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L. S. Wright, I. Pinilla, A. T. Steinhauer, R. L. Shearer, J. S. Meyer, E. E. Capowski, D. M. Gamm; Chx10 Expression Is Not Maintained in Long-Term Prenatal Human Retinal Neurosphere Cultures. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1265.
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To compare Chx10-positive cell populations in tissue sections obtained from human prenatal eyes of increasing gestational age with age-matched retinal progenitor cultures.
Intact postmortem human prenatal eyes from approximately 60 to110 days of gestation were obtained from the Birth Defects Laboratory (University of Washington), prepared for immunohistochemical analysis and immunostained with markers of neural and retinal proliferation and differentiation. Retinal neurosphere cultures were generated from neurosensory retina from postmortem eyes of identical gestational age, maintained in serum-free media supplemented with FGF2 and EGF for more than one month, prepared for immunocytochemical analysis and immunostained for those same markers.
Throughout the range of examined ages, Chx10 immunopositive cells were exclusively confined to the zone of proliferative progenitors within the outer retina. This population co-localized with well-established neural progenitor markers such Ki67, Sox2 and nestin, but no association was seen with post-mitotic retinal neuronal markers such as Pax6, HuC/D, recoverin or ß III tubulin. A population of cells with similar phenotype was present in early neurosphere cultures established from age-matched tissues. However, the Chx10+/Ki67 population was lost by one month in vitro with a concomitant decrease in neurogenesis, whereas a glial-restricted Chx10-/nestin+/Sox2+ cell became the predominant proliferating cell type in long-term cultures.
A population of Chx10+ cells with proliferative capacity, as determined by Ki67 colocalization, was confined as early as 60 days gestation to the outer retina and can be established in vitro. These Chx10+/Ki67+ cells were not capable of extended propagation; instead, a Chx10- glial-restricted progenitor became the principal cell type in long-term culture.
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