Viral-mediated gene transfer in the cochlea.

Gene transfer is an exciting new tool in medical therapy and scientific investigation, but only very recently has it begun to be developed in the auditory system. This paper describes in vivo and ex vivo experiments using an adenoviral vector (Ad. RSVntlacZ), which is a replication-deficient virus based on a human adenoviral (serotype 5) genomic backbone. The in vivo experiments demonstrate successful gene transfer into multiple types of cochlear cells. We observed a relatively efficient transduction, several weeks of sustained transgene expression and an absence of major lethal cytotoxicity in spiral ganglion and epithelial cells of the cochlea in adult animals. The ex vivo experiments were performed using fibroblasts transduced in vitro with Ad. RSVntlacZ. Two weeks after inoculation of the fibroblasts into the perilymph, we observed transplanted fibroblasts, which were adherent to the lining of the perilymphatic spaces, and were expressing the lacZ transgene. We speculate that, as the genetic basis of degenerative cochlear diseases is characterized on a mutational level, transgene expression will allow us to test hypotheses regarding the effects of specific genes on cochlear cell biology. Gene transfer will not only increase our understanding of the pathophysiology of hearing loss, but also may provide gene therapy for disease.

Presynaptic terminals in hyaline cells of normal and overstimulated chick inner ears.

Hyaline cells are non-sensory epithelial cells of the vibrating part of the basilar membrane of chicks; they receive an extensive efferent innervation. Although these anatomical features suggest roles in auditory transduction, very little is known about the function of these cells. One possible way to understand function is by lesion experiments. We used synapsin-specific antibodies to study changes that occur in the pattern of efferent innervation in hyaline cells after lesion of the sensory epithelium induced by acoustic overstimulation. We found only small changes in hyaline cells after such trauma. These included a small increase in size and a small decrease in density of nerve terminals on hyaline cells. This suggests that hyaline cells and their nerve terminals are less susceptible to acoustic trauma than hair cells. Using neurofilament-specific antibodies we found little or no trauma-induced change in the density of nerve fibres that cross the basilar papilla and reach the hyaline cell region. This finding suggested that trauma to the hair cells does not necessarily lead to changes in the efferent fibres that cross the papilla and extend into the hyaline cell region. Using the trauma and the morphological parameters studied here, it appears that a moderate lesion in the hair cell region in the avian inner ear does not influence the hyaline cells or their innervation.

Adenoviral-mediated gene transfer into guinea pig cochlear cells in vivo.

Loss of ganglion cells is a common and irreversible complication of hair cell loss in the cochlea. Gene transfer could potentially be used to prevent this neuronal degeneration and other pathologies in the cochlea. Human adenoviruses should provide a feasible gene transfer vehicle for transducing the quiescent cochlear neurons and organ of Corti epithelium. We now describe in vivo experiments in which a replication-deficient adenoviral vector, Ad.RSVntlacZ was injected into the perilymphatic fluid of six normal guinea pigs. Postoperative recovery of animals was complete. Inner ear tissues were assessed for histology and for presence of lacZ-positive cells 1 or 2 weeks after the injection. A large number of blue (lacZ-positive) cells were observed in the neural, epithelial and connective tissues of the cochlea. In four ears spiral ganglion cell infection exceeded 50%, throughout the length of the cochlear spiral. No major pathology was detected in the organ of Corti and other cochlear tissues, and no infection was present in the vestibular tissues or the contralateral cochlea. Immunocytochemical assessment of T cells revealed an increased in the number of lymphocytes in the connective tissue lining the perilymphatic spaces. We conclude that efficient gene transfer into multiple types of cochlear cells in vivo can be achieved without major morphological signs of pathology or toxicity.