Metabolic reprogramming of endothelial precursor cells in subretinal fibrosis

Therapeutic agents that target the vascular endothelial growth factor (VEGF) have achieved remarkable success in patients with the neovascular form of age-related macular degeneration (nAMD). An emerging clinical problem, however, is that many of the nAMD patients develop subretinal fibrosis (SRF) after receiving anti-VEGF therapy. SRF can cause irreversible structural damage to the retina and is a major vision- threatening complication with no effective treatment. The disease mechanisms of SRF in nAMD are largely unknown. Transforming growth factor beta (TGF-beta) is a major driver of fibrosis. The source of TGF-beta in SRF, and its main effector cells, have not been well defined. SRF can be modeled in mice with spontaneous or experimentally-induced choroidal neovascularization (CNV). In our published and preliminary studies, we found that mice with targeted deletion in the very low-density lipoprotein receptor (Vldlr) gene developed SRF when their CNV lesions regressed. Using single cell RNA sequencing, we identified endothelial precursor cells (EPCs) as a major cluster of cells that displayed markers of fibrosis. Similar findings were observed in JR5558 mice and in laser-induced CNV. EPCs have stem cell-like properties, and they are recruited to the choroidal and retinal neovessels to facilitate the vascular repair. In the subretinal microenvironment, EPCs gradually lose their cellular structures and transdifferentiate into fibroblast-like cells. We hypothesize that TGF-beta-mediated metabolic reprograming of EPCs is a key signaling event that contributes to the formation and progression of SRF after CNV. For the project proposed in this application, we will determine the roles of EPCs in mouse models of CNV and in human donor eye tissues with wet AMD. We will also examine the metabolic reprogramming of EPCs in response to TGF-beta. Furthermore, we will explore whether Muller cell-derived IL- 33 promotes TGF-beta production from macrophages, and whether inhibiting the IL-33 signaling suppresses SRF. Results from these studies will reveal novel molecular and cellular mechanisms of SRF, and define new targets for potential therapeutic intervention.