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AV Das, J James, X Zhao, WB Thoreson, I Ahmad; Molecular Characterization And Differentiation Potential Of Ciliary Body-derived Neural Stem Cells . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3688.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The ciliary body (CB)-derived neural progenitors display several properties reminiscent of stem cells. They are multipotential and have the ability to self-renew. They are able to differentiate into neurons displaying retinal phenotypes. Our objective is to identify genes that regulate proliferation and differentiation, and to characterize conditions that promote retinal differentiation of CB-derived progenitors. Methods: Dissociates from pigmented CB of adult rats were cultured in serum-free medium containing EGF (20ng/ml) and FGF2 (10ng/ml) for 5-7 days to obtain neurospheres. Differentiation was induced by withdrawing EGF and adding 0.5% FBS into the medium. The differential gene expression in the proliferating and differentiated neurospheres was studied using microarray technology. Biotin-labeled cRNA probes were made and hybridized to Affimetrix rat U34A chips and data were analyzed by Affimetrix GeneChip algorithms. To induce retina-specific differentiation the neurospheres were cultured with or without E3 Chick (to promote differentiation of early born neurons) or PN1 retinal cells (to promote differentiation of late born neurons) followed by immunocytochemical and RT-PCR analyses. Results: Microarray analysis showed differential regulation of several classes of genes in proliferation and differentiation conditions. Some of the genes whose levels were differentially regulated in differentiation condition includes Pax6 (4.5 fold decrease), mitogenic regulatory gene 322 (3 fold decrease), syntaxin, (2 fold increase), brain derived neurotrophic factor (4.1 fold increase) and PSD-associated protein-2 gene (3.4 fold increase). A significant proportion of CB-derived progenitor cells expressed Brain 3B (RGC marker) when co-cultured with E3 chick retinal cells and RetP1 (a rod photoreceptor specific marker) when co-cultured with PN1 retinal cells. Conclusion: The differentiation of CB-derived neural progenitors involves distinct classes of genes and their differentiation along retinal lineage can be influenced by extracellular cues. Both early born and late born retinal neurons can be generated in response to distinct culture conditions. Supported by NEI, Nebraska Research Initiatives and Foundation Fighting Blindness
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