Effects of an enhanced acoustic environment on residual hearing following chronic cochlear implantation and electrical stimulation in the partially deafened cat.

There is an increasing trend to provide cochlear implants for people with useful residual hearing, typically in the low frequency range (<2 kHz). These recipients typically use both electrical stimulation from their implant and acoustic stimulation that has been amplified with a hearing aid to access their residual hearing, so called electro-acoustic stimulation (EAS). However, a significant problem is the loss of residual hearing following implantation that can occur immediately following surgery or delayed over many months. One potential cause of the loss of residual hearing is the over stimulation of remaining hair cells due to the combination of an amplified acoustic input and direct electrical activation. This paper aims to test this hypothesis. Here, we have used a neonatal aminoglycoside-induced partial hearing cat model that resulted in a high frequency hearing loss (>4 kHz). Two separate cohorts of animals were implanted and received unilateral chronic electrical stimulation using clinical stimulators and speech processors over 5 months. To simulate potential over stimulation via a hearing aid, one cohort of animals were also exposed to an enhanced acoustic environment consisting of 80 dB SPL 4-talker babble presented 14 h per day. Hearing thresholds for both stimulated and unstimulated ears were measured throughout the implantation period. Cochleae were collected for histology to measure spiral ganglion neuron survival, hair cell survival and tissue response to chronic implantation and electrical stimulation. Consistent with clinical observations, cochlear implantation and stimulation resulted in an increase in threshold across the population. There was no significant effect of the enhanced acoustic environment on auditory thresholds or histological measures (hair cell survival, neuronal survival) of hearing, indicating that hair cell overstimulation was not a significant driver of loss of residual hearing.

Effects of chronic implantation and long-term stimulation of a cochlear implant in the partial hearing cat model.

The expansion of criteria for cochlear implantation has resulted in increasing numbers of cochlear implant subjects having some level of residual hearing. The present study examined the effects of implantation surgery and long-term electrical stimulation on residual hearing in a partially deafened cat model. Eighteen animals were partially deafened, implanted and chronically stimulated. Implantation resulted in a pronounced loss evident 2-weeks post implantation of up to 30-40 dB at 4 & 8 kHz which was statistically significant (2-way RM ANOVA (Time, Frequency): p(Time) = 0.001; p(Frequency) < 0.001; p(Time x Frequency) < 0.001)). Chronic stimulation resulted in a significant (RM ANOVA: p(Time) = 0.030) ongoing hearing loss, with 5 animals (∼30%) exhibiting an increase in threshold of 20 dB or more. Different loss profiles were evident with peripheral and central hearing assessments suggests that changes in 'central gain' may be occurring. Despite significant loss of hair cells and spiral ganglion neurons and distinct fibrous tissue growth in the scala tympani following implantation and long-term electrical stimulation, there were no significant correlations with any histological measures and ongoing hearing loss. The partially deafened, chronically stimulated cat model provides a clinically relevant model in which to further investigate the cause of the delayed hearing loss following cochlear implant surgery and use.

Gel-Mediated Electrospray Assembly of Silica Supraparticles for Sustained Drug Delivery.

Supraparticles (SPs) composed of smaller colloidal particles provide a platform for the long-term, controlled release of therapeutics in biomedical applications. However, current synthesis methods used to achieve high drug loading and those involving biocompatible materials are often tedious and low throughput, thereby limiting the translation of SPs to diverse applications. Herein, we present a simple, effective, and automatable alginate-mediated electrospray technique for the assembly of robust spherical silica SPs (Si-SPs) for long-term (>4 months) drug delivery. The Si-SPs are composed of either porous or nonporous primary Si particles within a decomposable alginate matrix. The size and shape of the Si-SPs can be tailored by controlling the concentrations of alginate and silica primary particles used and key electrospraying parameters, such as flow rate, voltage, and collector distance. Furthermore, the performance (including drug loading kinetics, loading capacity, loading efficiency, and drug release) of the Si-SPs can be tuned by changing the porosity of the primary particles and through the retention or removal (via calcination) of the alginate matrix. The structure and morphology of the Si-SPs were characterized by electron microscopy, dynamic light scattering, N2 adsorption-desorption analysis, and X-ray photoelectron spectroscopy. The cytotoxicity and degradability of the Si-SPs were also examined. Drug loading kinetics and loading capacity for six different types of Si-SPs, using a model protein drug (fluorescently labeled lysozyme), demonstrate that Si-SPs prepared from primary silica particles with large pores can load significant amounts of lysozyme (∼10 µg per SP) and exhibit sustained, long-term release of more than 150 days. Our experiments show that Si-SPs can be produced through a gel-mediated electrospray technique that is robust and automatable (important for clinical translation and commercialization) and that they present a promising platform for long-term drug delivery.

Treating hearing disorders with cell and gene therapy.

