Adult Ciliary Epithelial Cells, Previously Identified as Retinal Stem Cells with Potential for Retinal Repair, Fail to Differentiate into New Rod Photoreceptors

Abstract

The ciliary margin in lower vertebrates is a site of continual retinal neurogenesis and a stem cell niche. By contrast, the human eye ceases retinal neuron production before birth and loss of photoreceptors during life is permanent and a major cause of blindness. The discovery of a proliferative cell population in the ciliary epithelium (CE) of the adult mammalian eye, designated retinal stem cells, raised the possibility that these cells could help to restore sight by replacing lost photoreceptors. We previously demonstrated the feasibility of photoreceptor transplantation using cells from the developing retina. CE cells could provide a renewable source of photoreceptors for transplantation. Several laboratories reported that these cells generate new photoreceptors, whereas a recent report questioned the existence of retinal stem cells. We used Nrl.gfp transgenic mice that express green fluorescent protein in rod photoreceptors to assess definitively the ability of CE cells to generate new photoreceptors. We report that CE cells expanded in monolayer cultures, lose pigmentation, and express a subset of eye field and retinal progenitor cell markers. Simultaneously, they continue to express some markers characteristic of differentiated CE and typically lack a neuronal morphology. Previously reported photoreceptor differentiation conditions used for CE cells, as well as conditions used to differentiate embryonic retinal progenitor cells (RPCs) and embryonic stem cell-derived RPCs, do not effectively activate the Nrl-regulated photoreceptor differentiation program. Therefore, we conclude that CE cells lack potential for photoreceptor differentiation and would require reprogramming to be useful as a source of new photoreceptors. STEM CELLS

Adult Human Müller Glia Cells Are a Highly Efficient Source of Rod Photoreceptors

“…However, these cells exhibit reduced proliferation ability with only a minor fraction (below 10%) able to differentiate into photoreceptors. Moreover, latest studies provide evidence that these cells retain features of differentiated ciliary pigmented cells even after extensive in vitro culture and lack the potential to differentiate into mature photoreceptor cells [37,38]. Alternatively, important advances were achieved using human ES or iPS (induced-Pluripotent Stem Cells) for the generation of photoreceptor precursors.…”

Mobilizing Endogenous Stem Cells for Retinal Repair

“…A population of multipotent RPCs has been isolated from the ciliary epithelium (CE) of adult rodents and humans that shows the capacity to generate various retinal cell types in vitro [31][32] . However, their ability to proliferate and generate new retinal neurons, such as photoreceptors, appears to be limited in vivo [33][34] . Mitogens, including basic FGF, insulin, Wnt3a, and pigment epithelium-derived factor, are found to promote the proliferative potential of CE-derived RPCs [35][36][37][38][39] .…”

Sphere Formation Permits Oct4 Reprogramming of Ciliary Body Epithelial Cells into Induced Pluripotent Stem Cells

“…PCR and Real-time PCR were performed as previously described [1]. Previously published primer sets were used [1,9,18,19]. The mRNA level of target gene was normalized to the housekeeping gene HPRT.…”

“…However, the expression seems to be induced by growth factors in the medium. It remains unclear if these cells are bona fide quiescent retinal stem cells [8,9].…”