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K. Wallace, J. S. Meyer, A. D. Verhoeven, E. E. Capowski, L. S. Wright, D. M. Gamm; Factors Predicting Retinal Competence in Human Pluripotent Stem Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):727.
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© ARVO (1962-2015); The Authors (2016-present)
Human induced pluripotent stem (iPS) cell lines generate retinal progeny with varying efficiencies. We sought to identify factors expressed during early differentiation of human pluripotent stem cells that correlate with competency to produce retinal cell types.
Human iPS and ES cells were differentiated along the retinal lineage using a previously published protocol (1). Several human ES and iPS cell lines (vector and vector-free) were examined at day 10 of differentiation by quantitative PCR, immunocytochemistry and Western blot analysis for the presence of early anterior neuroepithelial and retinal markers and expression of Wnt, BMP and Nodal signaling components. Cultures were then treated with inhibitors of these pathways (Dkk1, Noggin and SB431542, respectively) to determine their effects on neural and retinal differentiation.
Consistent with previous observations, human iPS cell lines varied in their capacity to adopt a primitive retinal fate. ES and iPS lines that endogenously expressed high levels of Dkk1 and Noggin during early differentiation were the most efficient at producing primitive neuroepithelial/eye field cell populations (Pax6+, Rx+ and Lhx2+) at day 10. These populations in turn generated the largest populations of Vsx2+ neural retinal progenitors at day 20. Conversely, lines that did not express high levels of endogenous Dkk1 and/or Noggin failed to yield significant percentages of retinal cell types. Exogenous addition of Dkk1, Noggin and/or SB431542 improved retinal differentiation potential to a limited extent in some lines.
Retinal competence in human ES and iPS cell lines is predicted by early endogenous Dkk1 and Noggin expression. This information may be useful for screening iPS cell lines and optimizing their ability to produce retinal cell types.1. Meyer et al. (2009) PNAS 106:16698.
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