Hearing loss is an increasing problem for a substantial number of people and, with an aging population, the incidence and severity of hearing loss will become more significant over time. There are very few therapies currently available to treat hearing loss, and so the development of new therapeutic strategies for hearing impaired individuals is of paramount importance to address this unmet clinical need. Most forms of hearing loss are progressive in nature and therefore an opportunity exists to develop novel therapeutic approaches to slow or halt hearing loss progression, or even repair or replace lost hearing function. Numerous emerging technologies have potential as therapeutic options. This paper details the potential of cell- and gene-based therapies to provide therapeutic agents to protect sensory and neural cells from various insults known to cause hearing loss; explores the potential of replacing lost sensory and nerve cells using gene and stem cell therapy; and describes the considerations for clinical translation and the challenges that need to be overcome.

Hair cell regeneration after ATOH1 gene therapy in the cochlea of profoundly deaf adult guinea pigs.

The degeneration of hair cells in the mammalian cochlea results in permanent sensorineural hearing loss. This study aimed to promote the regeneration of sensory hair cells in the mature cochlea and their reconnection with auditory neurons through the introduction of ATOH1, a transcription factor known to be necessary for hair cell development, and the introduction of neurotrophic factors. Adenoviral vectors containing ATOH1 alone, or with neurotrophin-3 and brain derived neurotrophic factor were injected into the lower basal scala media of guinea pig cochleae four days post ototoxic deafening. Guinea pigs treated with ATOH1 gene therapy, alone, had a significantly greater number of cells expressing hair cell markers compared to the contralateral non-treated cochlea when examined 3 weeks post-treatment. This increase, however, did not result in a commensurate improvement in hearing thresholds, nor was there an increase in synaptic ribbons, as measured by CtBP2 puncta after ATOH1 treatment alone, or when combined with neurotrophins. However, hair cell formation and synaptogenesis after co-treatment with ATOH1 and neurotrophic factors remain inconclusive as viral transduction was reduced due to the halving of viral titres when the samples were combined. Collectively, these data suggest that, whilst ATOH1 alone can drive non-sensory cells towards an immature sensory hair cell phenotype in the mature cochlea, this does not result in functional improvements after aminoglycoside-induced deafness.

Viability of long-term gene therapy in the cochlea.

Gene therapy has been investigated as a way to introduce a variety of genes to treat neurological disorders. An important clinical consideration is its long-term effectiveness. This research aims to study the long-term expression and effectiveness of gene therapy in promoting spiral ganglion neuron survival after deafness. Adenoviral vectors modified to express brain derived neurotrophic factor or neurotrophin-3 were unilaterally injected into the guinea pig cochlea one week post ototoxic deafening. After six months, persistence of gene expression and significantly greater neuronal survival in neurotrophin-treated cochleae compared to the contralateral cochleae were observed. The long-term gene expression observed indicates that gene therapy is potentially viable; however the degeneration of the transduced cells as a result of the original ototoxic insult may limit clinical effectiveness. With further research aimed at transducing stable cochlear cells, gene therapy may be an efficacious way to introduce neurotrophins to promote neuronal survival after hearing loss.

Neurotrophin gene therapy for sustained neural preservation after deafness.

The cochlear implant provides auditory cues to profoundly deaf patients by electrically stimulating the residual spiral ganglion neurons. These neurons, however, undergo progressive degeneration after hearing loss, marked initially by peripheral fibre retraction and ultimately culminating in cell death. This research aims to use gene therapy techniques to both hold and reverse this degeneration by providing a sustained and localised source of neurotrophins to the deafened cochlea. Adenoviral vectors containing green fluorescent protein, with or without neurotrophin-3 and brain derived neurotrophic factor, were injected into the lower basal turn of scala media of guinea pigs ototoxically deafened one week prior to intervention. This single injection resulted in localised and sustained gene expression, principally in the supporting cells within the organ of Corti. Guinea pigs treated with adenoviral neurotrophin-gene therapy had greater neuronal survival compared to contralateral non-treated cochleae when examined at 7 and 11 weeks post injection. Moreover; there was evidence of directed peripheral fibre regrowth towards cells expressing neurotrophin genes after both treatment periods. These data suggest that neurotrophin-gene therapy can provide sustained protection of spiral ganglion neurons and peripheral fibres after hearing loss.

The effect of deafness duration on neurotrophin gene therapy for spiral ganglion neuron protection.

