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Victor Liao, Narine Harutyunyan, Jennifer Aparicio, David Cobrinik, Thomas C Lee; Evaluation of Human Embryonic Stem Cell-Derived Retina as a Potential Retinoblastoma Model. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2984.
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© ARVO (1962-2015); The Authors (2016-present)
Retinoblastoma is a childhood malignancy initiated by the loss of Rb function during human retinal development. Mouse models do not accurately mimic retinoblastoma since Rb1 loss alone does not induce retinal tumors, and retinal tumors generated by combined loss of Rb and Rb-related proteins have a distinct phenotype. Human embryonic stem cells (hESCs) are a potential source of immature retinal tissue that could be used to model retinoblastoma if it exhibits developmental properties similar to human fetal retina. Our objective is to characterize hESC-derived retina, particularly with respect to Rb expression, to ascertain its potential to accurately model retinoblastoma pathogenesis.
hESC-derived retinas were grown in 3-dimensional cultures initiated by reaggregation of dissociated hESC colonies into spheres which were differentiated using small molecules and matrigel as described (Nakano et al 2012). hESC-derived retinas were analyzed by immunofluorescence staining of cryosections after various times of differentiation.
Analogous to retinal tissue developing in vivo, retinal progenitor cells (RPCs) of the hESC-derived retinas were Chx10+,Ki67+, and Pax6+, underwent interkinetic nuclear migration, and prominently expressed Rb. The RPCs also differentiated into retinal cells that were appropriately positioned within stratified retinal layers and were produced in the characteristic sequence seen in vivo. For example, detection of Brn3+ ganglion cell precursors preceded that of CRX+ photoreceptor precursors. Likewise, cone markers (cone arrestion, L/M opsin) emerged prior to rod markers (NRL). Although CRX+ cone precursors expressed L/M opsin and cone arrestin, and exhibited structures resembling inner segments and rudimentary outer segments, Rb was not detected in these cells, at least up to 120 days in culture, the latest age examined. In contrast, maturing human fetal cones express high levels of Rb.
hESC-derived retinas recapitulated many processes of normal retinal development, including production of RPCs and differentiated retinal cells in stratified layers. RPCs in hESC-derived retinas exhibited high levels of Rb similar to what is seen in human fetal retina. However, the retinas failed to induce Rb expression in maturing cone precursors, bringing into question whether they sufficiently simulate in vivo retinal maturation to serve as a retinoblastoma model.
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