Transplantation of conditionally immortal auditory neuroblasts to the auditory nerve

Abstract

Cell transplantation is a realistic potential therapy for replacement of auditory sensory neurons and could benefit patients with cochlear implants or acoustic neuropathies. The procedure involves many experimental variables, including the nature and conditioning of donor cells, surgical technique and degree of degeneration in the host tissue. It is essential to control these variables in order to develop cell transplantation techniques effectively. We have characterized a conditionally immortal, mouse cell line suitable for transplantation to the auditory nerve. Structural and physiological markers defined the cells as early auditory neuroblasts that lacked neuronal, voltage-gated sodium or calcium currents and had an undifferentiated morphology. When transplanted into the auditory nerves of rats in vivo, the cells migrated peripherally and centrally and aggregated to form coherent, ectopic 'ganglia'. After 7 days they expressed beta 3-tubulin and adopted a similar morphology to native spiral ganglion neurons. They also developed bipolar projections aligned with the host nerves. There was no evidence for uncontrolled proliferation in vivo and cells survived for at least 63 days. If cells were transplanted with the appropriate surgical technique then the auditory brainstem responses were preserved. We have shown that immortal cell lines can potentially be used in the mammalian ear, that it is possible to differentiate significant numbers of cells within the auditory nerve tract and that surgery and cell injection can be achieved with no damage to the cochlea and with minimal degradation of the auditory brainstem response.

Cells transplanted onto the surface of the glial scar reveal hidden potential for functional neural regeneration

“…Animal experiments were conducted in accordance with the Guidelines for Animal Experiments at Kyoto University. Exposing of the auditory nerve of male Sprague-Dawley rats (P7-8 wk) was performed as previously reported (7,8,10,11). For compression, a nerve hook (tip diameter: 0.15 mm, No.…”

“…Temporal bones were obtained and processed as previously reported (7,8,10,11). The section angle was chosen so that the auditory nerve trunk, PVCN, and DCN were simultaneously included in one section as fully as possible by reference to figures 1 and 2 in Hackney et al (56).…”

“…We developed an in vivo model of the chronic gliotic environment in clinical patients by compressing the rat auditory nerve without breaching the fluid spaces that contain the sensory structures inside the cochlea (7)(8)(9)(10). In this model, spiral ganglion cells (the auditory neurons) degenerate selectively, but hair cells are preserved both morphologically and functionally (7,8,10,11). We then waited 5 wk before cell transplantation to allow the formation of a glial scar with the progressive degeneration of auditory neurons.…”

“…We used a murine auditory neuroblast cell line for transplantation (US/VOT-N33) (12). This cell line was derived from the ventral otocyst (inner ear anlage) of a mouse embryo at embryonic day ED10.5 and was selected because it expresses key markers for auditory sensory neurons and differentiates with the appropriate bipolar morphology in the ear both in vitro and in vivo (11,12).…”

The Development of a Stem Cell Therapy for Deafness

“…This area is highly porous, and may provide windows big enough for transplanted cells to migrate into Rosenthal's canal. However, the size of transplanted cells would be a major factor limiting cell migration through these bony pores (Sekiya et al 2007 ). A report by Parker et al ( 2007 ) described general distribution of cells throughout different cochlear compartments when using a perfusion system (Parker et al 2007 ).…”