RPE autonomous molecular events are sufficient for drusen biogenesis and extracellular matrix alterations in hiPSC-derived models of macular degeneration.

Abstract

Purpose : Patient-derived human induced pluripotent stem cells (hiPSCs) allow the investigation of a singular cell type in the disease pathology. The purpose of this study was to determine the specific role of retinal pigment epithelium (RPE) cell in macular degeneration pathogenesis. We accomplished this by evaluating the phenotypic and molecular alterations in patient-derived hiPSC-RPE model of multiple maculopathies affecting the RPE-extracellular matrix (ECM) complex.

Methods : hiPSC-RPE were derived from patients with inherited maculopathies, Sorsby’s fundus dystrophy (SFD) and Doyne honeycomb retinal dystrophy (DHRD), and adult onset macular degeneration (MD, no known genetic defect) as well as unaffected controls (family member, gene corrected line, random population). Ultrastructural, immunocytochemical and gene/protein expression analyses were performed to determine the 1) development and composition of sub-RPE deposits (drusen) 2) ECM alterations including altered matrix metalloproteinases (MMP) activity/expression and ECM protein accumulation and 3) complement activation in hiPSC-RPE monoculture model of SFD, DHRD and MD.

Results : Monocultures of patient hiPSC-RPE (SFD/DHRD/MD) aged for > 90 days in culture developed sub-RPE deposits that consisted of lipids (Nile red) and RPE synthesized proteins (ApoE, TIMP3). Importantly, no drusen deposits were seen in relatively young (~day 30) patient-hiPSC-RPE cultures. Serum exposure affected the composition of drusen in patient hiPSC-RPE cultures, with the inclusion of complement proteins (C3, membrane attack complex (MAC) or C5b-9) and vitronectin (VIT). Patient hiPSC-RPE models of macular degeneration (in the absence of exogenous stressors) also demonstrated accumulation of ECM proteins TIMP3, Col IV and laminin and altered expression/activity of several MMP and complement genes.

Conclusions : Using hiPSC-RPE from multiple patients with distinct maculopathies but similar pathological phenotype, we have established that RPE cells alone are sufficient to promote drusen deposition in a cell autonomous manner in maculopathies affecting RPE-ECM complex. Furthermore, we have for the first time illustrated in a human model system that similar molecular alterations (impaired ECM turnover, complement response) underlie distinct maculopathies (SFD, DHRD, MD).

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.