Modeling Late-Onset Retinal Degeneration with Human iPSCs: Insights into the Shared Pathogenesis of Macular Degenerations

Abstract

Purpose: Late-Onset Retinal Degeneration (L-ORD) is an autosomal dominant disorder caused by a S163R mutation in CTRP5. The disease is characterized by delayed dark adaptation, long anterior lens zonules, and drusen-like deposits in the sub-retinal pigment epithelium (RPE). The purpose of our study is to deduce the mechanism by which mutated CTRP5 causes retinal degeneration using RPE derived from induced pluripotent stem cells (iPSCs) specific to L-ORD patients and their healthy siblings.

Methods: iPSCs were generated from skin biopsies obtained from two affected and two unaffected siblings from a L-ORD family using Sendai virus based transfection of OCT3/4, SOX2, c-MYC, and KLF4 factors. iPSCs were characterized for expression of pluripotent markers by immunofluorescence and karyotyped for genetically stability. iPSCs were differentiated into RPE cells (iRPE) using a developmentally-guided differentiation protocol. The iRPE cells were authenticated by gene expression, immunofluorescence, TEM, and physiology.

Results: We successfully generated and authenticated iPSCs and iRPEs from a L-ORD family. We confirmed that patient iPSCs retained the S163R mutation in CTRP5 and control iPSCs retained the wild-type sequence. TEM images of healthy sibling- and L-ORD patient- iRPE confirmed their epithelial morphology, presence of apical microvilli, melanosomes, and tight junctions. iRPE monolayers also show electrophysiological responses that are typical of native RPE.<br /> <br /> Preliminary findings indicate L-ORD Patient i-RPE have impaired trans-epithelial resistance. This is consistent with the impaired adhesion of L-ORD RPE to Bruch’s membrane reported in the literature. Further investigation is currently underway to determine the disease mechanism.

Conclusions: The retinal degeneration in L-ORD patients resembles age-related macular degeneration (AMD), a multi-factorial late-onset disease. However, unlike AMD, L-ORD is a monogenic disease caused by a single missense (S163R) mutation in the CTRP5 protein, making it a powerful tool to study more genetically complex diseases such as AMD. Elucidating the mechanism by which mutated CTRP5 causes L-ORD will aid in understanding disease mechanisms in AMD and other retinal degenerations.