Deficiency of Endothelial Cxcr4 Reduces Reendothelialization and Enhances Neointimal Hyperplasia After Vascular Injury in Atherosclerosis-Prone Mice

Abstract

Objective— The Cxcl12/Cxcr4 chemokine ligand/receptor axis mediates the mobilization of smooth muscle cell progenitors, driving injury-induced neointimal hyperplasia. This study aimed to investigate the role of endothelial Cxcr4 in neointima formation. Approach and Results— β-Galactosidase staining using bone marrow x kinase ( Bmx ) -CreER T2 reporter mice and double immunofluorescence revealed an efficient and endothelial-specific deletion of Cxcr4 in Bmx-CreER T2+ compared with Bmx-CreER T2− Cxcr4-floxed apolipoprotein E–deficient ( Apoe −/− ) mice (referred to as Cxcr4 EC-KO ApoE −/− and Cxcr4 EC-WT ApoE −/− , respectively). Endothelial Cxcr4 deficiency significantly increased wire injury–induced neointima formation in carotid arteries from Cxcr4 EC-KO ApoE −/− mice. The lesions displayed a higher number of macrophages, whereas the smooth muscle cell and collagen content were reduced. This was associated with a significant reduction in reendothelialization and endothelial cell proliferation in injured Cxcr4 EC-KO ApoE −/− carotids compared with Cxcr4 EC-WT ApoE −/− controls. Furthermore, stimulation of human aortic endothelial cells with chemokine (C-X-C motif) ligand 12 (CXCL12) significantly enhanced their wound-healing capacity in an in vitro scratch assay, an effect that could be reversed with the CXCR4 antagonist AMD3100. Also, flow cytometric analysis showed a reduced mobilization of Sca1 + Flk1 + Cd31 + and of Lin − Sca1 + progenitors in Cxcr4 EC-KO ApoE −/− mice after vascular injury, although Cxcr4 surface expression was unaltered. No differences could be detected in plasma concentrations of Cxcl12, vascular endothelial growth factor, sphingosine 1-phosphate, or Flt3 (fms-related tyrosine kinase 3) ligand, all cytokines with an established role in progenitor cell mobilization. Nonetheless, double immunofluorescence revealed a significant reduction in local endothelial Cxcl12 staining in injured carotids from Cxcr4 EC-KO ApoE −/− mice. Conclusions— Endothelial Cxcr4 is crucial for efficient reendothelialization after vascular injury through endothelial wound healing and proliferation, and through the mobilization of Sca1 + Flk1 + Cd31 + cells, often referred to as circulating endothelial progenitor cells.