Purchase this article with an account.
Dominic Shayler, Hardeep Pal Singh, Narine Harutyunyan, David Cobrinik; Altered RB and p27 expression patterns in hESC-derived versus fetal retinal cone-precursors. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3587.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Methods have recently been developed to generate retinal tissue from human embryonic stem cells (hESCs). These hESC-derived “retinal spheres” potentially could model aspects of human retinal development and disease more accurately than live animal models. For example, in human but not mouse retinas, maturing cone arrestin (CA)-positive fetal cone precursors have been shown to upregulate RB protein, MDM2 and MYCN, and to downregulate p27.These changes have been implicated in the human cone precursor response to RB loss in retinoblastoma development (Xu et al., 2014, Nature). To investigate whether hESC-derived cone precursors display similar features to fetal cone precursors, as might be needed in a retinoblastoma model, RB, MDM2, MYCN and p27 were examined in spheres of multiple ages.
H9 embryonic stem cells were differentiated into retinal tissue as described (Nakano et al. 2012). Retinal spheres were collected from multiple preparations, frozen sectioned and immunostained. p27, RB and CA expression was observed at days 27, 41, 60/69, 97 and 132, while MDM2 and CA expression was observed at the same ages as well as at day 123. Fetal retina of weeks 15 and 21 served as positive controls. All sections were imaged under confocal microscopy.
The earliest CA-positive cells appeared at day 41. Older spheres displayed greater numbers of CA-positive cells with increasing numbers on the apical surface. p27 was downregulated in these cells in fetal retina but not in spheres. RB was expressed at higher levels in cone precursors compared to other retinal neuron cell types in fetal retina, but was expressed at lower levels at all ages in hESC-derived tissue. MDM2 was expressed at higher levels in CA-positive cells than other retinal cell types both in retinal spheres and in fetal retina.
These data demonstrate that developing CA-positive cells do not share all protein expression patterns with fetal retina. MDM2 displayed similar expression in both systems, but RB was not prominent and p27 remained highly expressed in cone precursors as compared to other retinal cell types in the sphere system. This indicates that there is a difference in expression of proteins that are relevant to retinoblastoma development in the hESC-derived retina model.
This PDF is available to Subscribers Only