Abstract
The availability of in vitro models of the human retina in which to perform pharmacological and toxicological studies is an urgent and unmet need. An essential step for developing in vitro models of human retina is the ability to generate laminated, physiologically functional, and light‐responsive retinal organoids from renewable and patient specific sources. We investigated five different human‐induced pluripotent stem cell (iPSC) lines and showed a significant variability in their efficiency to generate retinal organoids. Despite this variability, by month 5 of differentiation, all iPSC‐derived retinal organoids were able to generate light responses, albeit immature, comparable to the earliest light responses recorded from the neonatal mouse retina, close to the period of eye opening. All iPSC‐derived retinal organoids exhibited at this time a well‐formed outer nuclear like layer containing photoreceptors with inner segments, connecting cilium, and outer like segments. The differentiation process was highly dependent on seeding cell density and nutrient availability determined by factorial experimental design. We adopted the differentiation protocol to a multiwell plate format, which enhanced generation of retinal organoids with retinal‐pigmented epithelium (RPE) and improved ganglion cell development and the response to physiological stimuli. We tested the response of iPSC‐derived retinal organoids to Moxifloxacin and showed that similarly to in vivo adult mouse retina, the primary affected cell types were photoreceptors. Together our data indicate that light responsive retinal organoids derived from carefully selected and differentiation efficient iPSC lines can be generated at the scale needed for pharmacology and drug screening purposes. stem cells 2018;36:1535–1551
Highlights
Visual impairment affects 285 million people worldwide, with 90% of cases arising from chronic diseases [1]
This variability was corroborated by the gene expression analysis which indicated that the induced pluripotent stem cell (iPSC) lines which were more efficient at giving rise to retinal organoids (WT1 and WT2) had higher expression of photoreceptor progenitor (VSX2), photoreceptor precursor (CRX and RCVRN), and retinal ganglion cell (MATH5) markers compared with the less efficient iPSC lines (Fig. 1C)
No apparent retinal-pigmented epithelium (RPE) was observed in the organoids from any of the five iPSC lines at this time point, the expression of RPE marker (MITF) was higher in the two iPSC lines which were more efficient at giving rise to RPE (WT3 and age-related macular degeneration (AMD)) during the course of differentiation
Summary
Visual impairment affects 285 million people worldwide, with 90% of cases arising from chronic diseases [1]. 80% arise in the aged population, and with a continued increase in life expectancy, the numbers affected by vision loss are set to rise [2, 3]. As the retina does not readily regenerate, diseases such as age-related macular degeneration (AMD), retinitis pigmentosa (RP), and glaucoma cause cell loss and result in irreversible profound visual impairment. For treatments to be developed, in vitro models of human retina that encompass all the retinal cell types and which are electrophysiologically responsive to light are required to screen novel substances for pharmacological and toxicological effects. Most drug studies are currently performed in vivo using rodent models, but this approach is far from optimal because there are fundamental structural and functional differences between rodent and human retina.
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