Abstract
Purpose :
Choroidal neovascularization (CNV) is a hallmark of neovascular age-related macular degeneration (nAMD). In the aging population, aberrant growth of choroidal vessels through the Bruch’s membrane, termed CNV, results in loss of central vision. During active neovascularization, CNV is associated with inappropriate levels of apoptosis in multiple cell types including choroidal endothelial cells (ChEC). Bim is a pro-apoptotic Bcl-2 family member and is essential for apoptosis of cells exposed to dexamethasone and many other drugs or decreased pro-survival factors such as VEGF. Thus, gaining a comprehensive understanding of the role varied Bim expression levels play in modulation of ChEC phenotype and their response to various therapeutics will allow more tailored treatment modalities.
Methods :
To determine the impact of Bim expression on ChEC phenotype, ChEC were isolated from Bim +/+ and Bim -/- mice. Flow cytometry was utilized to ensure expression of appropriate EC markers in these cells. The impact of VEGF signaling cell survival, proliferation, migration, and inflammatory cytokine expression was assessed. Cell migration was assessed using a scratch wound assay. VEGF and inflammatory cytokine expression was determined by qPCR. Cell survival was examined using a Caspase-Glo 3/7-assay. Cell proliferation was assessed using a Click-iT-EdU. Capillary morphogenesis was evaluated by plating cells in Matrigel. The mouse model of laser induced CNV was used to assess the impact of anti-VEGF treatment in 3-month-old Bim+/+ and Bim-/- mice.
Results :
ChEC lacking Bim expression demonstrated increased VEGF, osteopontin and inflammatory cytokines Rantes/Ccl5, Mcp1/Ccl2 and IL6 expression. Bim -/- ChEC were more proliferative and demonstrated an increased capacity to undergo capillary morphogenesis. Anti-VEGF disrupted capillary morphogenesis effectively in Bim +/+ ChEC but its effectiveness in Bim -/- cells was significantly reduced. Although anti-VEGF treatment mitigated CNV in Bim+/+ mice it failed to mitigate CNV in Bim-/- mice.
Conclusions :
Reduced levels of Bim expression impacts the effectiveness of anti-VEGF to disrupt ChEC capillary morphogenesis. This is likely due to increased proliferation, levels of VEGF, extracellular matrix proteins and inflammatory cytokines expression, and could also explain the lack of anti-VEGF response in a significant portion of nAMD patients with varied Bim expression.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.