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Updated: May 8, 2026
A Mouse Model for Laser-induced Choroidal Neovascularization
Published on: December 27, 2015
VEGF receptor blockade markedly reduces retinal microglia/macrophage infiltration into laser-induced CNV
Hu Huang1, Rachel Parlier, Ji-Kui Shen
1Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America.
Abstract:
Although blocking VEGF has a positive effect in wet age-related macular degeneration (AMD), the effect of blocking its receptors remains unclear. This was an investigation of the effect of VEGF receptor (VEGFR) 1 and/or 2 blockade on retinal microglia/macrophage infiltration in laser-induced choroidal neovascularization (CNV), a model of wet AMD. CNV lesions were isolated by laser capture microdissection at 3, 7, and 14 days after laser and analyzed by RT-PCR and immunofluorescence staining for mRNA and protein expression, respectively. Neutralizing antibodies for VEGFR1 or R2 and the microglia inhibitor minocycline were injected intraperitoneally (IP). Anti-CD11b, CD45 and Iba1 antibodies were used to confirm the cell identity of retinal microglia/macrophage, in the RPE/choroidal flat mounts or retinal cross sections. CD11b(+), CD45(+) or Iba1(+) cells were counted. mRNA of VEGFR1 and its three ligands, PlGF, VEGF-A (VEGF) and VEGF-B, were expressed at all stages, but VEGFR2 were detected only in the late stage. PlGF and VEGF proteins were expressed at 3 and 7 days after laser. Anti-VEGFR1 (MF1) delivered IP 3 days after laser inhibited infiltration of leukocyte populations, largely retinal microglia/macrophage to CNV, while anti-VEGFR2 (DC101) had no effect. At 14 days after laser, both MF1 and DC101 antibodies markedly inhibited retinal microglia/macrophage infiltration into CNV. Therefore, VEGFR1 and R2 play differential roles in the pathogenesis of CNV: VEGFR1 plays a dominant role at 3 days after laser; but both receptors play pivotal roles at 14 days after laser. In vivo imaging demonstrated accumulation of GFP-expressing microglia into CNV in both CX3CR1(gfp/gfp) and CX3CR1(gfp/+) mice. Minocycline treatment caused a significant increase in lectin(+) cells in the sub-retinal space anterior to CNV and a decrease in dextran-perfused neovessels compared to controls. Targeting the chemoattractant molecules that regulate trafficking of retinal microglia/macrophage appears to be a compelling therapeutic strategy to control CNV and treat wet AMD.
Insights
Blocking VEGF receptor 1 (VEGFR1) significantly reduced microglia/macrophage infiltration in wet age-related macular degeneration (AMD) models. Both VEGFR1 and VEGFR2 inhibition were crucial at later stages for controlling choroidal neovascularization (CNV).
Area of Science:
- Ophthalmology
- Immunology
- Molecular Biology
Background:
- Vascular Endothelial Growth Factor (VEGF) pathway blockade is effective for wet age-related macular degeneration (AMD).
- The specific roles of VEGF receptors (VEGFR1 and VEGFR2) in microglia/macrophage infiltration in choroidal neovascularization (CNV), a model for wet AMD, are not fully understood.
Purpose of the Study:
- To investigate the impact of VEGFR1 and/or VEGFR2 blockade on retinal microglia/macrophage infiltration in a laser-induced CNV model.
- To elucidate the differential roles of VEGFR1 and VEGFR2 in the pathogenesis of CNV.
Main Methods:
- CNV lesions were analyzed using laser capture microdissection, RT-PCR, and immunofluorescence staining at 3, 7, and 14 days post-laser.
- Neutralizing antibodies against VEGFR1 (MF1) or VEGFR2 (DC101) were administered intraperitoneally.
- Microglia/macrophage infiltration was assessed using antibodies against CD11b, CD45, and Iba1, and quantified via cell counting.
Main Results:
- VEGFR1 mRNA and protein were expressed at all stages, while VEGFR2 was detected only at later stages.
- Anti-VEGFR1 (MF1) inhibited leukocyte infiltration at 3 days post-laser, whereas anti-VEGFR2 (DC101) had no effect at this time point.
- By 14 days post-laser, both MF1 and DC101 markedly inhibited microglia/macrophage infiltration into CNV.
Conclusions:
- VEGFR1 plays a dominant role in early CNV pathogenesis (3 days), while both VEGFR1 and VEGFR2 are pivotal in later stages (14 days).
- Targeting chemoattractant molecules regulating retinal microglia/macrophage trafficking presents a promising therapeutic strategy for CNV and wet AMD.
