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T. Inoue, B. L. K. Coles, F. Tremblay, G. Prusky, Y. Tano, D. van der Kooy; Human Adult Retinal Stem Cells With Overexpression of OTX2/CRX and Suppression of CHX10 Can Differentiate Photoreceptor in vitro and in vivo. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4077.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal stem cells (RSCs) are present in the ciliary margin of the adult human eye and can give rise to all retinal cell types, but only a minority of these become photoreceptors when differentiated in vitro. Our purpose is to drive RSC progeny down the photoreceptor lineage.
To enhance the number of photoreceptors that differentiate from human RSCs, we transferred several critical genes shown to be important for retinal development into RSCs in vitro. These gene-modulated human RSC clonal sphere colonies were differentiated in vitro and transplanted into the mouse eye in vivo. Furthermore, these modurated-hRSCs were transplanted into the transducin-mutant mice eye to serve as a model for photoreceptor disease, and then function analysed using electrophysiological and behavioral tests.
Transfection with a CHX10-dominant negative construct (CHX10DN, 35.9%), an OTX2 (31.5%) constuct, or a CRX (26.8%) construct significantly increased the photoreceptor progeny of human RSCs compared to control transfections(11.8%). The optimal gene combination for enhancement of photoreceptor differentiation was the overexpression of OTX2 and CRX along with suppression of CHX10 (CHX10DN/OTX2/CRX, such that 60.6% of the progeny of single RSCs now became photoreceptors). Transplanted CHX10DN/OTX2/CRX transfected-human RSC progeny improved integration and differentiation to photoreceptors (91.3%) in the CD1 mouse eye compared to control (44.8 % photoreceptors). These human RSCs transplanted into the tranducin mutant eye showed functional recovery both in b-wave amplitudes in the electroretinogram and in spatial frequency sensitivity in a behaivioral test.
We found that the best gene combination for enhancement of photoreceptor differentiation from clonal human RSC progeny, was the overexpression of the genes OTX2 and CRX and the suppression of CHX10. The gene modulation of human RSC may provide a source of photoreceptor cells that can be used in the treatment of photoreceptor disease.
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