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Y Arsenijevic, N Taverney, C Kostic, L Zografos, D Schorderet, F Munier; Adult Human Eye Sclera Cells Have in Vitro Capacity to Generate Neurons Over Time . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3899.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Isolation of adult human cells able to produce neurons is of particular interest to have an in vitro model of human neurogenesis and to provide cells for neural tissue repair. Several eye tissues have as origin the neuroectoderm: the retina including RPE cells, the iris, and the melanocytes located in the choroids and the sclera. Previous observations in the sixties suggest that some eye melanoma might generate neurons. In this study we investigate the possibility of generating neurons in vitro from human eye-derived melanocytes. Methods: Eyes from deceased donors constitute our tissue source. During dissection, melanocytes are scratched off the sclera to produce high melanocyte density cultures. Cells were plated in the presence of EGF and FBS. Results: After one week in growth medium, melanocytes proliferated in clusters generating a monolayer of pigmented and non-pigmented cells. The first step of growth took one month. Cells were transferred on laminin substrate to induce cell differentiation in presence of various neurotrophic factors. 0,3% to 5% (depending on the stimulations) of total cells presented typical neuronal morphology and expressed b-tubulin, an early neuronal marker. Throughout passages (6 to 11) and donors (n=7), we observed a similar percentage of neurons. RT-PCR analysis confirmed the presence of other neuronal genes like calbindin and neurofilament. To determine whether these neurons were produced in vitro via quiescent eye progenitor recruitment or by a transdifferentiation process, we investigated the presence of precursors either in adult choroid slices, after tissue dissociation, or in our cultures. We observed the appearance of the expression of the nestin precursor marker in culture, as well as the expression of hes-1; after differentiation at early passages, neurons contained pigments. These observations contrast with the absence of neurons, and neural precursors in in vivo sclera and choroids. Altogether these features suggest for a transdifferentiation process of a population of melanocytes. Conclusions: The cue identification allowing cell proliferation and leading to a neuronal phenotype from a differentiated cell should have potent implications for identification of the genes involved in transdifferentiation and cell renewal. Supported by the Swiss National Science Foundation PNR46.
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