Metabolism of AMD iPSC-derived RPE

Project Summary/Abstract
Age-related macular degeneration (AMD) is a leading cause of visual impairment and blindness in adults over
the age of 65 and is expected to affect ~288 million people worldwide by the year 2040. Recently, induced
pluripotent stem cells (iPSC)-derived RPE generated from AMD patients and those with phenotypically similar
monogenic diseases have been shown to approximate elements of AMD disease phenotype in culture,
including the formation of sub-RPE deposits resembling drusen, dysregulated complement, and mitochondrial
dysfunction. Our groups and others have measured metabolite usage, glycolysis, mitochondrial function, and
lipid metabolism in a variety of iPSC RPE model systems. While in vitro RPE models show significant promise
in the discovery of disease mechanisms and therapeutic targets, there is also increasing awareness of
potential limitations, including reproducibility across model systems and fidelity to native conditions. A
comprehensive review of recent iPSC RPE studies shows that the most used traditional culture media are
highly diverse in nutrient and metabolite content which may significantly alter RPE metabolism. Moreover,
multiple types of plating substrates used could contribute to the variability in nutrient environments. A lack of
consensus on baseline nutrient environments and knowledge of their impact on RPE metabolism makes
comparisons between findings challenging. The goal of this proposal is to characterize the metabolic and
disease-relevant phenotypic profiles of AMD iPSC RPE cells in three distinct and commonly used traditional
media and physiological medium closely approximating the composition of human blood. Two AMD iPSC RPE
lines and their CRISPR-corrected isogenic controls will be used in this study. RPE will be differentiated from
one NIH/NYSCF AREDS2 subject iPSC line with multiple known high-risk alleles, selected to gender and
complotype-match RPE lines generated from an individual with early onset macular drusen (EOMD). A splicing
mutation in the CFH gene results in this severe subtype of AMD, and our preliminary data show that EOMD
iPSC RPE display AMD disease-relevant features, including complement dysregulation, sub-RPE deposit
formation, and altered metabolism. iPSC RPE will be cultured on twp substrates (Matrigel®, vitronectin), and
maintained in four media preparations (MEM-α based, DMEM/F-12 based, X-VIVO 10TM and PlasmaxTM). This
project aims to determine the impact of culture microenvironment on AMD and EOMD iPSC RPE metabolism
and disease phenotype. The outcome of this project will be a new and more comprehensive understanding of
how traditional and physiologic media influence the metabolic profile and phenotypic characteristics of normal
and diseased RPE cells. This new understanding will aid in the interpretation of metabolite studies across
model systems and help to inform the design of more physiologic cell culture media for future studies.

Grant Number: 5U01EY034591-03
NIH Institute/Center: NIH
Principal Investigator: Jennifer Chao