Purchase this article with an account.
Barbara Corneo, Timothy Blenkinsop, Patricia Lederman, Nathan Boles, Janmeet Saini, Gretchen Kusek, Sunita L D'Souza, Christoph Schaniel, Jeffrey Stern, Sally Temple; Generation Of iPSCs From Adult Proliferating RPE Stem Cells. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2225.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
We have recently shown that RPE (retinal pigment epithelium) cells isolated from adult cadaver donor eyes contains a subpopulation (around 3%) of cells that are highly proliferative, self-renewing and multipotent, defining the RPE stem cell (RPESC). RPESCs can proliferate and differentiate to produce a cobblestone RPE-like epithelium in vitro. In this study, we use adult RPESCs as a unique cell source for iPSC production. The resulting RPESC-derived iPSCs can be differentiated back into RPE cells, producing an essentially unlimited cell supply. Because the RPE is the disease-affected tissue in Age-Related Macular Degeneration (AMD), the leading cause of blindness in people over 50, these RPESC-derived cells may produce a superior iPSC-based model of AMD compared to iPSCs derived from other tissues.
We routinely isolate adult RPE from cadaver donor eyes of different ages and gender. iPSC were derived from human adult RPESCs using a Sendai virus-based reprogramming strategy. The resulting cells were characterized by standard protocols, including immunostaining, FACS, qPCR, in vitro differentiation into the three germ layers and teratoma formation in vivo.
Adult human RPESC-derived RPE cells were successfully used as donor tissue to derive iPSCs expressing pluripotency markers, including Tra-1-60, Nanog and SSEA4, and able to give rise to the three germ layers in vitro and in vivo. No residual donor RPE cells were detected in the iPSC cultures. Moreover, we obtained iPSCs from RPESCs isolated from AMD cadaver donor eyes.
We were able to obtain iPSC from elderly donor using a mix of 4 factors, demonstrating that we can successfully reprogram RPE from donors with ages ranging from 56 to 91 years old. To our knowledge, these are the first adult human CNS-derived iPSC, and the collection is particularly valuable as it includes elderly and AMD donors. In contrast to other sources of CNS stem cells in the hippocampal dentate gyrus and the subventricular zone, the RPESCs can be obtained relatively easily from live patients with minimal surgery, or from donor cadaver eyes. RPESC-derived iPSC could be used to reveal the molecular pathways involved with aging and that underlie AMD. The essentially unlimited RPESC-iPSC supply makes large-scale studies and drugs screens possible.
This PDF is available to Subscribers Only