Neutrophils promote laser-induced choroidal neovascularization by increasing pro-inflammatory cytokines secretion and cell cycle arrest in retinal pigment epithelium.

Abstract

Inflammation is hypothesized to have essential functions in the development of wet age-related macular degeneration (AMD). Polymorphonuclear neutrophils (PMNs), recognized as major players in inflammation, are typically the first leukocytes to be recruited to an inflammatory site. Previous studies have identified neutrophil aggregates in the lesion site of the choroidal neovascularization model, and systemic depletion of neutrophils in adult mice is associated with reduced choroidal neovascularization (CNV) area, suggesting a pivotal role of PMNs in CNV pathogenesis. Here, we investigate the role of neutrophils in promoting CNV, a key feature of wet AMD. The malfunction and demise of retinal pigment epithelium cells are essential elements in CNV pathogenesis. Our hypothesis posits that neutrophils exacerbate CNV by influencing pro-inflammatory cytokines secreted by retinal pigment epithelium (RPE) cells. Using in vivo laser-induced CNV models with mice and in vitro experiments with the human ARPE-19 cell line, we demonstrated that co-culturing neutrophils with ARPE-19 cells induces an increase in pro-inflammatory cytokines and leads to S-phase cell cycle arrest, potentially through the induction of double-strand breaks (DSBs). Further exploration of this interaction revealed a potential pathway involving reactive oxygen species (ROS) and microRNA-23a, wherein PMNs induce DSBs by initiating the downregulation of LB1 via microRNA-23a. Additionally, we found that dHL-60 cell line could serve as a substitute for primary PMNs, highlighting its potential as a valuable tool in experimental models involving interactions with retinal cells. Our findings underscore the significant role of neutrophils in CNV pathogenesis, providing insights into potential therapeutic targets for wet AMD.