Identification and purification of retinal stem cells by the expression pattern of specific genes.

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Neural retina is an important target organ for regenerative medicine, and isolation and expansion of retinal progenitor cells are critical issues from both scientific and clinical views. However, the characters of the immature retinal cells are not elucidated because of the lack of prospective approach to identify retinal progenitor cells.We screened the expression pattern of cell surface proteins in mouse immature retina by flow cytometer using about 120 antibodies against different membrane proteins. Among them, 20 antibodies recognized sub-populations of immature retina, and we examined the proliferation and differentiation abilities of purified those sub-populations of retina.We identify the SSEA-1 (CD15), which is expressed in ES cells and neural stem cells, as a novel surface antigen to define immature retinal progenitor cells. SSEA-1 positive cells are in the peripheral region of retina of the E17 embryo and then dramatically disappeared along with retinal development. SSEA-1 st … More rong positive cells were Ki-67 antigen positive and had prolonged proliferation activities than that of SSEA-1 negative cells in reaggregation culture. Moreover, differentiation of SSEA-1 cells into late born retinal subtypes took longer period, suggesting that these cells are at more immature stage than SSEA-1 negative cells.To obtain new cell markers defining retinal stem cells, we took an approach of proteomics. Membrane fractions were purified from embryonic and adult retina, and the profile of the expression pattern of membranous proteins were examined by shotgun analyses on a nanoflow LC-MS/MS system. Several proteins which were expressed specifically in embryo or adult retina were identified, and biological functions of these proteins are currently examined by si-RNA.Taken together, it is the first report showing the finding of a surface marker to determine regionally restricted immature subset of progenitor cells of neural retina and may serve an excellent tool for clinical use of retinal progenitor cells. Less