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M. Kayama, M. S. Kurokawa, Y. Ueda, H. Ueno, Y. Kumagai, C. Masuda, E. Takada, R. Tago, S. Ueno, N. Suzuki; Subretinal Transplantation of Embryonic Stem Cell Derived Neuroretinal Cells to Mice With Optic Nerve Injury. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3549.
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© ARVO (1962-2015); The Authors (2016-present)
Glaucoma is caused by irreversible damage of an optic nerve. Because an optic nerve is a bundles of retinal ganglion cell axons, retinal ganglion cell replacement therapy may become alternative modality for improving the visual acuity. Here, we tried to repair damaged optic nerves by subretinal transplantation of embryonic stem (ES) cell derived neuroretinal progenitor cells.
Mouse ES cells were transfected with pax6 gene, which was an essential transcription factor for early eye development. The pax6 transfected cells were cultured in the G418 containing media and limiting dilution culture was conducted to select cloned neural progenitors. The growing neural cells were used for in vitro characterization and for neuroretinal cell graft. Optic nerves of C57BL/6 female mice were injured 4 mm before optic chiasmata. The pax6 transfected cells were transplanted to subretinal layer of the eye with optic nerve injury. One month after the transplantation, the transplanted eyes were assessed by immunofluorescence, anterograde axon tracing and light reflex test.
We obtained cloned cell lines of the pax6 transfected cells which express neuroretina related molecules including nestin, neurofilament middle chain (NFM), Brn3, Thy1, protein kinase C, Crx, and opsin. These cells transplanted to subretinal layer of the optic nerve injured mice have kept alive in the grafted eye one month after the transplantation. The transplantedcells elongated axons and expressed NFM and Brn3. The pax6 transfected cells elongated axons across the optic chiasm to the superior colliculus (SC). The pax6 transfected cells connect the axons with recipient photoreceptor layer, accompanying synaptophysin expression. Cholera toxin-B (CTB) used as an axon tracer was injected into the grafted eyes. We found that CTB reached the SC, suggesting the neural connection was reconstructed between the cell transplanted retina and SC. The pax6 transfected cell transplanted mice with optic nerve injury restored light reflex from 21 days after the transplantation. In contrast, the PBS injected control mice did not restore the light reflex until 28 days after the transplantation.
Transplantationof ES cell derived neuroretinal progenitor cells may be applicable as a novel therapeutic approach for glaucoma.
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