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Christopher Hampton, Min Jae Song, Roba Dejene, Devika Bose, Russ Quinn, Kapil Bharti; Hypoxia of Retina Pigment Epithelium Induces Type 1 CNV-like Morphology within 3D Engineered iPSC-RPE/“Choroid” Tissues. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3272.
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© ARVO (1962-2015); The Authors (2016-present)
Hypoxia has been reported as a critical driving factor of choroidal neovascularization (CNV) in the wet form of age related macular degeneration (wet AMD). We induce hypoxia inducible factor 1 alpha (HIF-1α) in iPSC derived RPE to study underlying mechanisms of CNV initiation/progression using engineered 3D tissues containing RPE and “choroid”. This model is being used to understand pathogenesis of wet AMD and for drug testing.
We developed choroid-like 3D vascularized tissue structure using bioprinting technology. Optimal “Bioink”, which contains endothelial cells (ECs), pericytes, and fibroblasts embedded in hydrogel, was printed on one side of a biodegradable polymer scaffold. iPSC derived RPE monolayer was formed on the other side of the scaffold. The scaffold served as a temporary Bruch’s membrane until RPE and endothelial cells produced extracellular matrix (ECM) proteins. DMOG (100µM) was added to only the apical side of the RPE to mimic local hypoxia surrounding RPE. After two weeks of DMOG treatment, we measured cytokine secretion from RPE, transepithelial/tissue resistance (TER), and angiogenesis of “choroid”.
RPE polarity was confirmed by the presence of apical processes and apical EZRIN expression. Fenestration of “choriocapillaris” was confirmed with EM and expression of a fenestration marker, FELS. DMOG treatment significantly induced secretion of proangiogenic factors, such as vascular endothelial growth factor (VEGF), in the basal region of RPE. The stressed RPE increased angiogenesis of “choriocapillaris” and significantly increased vessel growth in the subRPE region, similar to “wet” AMD.
We successfully demonstrated that DMOG induced HIF-1α in RPE causes a type I CNV like vessel growth pattern within RPE/“choroid” tissues. This model shows that increased secretion of VEGF and likely other proangiogenic factors by RPE are critical in directing CNV toward RPE. HIF-1α acts upstream of VEGF expression within RPE. Interestingly, RPE electrical resistance barrier function is elevated with DMOG treatment, which is presumably a part of the RPE’s protective mechanism. We haven’t yet seen advanced CNV (type 2 or 3) in this model. Other types of stressors and gene modification of iPSC-RPE will be tested to understand disease pathways involved in wet AMD.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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