Regeneration of Retinal Ganglion Cells Induced by suppression of Rho–Associated Protein Kinase

Abstract

Abstract: : Purpose: To elucidate the role of Rho–associated coiled coil–forming protein kinase (ROCK) on neurite outgrowth in purified retinal ganglion cells (RGCs) and on axonal regeneration of axotomised RGCs into a peripheral nerve graft in the rat eye. Methods: Rat RGCs were purified from dissociated retinal cells by panning method and cultured in a medium containing neurotrophic factors. Neurite outgrowth of RGCs was observed by microscope after cultivation with a specific ROCK inhibitor, Y39983. As an in vivo study, autologous sciatic nerve was transplanted to the axotomised optic nerve stump of rat. Six weeks after transplantation, a fluorescent dye was applied to the grafted sciatic nerve to retrogradely label the RGCs with extended axons in the graft. Intravitreal injection of Y39983 was administrated and the number of retrogradely labeled RGCs was estimated by microscopic observation of retinal wholemount preparations. Results: The ROCK inhibitor increased and elongated neurites in isolated RGCs. Results of nerve transplantation model showed that the density of retrogradely labeled RGCs was increased by the ROCK inhibitor in eyes whose optic nerve has been cut and sciatic nerve graft was used to replace the cut optic nerve. Number of ganglion cells which were retrogradely labeled with the dye significantly increased in animals treated with the ROCK inhibitor. Conclusion: The present data demonstrate that ROCK inhibitor promoted neurite outgrowth of purified RGCs and significantly enhanced the regeneration of RGCs with axons into peripheral nerve graft used to replace the cut optic nerve. Thus, the regeneration of nerve fiber enhanced by Y39983 in RGCs provides a possibility of therapy for degenerated diseases of RGCs, such as glaucoma and retinopathy.

Keywords: retina • retinal culture • regeneration