Modeling the cell specific origins of MacTel using a large patient iPSC-RPE library

Abstract

Purpose : Macular Telangiectasia type 2 (MacTel) is a rare inherited macular degenerative disease with presumed polygenic inheritance and variable penetrance. Modeling this disease has proven challenging as there are no direct genetic or environmental links to the disease. Recent GWAS analyses and metabalomics screens have indicated a possible defect in serine metabolism in patients, and histological studies on MacTel patient retinas suggest a possible defect in the retinal pigmented epithelium (RPE). In absence of a genetic model to probe this complex disease we have generated a library of iPSC-derived RPE from 20 individuals including MacTel patients, unaffected family members, and unaffiliated controls. Patient derived RPE were assayed for expression profiles, metabolic function, and phagocytic capacity.

Methods : We generated and validated iPSCs from 20 MacTel patients, unaffected family members, and unaffected controls. 2-3 iPSC clones per patient were used to control for variability stemming from iPSC derivation. Clones were differentiated into RPE using activin and nicotinamide based differentiation protocols and RPE were postmitotically matured for up to 4 months. Maturity was determined by longitudinal studies in polar secretion of VEGF and PEDF for each line as well as complete functionality in phagocytosis assays.

Results : To supplement genetic studies on MacTel patients the iPSC-RPE patient library was screened using RNA-seq to detect difference in expression profiles linked to MacTel. We probe for defects in serine metabolism using stable isotope tracing of carbon 13 labeled glucose to quantify dysfunction in central carbon metabolism, which includes serine biosynthesis and subsequent serine metabolism. We further tested for metabolic dysfunction in bioenergetic assays using Seahorse to measure cellular respiration under various stressors. We also tested for physiological defects in phagocytosis by quantifying the phagocytosis rate of porcine outersegments.

Conclusions : To determine the possible contribution of defects in RPE to MacTel we provide a comprehensive analysis of gene expression, metabolism, energetics, and cell function and integrate them with WES analysis of patient genomes to determine a cell specific origin a complex inherited disease.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.