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J Liu, X Zhao, I Ahmad; Retina-Specific Potential of Embryonic Stem (ES) Cell-Derived Neural Progenitors . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3689.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Embryonic stem cells are rich and renewable source of neural progenitors. We have shown previously that ES cell-derived neural progenitors have the potential to acquire retinal phenotypes (Zhao and Ahmad, 2001 ARVO Abstract 1066). We have studied these cells, generated by two different methods, in co:culture conditions and upon overexpression of photoreceptor regulating genes in order to improve the efficiency of their differentiation into retinal cell types. Methods: Mouse D3 ES cells were cultured and induced to generate neural progenitors using either retinoic acid (Bain et al., 1995 Dev. Bio. 168:342-357) or by ITSFn/FGF2 (Okabe et al., 1996 Mech. Dev. 59:89-102). ES cell-derived neural progenitors were co:cultured with excess of embryonic retinal cells. In addition, they were transfected with recombinant vector expressing Crx and Nrl, before co:culture. Results: Both ITSFn/FGF2- and RA- based methods caused neural induction in ES cells. However, the former generated more neural progenitors than the later (86.3% Vs 63%). A small subset of BrdU+, ES cell-derived neural progenitors expressed photoreceptor-specific markers, rhodopsin and IRBP, when co-cultured with retinal cells. The proportion of cells with photoreceptor-specific markers was higher in the ITSFn/ FGF2 group and increased from 0.7% to 2% when the duration of culture was extended from 5 days to 10 days. In addition, subsets of BrdU+ cells were detected that expressed retinal markers such as PKC, mGluR6, RPF1 and Islet1. However, these markers are also expressed in other brain regions. The proportion of BrdU+ cells, expressing one of these markers was considerably high than those expressing rhodopsin, at the end of 5th day of co:culture [ 1.3% (Islet1), 4% (RPF1), 6.4% (mGluR6) and 21.7% (PKC)]. Experiments are currently underway to analyze the effect of overexpression of Nrl and Crx on retina-specific differentiation of ES cell-derived neural progenitors. Conclusion: ES cell-derived neural progenitors can be induced to acquire retinal phenotypes, as ascertained by the expressing of photoreceptor specific markers. The efficiency of retinal differentiation improved with ITSFn/ FGF2-induced neural progenitors and by extending the duration of co:culture. Identification of conditions that promote retina-specific differentiation of ES cells will lead to a renewable source of retinal progenitors that can be used for understanding retinal development and for cell therapy to treat retinal degeneration. Supported by UNMC Dean's grant and Foundation Fighting Blindness.
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