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A.V. Das, G.V. Hegde, K. Mallya, I. Ahmad; Wnt Signaling Regulates the Differentiation of Retinal Stem Cells/Progenitors Into RGCs . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3232.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Wnt signaling is known to modulate multiple developmental events, such as patterning, cell proliferation, self renewal and cell fate determination. Various components of the canonical Wnt signaling pathway are expressed in the developing retina and cultured retinal stem cells/progenitors. We have previously shown the involvement of Wnt signaling in the maintenance of retinal progenitors (Das et al., 2004). Here, we present evidence that Wnt signaling regulates the differentiation of retinal stem cells/progenitors into RGCs. Methods: We examined the temporal expression of the components of canonical Wnt pathway and its modulators in the retina by RT–PCR analysis and also by in situ hybridization. To study the involvement of Wnt signaling in the regulation of RGC differentiation in vitro, the signaling was perturbed in the E14 neurospheres, using Wnt3a/Wnt2b/Fzd8CRD/Lef1 siRNA, and cells were analyzed for the expression of RGC regulators and markers. The accentuation/attenuation of canonical Wnt pathway was determined by measuring levels of cytosolic b–catenin and Lef1. Finally, to understand the involvement of Wnt signaling in the spatial differentiation of RGCs, we exposed the E14 whole retinal explants to Wnt3a/Wnt2b/Fzd8CRD/Lef1 siRNA under conditions which promote RGC differentiation and the expression of RGC regulators and markers was analyzed. Results: Components belonging to the canonical Wnt pathway (i.e., Wnt proteins, Frizzled receptors and Lef1) and their modulators (Sfrps and Dkks) were expressed in the retina. Their involvement in retinal development was reflected by their temporal pattern of expression during retinal histogenesis. The activation of Wnt signaling promoted the cell proliferation and the attenuation of Wnt signaling promoted differentiation of the retinal stem cells/progenitors. Attenuation of the Wnt signaling using Lef1 siRNA resulted in the decrease in the Lef1 expression followed by a significant increase in the expression of RGC markers, Brn3b and RPF1. Conclusions: Taken together, our data suggest that the canonical Wnt pathway play an important role in the regulation of the cell fate determination of retinal stem cells/progenitors, specifically into RGCs.
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