Comparison of older and younger fibroblast in the homeostasis of 3D bioprinted oBRB model

Abstract

Purpose : Dry age-related macular degeneration (AMD) is a multifactorial disease associated with cellular senescence and chronic inflammation. Choroid fibroblasts have been thought to contribute to maintaining the integrity and homeostasis of the outer blood-retinal barrier (oBRB), although their role isn’t fully characterized. Here, we aim to investigate the role of choroidal fibroblasts in oBRB maturation and function using an in vitro 3D bioprinted oBRB model.

Methods : The 3D bioprinted vascularized choroid was generated using GFP expressing endothelial cells, choroid pericytes, and fibroblasts derived either from iPSC, younger, or older eye tissue. After one week of vasculature formation, retinal pigment epithelial (RPE) cells were seeded on the apical side of bioprinted choroid and matured for additional four weeks. Capillary density and maturation were quantified and RPE pigmentation, polygonal morphology, and apical process density were analyzed.

Results : Older fibroblasts in the model caused hypopigmentation, enlargement, elongation, and reduced apical process density in RPE cells, along with impaired vascularization within the 3D-choroid. On the other hand, inclusion of iPSC-derived fibroblasts or younger eye derived fibroblasts demonstrated greater TER measurements with improved capillary network and RPE morphology. Immunostained images after five weeks of tissue maturation disclosed that iPSC-derived fibroblasts self-assembled within the 3D choroid to form a basal boundary where little to no fibroblasts were detected in the capillaries or transwell membrane. In the case of the older fibroblast, there was an assembly of a confluent basal layer, however, a great number of fibroblasts were present in the capillaries and transwell membranes, which suggests that this cell type promotes vascular degeneration.

Conclusions : Our results indicate that numbers and age (or senescence) of fibroblasts in choroid play a role in maintaining capillary density and RPE maturity. Future work will focus on what gene expression changes in older versus younger fibroblasts are associated with their differential behavior in 3D oBRBs.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.