly reduced both pathologic Tau accumulation and indications of ongoing neurodegeneration in the cochlea and the auditory cortex. These results demonstrate that a combination of HPN-07 and NAC administrated shortly after a blast exposure can serve as a potential therapeutic strategy for preserving auditory function among military personnel or civilians with blast-induced traumatic brain injuries.

The permeability of SPION over an artificial three-layer membrane is enhanced by external magnetic field.

BACKGROUND: Sensorineural hearing loss, a subset of all clinical hearing loss, may be correctable through the use of gene therapy. We are testing a delivery system of therapeutics through a 3 cell-layer round window membrane model (RWM model) that may provide an entry of drugs or genes to the inner ear. We designed an in vitro RWM model similar to the RWM (will be referred to throughout the paper as RWM model) to determine the feasibility of using superparamagnetic iron oxide (Fe3O4) nanoparticles (SPION) for targeted delivery of therapeutics to the inner ear. The RWM model is a 3 cell-layer model with epithelial cells cultured on both sides of a small intestinal submucosal (SIS) matrix and fibroblasts seeded in between. Dextran encapsulated nanoparticle clusters 130 nm in diameter were pulled through the RWM model using permanent magnets with flux density 0.410 Tesla at the pole face. The SIS membranes were harvested at day 7 and then fixed in 4% paraformaldehyde. Transmission electron microscopy and fluorescence spectrophotometry were used to verify transepithelial transport of the SPION across the cell-culture model. Histological sections were examined for evidence of SPION toxicity, as well to generate a timeline of the position of the SPION at different times. SPION also were added to cells in culture to assess in vitro toxicity. RESULTS: Transepithelial electrical resistance measurements confirmed epithelial confluence, as SPION crossed a membrane consisting of three co-cultured layers of cells, under the influence of a magnetic field. Micrographs showed SPION distributed throughout the membrane model, in between cell layers, and sometimes on the surface of cells. TEM verified that the SPION were pulled through the membrane into the culture well below. Fluorescence spectrophotometry quantified the number of SPION that went through the SIS membrane. SPION showed no toxicity to cells in culture. CONCLUSION: A three-cell layer model of the human round window membrane has been constructed. SPION have been magnetically transported through this model, allowing quantitative evaluation of prospective targeted drug or gene delivery through the RWM. Putative in vivo carrier superparamagnetic nanoparticles may be evaluated using this model.

Efficacy of the antioxidant N-acetylcysteine (NAC) in protecting ears exposed to loud music.

Antioxidants have been reported to be effective in reducing acoustic trauma in animal models but have not been studied in humans. In this study, the antioxidant N-acetylcysteine (NAC) was evaluated to determine if it would reduce temporary changes in auditory function as a result of exposure to loud music in humans. Pure-tone thresholds and distortion product otoacoustic emissions (DPOAEs) were collected in 31 normal-hearing participants, using a randomized, double-blind, placebo-controlled design, before and after two hours of live music in a nightclub. Using repeated measures analysis of variance, no statistically significant differences were found between participants who received NAC versus a placebo for any of the outcome measures. Across all subjects, the largest pure-tone threshold shift occurred at 4 kHz. DPOAE measures were characterized by reductions in amplitude and a trend for shorter group delay values. When the 3 and 4 kHz data were examined by imposing specific criteria of greater than 2 dB DPOAE amplitude reductions and 10 dB or greater pure-tone threshold shifts, DPOAE reductions occurred more often at 3 kHz, and pure-tone shifts occurred more often at 4 kHz.

Ameliorative Effects of Antioxidants on the Hippocampal Accumulation of Pathologic Tau in a Rat Model of Blast-Induced Traumatic Brain Injury.

Traumatic brain injury (TBI) can lead to early onset dementia and other related neurodegenerative diseases. We previously demonstrated that damage to the central auditory pathway resulting from blast-induced TBI (bTBI) could be significantly attenuated by a combinatorial antioxidant treatment regimen. In the current study, we examined the localization patterns of normal Tau and the potential blast-induced accumulation of neurotoxic variants of this microtubule-associated protein that are believed to potentiate the neurodegenerative effects associated with synaptic dysfunction in the hippocampus following three successive blast overpressure exposures in nontransgenic rats. We observed a marked increase in the number of both hyperphosphorylated and oligomeric Tau-positive hilar mossy cells and somatic accumulation of endogenous Tau in oligodendrocytes in the hippocampus. Remarkably, a combinatorial regimen of 2,4-disulfonyl α-phenyl tertiary butyl nitrone (HPN-07) and N-acetylcysteine (NAC) resulted in striking reductions in the numbers of both mossy cells and oligodendrocytes positively labeled for these pathological Tau immunoreactivity patterns in response to bTBI. This treatment strategy represents a promising therapeutic approach for simultaneously reducing or eliminating both primary auditory injury and nonauditory changes associated with bTBI-induced hippocampal neurodegeneration.