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Updated: Aug 9, 2025
Digestion of Whole Mouse Eyes for Multi-Parameter Flow Cytometric Analysis of Mononuclear Phagocytes
Published on: June 17, 2020
CD11c+ macrophages are proangiogenic and necessary for experimental choroidal neovascularization
Steven Droho1, Amrita Rajesh1, Carla M Cuda2
1Department of Ophthalmology and.
Abstract:
Patients with neovascular AMD (nAMD) suffer vision loss from destructive angiogenesis, termed choroidal neovascularization (CNV). Macrophages are found in CNV lesions from patients with nAMD. Additionally, Ccr2-/- mice, which lack classical monocyte-derived macrophages, show reduced CNV size. However, macrophages are highly diverse cells that can perform multiple functions. We performed single-cell RNA-Seq on immune cells from WT and Ccr2-/- eyes to uncover macrophage heterogeneity during the laser-induced CNV mouse model of nAMD. We identified 12 macrophage clusters, including Spp1+ macrophages. Spp1+ macrophages were enriched from WT lasered eyes and expressed a proangiogenic transcriptome via multiple pathways, including vascular endothelial growth factor signaling, endothelial cell sprouting, cytokine signaling, and fibrosis. Additionally, Spp1+ macrophages expressed the marker CD11c, and CD11c+ macrophages were increased by laser and present in CNV lesions. Finally, CD11c+ macrophage depletion reduced CNV size by 40%. These findings broaden our understanding of ocular macrophage heterogeneity and implicate CD11c+ macrophages as potential therapeutic targets for treatment-resistant patients with nAMD.
Insights
Researchers uncovered specific macrophage subtypes driving wet age-related macular degeneration (AMD). Depleting CD11c+ macrophages significantly reduced choroidal neovascularization (CNV), suggesting a new therapeutic target for vision loss in AMD patients.
Area of Science:
- Ophthalmology
- Immunology
- Cell Biology
Background:
- Neovascular age-related macular degeneration (nAMD) causes vision loss through choroidal neovascularization (CNV).
- Macrophages infiltrate CNV lesions, and their absence in Ccr2-/- mice reduces CNV size, suggesting a role in nAMD pathogenesis.
- Macrophage populations are diverse and functionally heterogeneous, necessitating deeper investigation into their specific roles in ocular disease.
Purpose of the Study:
- To investigate macrophage heterogeneity in a laser-induced mouse model of nAMD using single-cell RNA-Seq.
- To identify specific macrophage subsets involved in the development of choroidal neovascularization.
- To evaluate the therapeutic potential of targeting identified macrophage populations in nAMD.
Main Methods:
- Single-cell RNA-sequencing of immune cells from wild-type (WT) and Ccr2-/- laser-injured eyes.
- Bioinformatic analysis to identify and characterize distinct macrophage clusters.
- Immunohistochemistry to detect CD11c+ macrophages in CNV lesions.
- Experimental depletion of CD11c+ macrophages to assess their impact on CNV size.
Main Results:
- Twelve distinct macrophage clusters were identified, including a proangiogenic Spp1+ macrophage subset enriched in WT lasered eyes.
- Spp1+ macrophages exhibited transcriptomes associated with vascular endothelial growth factor signaling, endothelial cell sprouting, cytokine signaling, and fibrosis.
- CD11c+ macrophages, a subset of Spp1+ macrophages, were increased by laser injury and found within CNV lesions.
- Depletion of CD11c+ macrophages resulted in a significant 40% reduction in CNV size.
Conclusions:
- Ocular macrophage populations exhibit significant heterogeneity relevant to nAMD pathogenesis.
- CD11c+ macrophages play a critical proangiogenic role in choroidal neovascularization.
- Targeting CD11c+ macrophages presents a promising therapeutic strategy for treatment-resistant nAMD.
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