Modeling Human Retinal Development with Patient-Specific Induced Pluripotent Stem Cells Reveals Multiple Roles for Visual System Homeobox 2

Abstract

Human induced pluripotent stem cells (hiPSCs) have been shown to differentiate along the retinal lineage in a manner that mimics normal mammalian development. Under certain culture conditions hiPSCs form optic vesicle-like structures (OVs), which contain proliferating progenitors capable of yielding all neural retina (NR) cell types over time. Such observations imply conserved roles for regulators of retinogenesis in hiPSC-derived cultures and the developing embryo. However, whether and to what extent this assumption holds true has remained largely uninvestigated. We examined the role of a key NR transcription factor, Visual System Homeobox 2 (VSX2), using hiPSCs derived from a patient with microphthalmia caused by an R200Q mutation in the VSX2 homeodomain region. No differences were noted between (R200Q)VSX2 and sibling control hiPSCs prior to OV generation. Thereafter, (R200Q)VSX2 hiPSC-OVs displayed a significant growth deficit compared to control hiPSC-OVs, as well as increased production of retinal pigmented epithelium (RPE) at the expense of NR cell derivatives. Furthermore, (R200Q)VSX2 hiPSC-OVs failed to produce bipolar cells, a distinctive feature previously observed in Vsx2 mutant mice. (R200Q)VSX2 hiPSC-OVs also demonstrated delayed photoreceptor maturation, which could be overcome via exogenous expression of wildtype VSX2 at early stages of retinal differentiation. Finally, RNAseq analysis on isolated hiPSC-OVs implicated key transcription factors and extracellular signaling pathways as potential downstream effectors of VSX2-mediated gene regulation. Our results establish hiPSC-OVs as versatile model systems to study retinal development at stages not previously accessible in humans, and support the bona fide nature of hiPSC-OV-derived retinal progeny.

An Advocacy for the Use of 3D Stem Cell Culture Systems for the Development of Regenerative Medicine: An Emphasis on Photoreceptor Generation

“…Taking advantage of this multi-step protocol combining suspension and adherent cell cultures, Meyer et al confirmed the possibility of employing hES cells able to produce an enriched population of photoreceptor cells and RPE at the earliest stage of retinal specification that closely mimics the key developmental stages of retinogenesis. Later on, other papers published by the same group validated these results [59,60].…”

“…Taking advantage of this multi-step protocol combining suspension and adherent cell cultures, Meyer et al confirmed the possibility of employing hES cells able to produce an enriched population of photoreceptor cells and RPE at the earliest stage of retinal specification that closely mimics the key developmental stages of retinogenesis. Later on, other papers published by the same group validated these results [59,60].In 2011, the first striking protocol using a 3D culture system during the whole experiment has been developed by the Sasai's group to differentiate mouse ES into optic cups forming well-structured retina tissues [61] (Figure 1). This masterwork has been briefly followed by a second one exclusively focused on human ES [62].…”

iPS Cells for Modelling and Treatment of Retinal Diseases

“…Upon further differentiation, photoreceptor-like cells and RPE were derived from the vesicle-like structures. Similar optic vesicle-like structures have also been generated from lymphocyte-iPSC [4,36]. Zhong et al recently reported three-dimensional laminated retinal cups generated from human iPSC with distinct populations of neural retinal cells interacting through synaptic junctions and photoreceptor cells capable of forming outer segment discs and responding to light [57].…”

Ips Progression a Decade Devoted

“…These 3D functional organoids have improvised our understanding of early human development and improved drug screening methodologies organoids including mini gut, mini heart (repopulating decellularized mice heart through iPS), mini brain (developing cortex, central telencephalon and choroid plexus), small lungs, mini eyes (optic vesicle like structures) have been formulated for disease remodeling purposes [82][83][84][85][86][87][88][89].…”