Multiomics data reveal microglia's promotion for choroidal neovascularization in endothelial cells

Choroidal neovascularization (CNV) stands as one of the leading causes of blindness worldwide, driven by the dysregulation of key signaling pathways, including vascular endothelial growth factor (VEGF) and transforming growth factor-β (TGF-β). This study aimed to elucidate the specific cell types within the retina and retinal pigment epithelium (RPE)/choroid complex that contribute to CNV progression, as well as to explore how the expression levels of cysteine-rich protein 2 (CSRP2), a downstream effector, are altered in the pathological mechanisms underlying CNV. By investigating these molecular and cellular dynamics, we seek to provide deeper insights into the disease's progression and identify potential therapeutic targets. Retinal tissues, RPE/choroid complexes, and blood samples from both age-related macular degeneration (AMD) caused CNV patients and healthy controls were obtained from the GEO database for differential gene expression analysis. Integrated analysis of tissue and blood samples from wet AMD patients and healthy controls identified CSRP2 as a critical biomarker gene associated with pathogenesis. To uncover potential underlying mechanisms, we conducted immune infiltration analysis and further validated our findings using single-cell RNA sequencing (scRNA-seq) data from the GEO database. Additionally, scRNA-seq data were utilized to investigate cell-cell communication networks and perform Gene Set Enrichment Analysis (GSEA). scRNA-seq analysis demonstrated that CSRP2 was significantly upregulated in microglia and endothelial cells, with concurrent activation of the VEGF and TGF-β signaling pathways. Microglia emerged as a central hub for outgoing interactions, while endothelial cells were identified as the primary target of incoming signals within these pathways. GSEA further implicated CSRP2 in CNV progression, highlighting its role in angioimmunoblastic regulated by VEGF and TGF-β signaling pathways. In the in-vitro experiments, we found that activated microglia stimulated VEGFA, TGF-β and CSRP2, which enhanced angiogenesis, migration, proliferation, permeability, and altered the phenotype of co-cultured choroidal endothelial cells. These findings underscore the pivotal involvement of CSRP2 in mediating cellular crosstalk and signaling dynamics critical to CNV development. Microglia and endothelial cells emerged as the primary cell clusters interacting under this signaling regulation, driving angiogenesis and contributing to the pathological progression of CNV. The findings provide promise alternative therapy for CNV patients casued by AMD.