Monocyte chemoattractant protein 1 and fractalkine play opposite roles in angiogenesis via recruitment of different macrophage subtypes

AIM： To explore the interaction between macrophages and chemokines [monocyte chemoattractant protein 1（MCP-1/CCL2） and fractalkine/CX3CL1] and the effects of their interaction on neovascularization.METHODS： Human peripheral blood mononuclear cells, donated by healthy volunteers, were separated and cultured in RPMI-1640 medium containing 10% fetal bovine serum, then induced into macrophages by stimulation with 30 μg/L granulocyte macrophage-colony stimulating factor（GM-CSF）. The expression of CCR2 and/or CX3CR1 in the macrophages was examined using flow cytometry. Macrophages were then stimulated with recombinant human CCL2（rh-CCL2） or recombinant human CX3CL1（rh-CX3CL1）. The expression of angiogenesis-related genes, including VEGF-A, THBS-1 and ADAMTS-1 were examined using real-time quantitative polymerase chain reaction（PCR）. Supernatants from stimulated macrophages were used in an assay of human retinal endothelial cell（HREC） proliferation. Finally, stimulated macrophages were cocultured with HREC in a migration assay.RESULTS： The expression rate of CCR2 in macrophages stimulated by GM-CSF was 42%±1.9%. The expression rate of CX3CR1 was 71%±3.3%. Compared with vehicle-treated groups, gene expression of VEGF-A in the macrophages was greater in 150 mg/L CCL2-treated groups（P〈0.05）, while expression of THBS-1 and ADAMTS-1 was significantly lower（P〈0.05）. By contrast, compared with vehicle-treated groups, expression of VEGF-A in 150 mg/L CX3CL1-treated groups was significantly lower（P〈0.05）, while expression of THBS-1 and ADAMTS-1 was greater（P〈0.05）. Supernatants from CCL2 treated macrophages promoted proliferation of HREC（P〈0.05）, while supernatants from CX3CL1-treated macrophages inhibited the proliferation of HREC（P〈0.05）. HREC migration increased when co-cultured with CCL2-treated macrophages, but decreased with CX3CL1-treated macrophages（P〈0.05）.CONCLUSION： CCL2 and CX3CL1 exert different effects in regulation of macrophage in expression of angiogenesisrelated factors, including VEGF-A, THBS-1 and ADAMTS-1. Our findings suggest that CCL2 and CX3CL1 may be candidate proteins for further exploration of novel targets for treatment of ocular neovascularization.