Abstract 3666: Major Role of Ly6chi/7/4hi/ccr2+ Monocytes in Post-ischemic Neovascularization
Circulating mouse monocytes consist of two main subsets, Ly6CHi7/ 4HiCCR2+CX3CR1loCCR5+ and Ly6Clo7/4loCCR2-CX3CR1HiCCR5+ monocytes.We sought to investigate their respective role in postischemic neovascularization and unravel the mechanisms involved in their recruitment to ischemic tissues. Postischemic neovascularization was impaired in MCP-1−/− or CCR2−/− but not in CCR5−/− or CX3CR1−/− mice with operatively-induced hindlimb ischemia (n=10 per groups). Overexpression of MCP-1 in the ischemic muscle by plasmid electrotransfer improved foot perfusion, angiographic score and capillary density by 1.4-, 1.8- and 1.4-fold, respectively (p<0 .01 versus control). In contrast, overexpression of Fractalkine or Rantes, the ligands of CX3CR1 and CCR5 respectively, had no significant effects (n=10 per groups) suggesting that recruitment of 7/4HiCCR2+ monocytes play a significant role in this setting. In this line, flow cytometry analysis revealed that monocytes infiltration in ischemic muscle peaked at day 3 and dropped to levels comparable to those in non ischemic muscle on day 7. 7/4Hi monocytes predominated from day 1 to day 3 (52% of infiltrated Cd11b+ cells) whereas 7/4lo prevailed from day 7 onward. MCP-1 or CCR2 deficiency fully abrogated infiltration of both monocytes subsets, while overexpression of MCP-1 in the ischemic muscle selectively increased 7/4Hi monocytes infiltration (287% of wild-type, p<0.05). In contrast, monocytes subsets infiltration was slightly reduced in CX3CR1−/− mice (p<0.05 versus MCP-1−/−) and unchanged in CCR5−/− mice. Interestingly, MCP-1 deficiency or upregulation also led to changes in monocyte subtypes levels in both bone marrow and blood. MCP-1/CCR2 signaling plays a major role in monocytes mobilization from the bone marrow and recruitment to the ischemic tissue. We also suggest that inflammatory 7/4Hi/CCR2 monocytes represent a major component of the inflammatory response in post-ischemic neovascularization.