A cochlear implant can restore hearing function by electrically exciting spiral ganglion neurons (SGNs) in the deaf cochlea. However, following deafness SGNs undergo progressive degeneration ultimately leading to their death. One significant cause of SGN degeneration is the loss of neurotrophic support that is normally provided by cells within the organ of Corti (OC). The administration of exogenous neurotrophins (NTs) can protect SGNs from degeneration but the effects are short-lived once the source of NTs has been exhausted. NT gene therapy, whereby cells within the cochlea are transfected with genes enabling them to produce NTs, is one strategy for providing a cellular source of NTs that may provide long-term support for SGNs. As the SGNs normally innervate sensory cells within the OC, targeting residual OC cells for gene therapy in the deaf cochlea may provide a source of NTs for SGN protection and targeted regrowth of their peripheral fibers. However, the continual degeneration of the OC over extended periods of deafness may deplete the cellular targets for NT gene therapy and hence limit the effectiveness of this method in preventing SGN loss. This study examined the effects of deafness duration on the efficacy of NT gene therapy in preventing SGN loss in guinea pigs that were systemically deafened with aminoglycosides. Adenoviral vectors containing green fluorescent protein (GFP) with or without genes for Brain Derived Neurotrophic Factor (BDNF) and Neurotrophin-3 (NT3) were injected into the scala media (SM) compartment of cochleae that had been deafened for one, four or eight weeks prior to the viral injection. The results showed that viral transfection of cells within the SM was still possible even after severe degeneration of the OC. Supporting cells (pillar and Deiters' cells), cells within the stria vascularis, the spiral ligament, endosteal cells lining the scala compartments and interdental cells in the spiral limbus were transfected. However, the level of transfection was remarkably lower following longer durations of deafness. There was a significant increase in SGN survival in the entire basal turn for cochleae that received NT gene therapy compared to the untreated contralateral control cochleae for the one week deaf group. In the four week deaf group significant SGN survival was observed in the lower basal turn only. There was no increase in SGN survival for the eight week deaf group in any cochlear region. These findings indicated that the efficacy of NT gene therapy diminished with increasing durations of deafness leading to reduced benefits in terms of SGN protection. Clinically, there remains a window of opportunity in which NT gene therapy can provide ongoing trophic support for SGNs.

Effects of localized neurotrophin gene expression on spiral ganglion neuron resprouting in the deafened cochlea.

A cochlear implant may be used to electrically stimulate spiral ganglion neurons (SGNs) in people with severe sensorineural hearing loss (SNHL). However, these neurons progressively degenerate after SNHL due to loss of neurotrophins normally supplied by sensory hair cells (HCs). Experimentally, exogenous neurotrophin administration prevents SGN degeneration but can also result in abnormal resprouting of their peripheral fibers. This study aimed to create a target-derived neurotrophin source to increase neuron survival and redirect fiber resprouting following SNHL. Adenoviral (Ad) vectors expressing green fluorescent protein (GFP) alone or in combination with brain-derived neurotrophic factor (BDNF) or neurotrophin-3 (NT3) were injected into the cochlear scala tympani or scala media of guinea-pigs (GPs) deafened via aminoglycosides for 1 week. After 3 weeks, cochleae were examined for gene expression, neuron survival, and the projection of peripheral fibers in response to gene expression. Injection of vectors into the scala media resulted in more localized gene expression than scala tympani injection with gene expression consistently observed within the partially degenerated organ of Corti. There was also greater neuron survival and evidence of localized fiber responses to neurotrophin-expressing cells within the organ of Corti from scala media injections (P < 0.05), a first step in promoting organized resprouting of auditory peripheral fibers via gene therapy.

Polypyrrole-coated electrodes for the delivery of charge and neurotrophins to cochlear neurons.

Sensorineural hearing loss is associated with gradual degeneration of spiral ganglion neurons (SGNs), compromising hearing outcomes with cochlear implant use. Combination of neurotrophin delivery to the cochlea and electrical stimulation from a cochlear implant protects SGNs, prompting research into neurotrophin-eluting polymer electrode coatings. The electrically conducting polypyrrole/para-toluene sulfonate containing neurotrophin-3 (Ppy/pTS/NT3) was applied to 1.7 mm2 cochlear implant electrodes. Ppy/pTS/NT3-coated electrode arrays stored 2 ng NT3 and released 0.1 ng/day with electrical stimulation. Guinea pigs were implanted with Ppy/pTS or Ppy/pTS/NT3 electrode arrays two weeks after deafening via aminoglycosides. The electrodes of a subgroup of these guinea pigs were electrically stimulated for 8 h/day for 2 weeks. There was a loss of SGNs in the implanted cochleae of guinea pigs with Ppy/pTS-coated electrodes indicative of electrode insertion damage. However, guinea pigs implanted with electrically stimulated Ppy/pTS/NT3-coated electrodes had lower electrically-evoked auditory brainstem response thresholds and greater SGN densities in implanted cochleae compared to non-implanted cochleae and compared to animals implanted with Ppy/pTS-coated electrodes (p<0.05). Ppy/pTS/NT3 did not exacerbate fibrous tissue formation and did not affect electrode impedance. Drug-eluting conducting polymer coatings on cochlear implant electrodes present a clinically viable method to promote preservation of SGNs without adversely affecting the function of the cochlear implant.