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S. Bhattacharya, A.V. Das, K.H. Cowan, I. Ahmad; Role of ABCG2 Transporter in Notch and CKIT-Mediated Maintenance of Retinal Stem Cells . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1668.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We have demonstrated that the retinal stem cells/progenitors can be prospectively identified as side population (SP) cells using the Hoechst dye efflux assay. It has been observed that the preferential exclusion of Hoechst dye by stem cells is mediated by ATP-binding cassette transporter, abcg2. It is thought that abcg2 may provide protection to stem cells by excluding substances, potentially harmful to DNA. Therefore, abcg2 transporter besides being a marker of stem cells may have important function(s) in their maintenance. It is known that signaling mediated by Notch and c-Kit RTK is involved in the maintenance of stem cells. We wanted to know if c-Kit and Notch receptors mediated maintenance of stem cells involved abcg2 gene. Methods: Neural stem cells/progenitors were enriched from embryonic day18 (E18) retina as SP cells by Hoechst dye efflux assay as previously described (Bhattacharya et al, 2002, ARVO abstract). To determine the involvement of abcg2 in the maturation of E18 stem cells/progenitors, levels of abcg2 transcripts were analyzed in response to perturbation of Notch and c-Kit-RTK signaling. Perturbation of Notch signaling was achieved by using DAPT, a γ-secretase inhibitor and expression of constitutively activated Notch intracellular domain (NICD). Perturbation of c-Kit RTK signaling was achieved by stem cell factor (SCF) or c-Kit antibody. The functional analysis of abcg2 gene was carried out using the abcg2 retroviral construct. Results: Transcripts corresponding to abcg2 gene were expressed in E18 retinal SP cells. The levels of expression were decreased in differentiation condition as compared to those in proliferation condition. abcg2 transcripts were detected throughout retinal histogenesis and disappeared at postnatal day 6 (PN6), a stage when specification of retinal cells is almost over, suggesting an association of abcg2 expression with progenitor population in vivo. Forced expression of abcg2 expanded the SP cell population and its expression was influenced in response to perturbation of Notch and c-Kit signaling. Conclusions: ABCG2, besides being a marker of retinal stem cells, may be a factor participating in the maintenance of retinal stem cells by regulating their homeostasis.
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