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I Ahmad, J James, AV Das; Late Retinal Stem Cells/Progenitors Can Give Rise to Early Born Retinal Neurons . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3685.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The generation of retinal cells involves two different neural stem cell/progenitor populations. The early neural stem cells/progenitors give rise to retinal ganglion cells (RGCs), cone photoreceptors and horizontal cells between E11 and E14 (early neurogenesis) and the late neural stem cells/progenitors generate the majority of rod photoreceptors, amacrine cells, bipolar cells and the Muller glia E18 onward (late neurogenesis). We wanted to know if the late neural stem cells/progenitors retain the potential to give rise to early born neurons, particularly RGCs. Methods: Neural stem cells/progenitors, isolated from E18 retina, were expanded in vitro in the presence of EGF and FGF2 as previously described (Ahmad et al., 1999 Brain Res. 831:1-10). Cells were exposed to BrdU to tag the dividing cells. BrdU-tagged neural stem cells/progenitors were co-cultured with excess of cells from E3 chick retina and forebrain. RGCs differentiate in E3 chick retina, therefore, this stage of retina was presumed to contain factors that promote RGC differentiation. The acquisition of RGC phenotype by BrdU-tagged neural stem cells/progenitors was ascertained by immunocytochemical analysis using antibodies against RPF1, Brn3B and Islet1. The RGC differentiation was corroborated by RT-PCR analyses of transcripts corresponding to RPF1, Brn3B and Islet1 genes. In order to understand the underlying mechanism of RGC differentiation, expression of bHLH factors, particularly Ath5, was studied. Results: Differentiation of BrdU-tagged stem cells/progenitors by mitogen withdrawal and addition of serum resulted into cells that expressed markers corresponding to late born neurons such as rhodopsin (rod photoreceptors), syntaxin (amacrine cells) and PKC (bipolar cells). However, BrdU cells expressing RGC markers were not detected either in proliferative or differentiation conditions. When BrdU-tagged stem cells/progenitors were co-cultured with E3 chick retina a small proportion of these cells were found expressing immunoreactivities corresponding to RPF1, Brn3B and Islet1. Results of immunocytochemical analyses were corroborated by RT-PCR on stem cells/progenitors co-cultured with retinal cells across a membrane barrier. Conclusion: A subset of late retinal stem cells/progenitors retain the potential to give rise to early born retinal neurons. They can differentiate into RGCs when exposed to an evolutionarily conserved diffusible factor(s) present in the embryonic retina. Support: NEI
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