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R. Kohno; Characerization of neurosphere–like populations derived from ciliary body and peripheral retina of adult mice . Invest. Ophthalmol. Vis. Sci. 2004;45(13):5382.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Neural stem/ progenitor cells from central nervous system (CNS), as well as from ciliary body (CB) and peripheral retina (PR), are likely to be a promising candidate for a cell source of the regenerative therapy for neurodegenerative disorders. It is well–known that such populations proliferate in liquid culture as cell clusters, namely, so–called "neurospheres", however, little information is now available regarding the characteristics of PR–derived retinal neurospheres. The objective of the current study is to characterize retinal neurospheres, directly compared with those of CNS–derived neurospheres. Methods: Retinal neurospheres were isolated from the ciliary body and peripheral retina of adult C57BL/6 mice or nestin–GFP transgenic mice (6 weeks old, respectively), and cultured for 14 days in the presence of EGF and FGF–2. Ultrastructure and the process of sphere formation were examined by transmission electron microscopy (TEM) and Time–laps system, respectively, compared with those of CNS–derived neurospheres. Results: Unlikely to CNS–derived neurospheres, TEM demonstrated that retinal neurospheres were contacted to each other by desmosome, tight junction and adherens junction, findings similar to those seen in epithelial cells. Time–laps analysis revealed that adult CB and PR–derived spheres were grown by sphere–sphere interaction or involving scattered cells, resulted in neurosphere–like appearance. Termination of the cell cycle by aphidicolin did not significantly affect the growth and increase of number of the retinal neurospheres, until 2 days after treatment. No GFP positive cells was observed at the ciliary body and peripheral retina in the flat–mounted ocular tissue of nestin–GFP transgenic mice, however, flow–cytometry analysis revealed that approximately 3–6% GFP positive cells appreared in neurospheres after 14 days cultivation, suggesting that the formation of retinal spheres from adult mice promoted cells to dedifferentiation– neurodifferentiation. Conclusions: These results suggest that neurosphere–like fomation of CB and PR cells from adult retina, so–called retinal neurospheres, is likely to be essentially different from the neurospheres from CNS or embryonic retina, in views of ultrastructure as well as their growing process. The current results also suggest that immature neural cells may be obtained by sphere formation from terminally differentiated cells of adult ocular tissue, suggesting the potency of possible cell source for regenerative medicine to treat neurodisorders.
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