Relevance of Monocytic Features for Neovascularization Capacity of Circulating Endothelial Progenitor Cells

TLDR

Transplantation of ex vivo expanded circulating endothelial progenitor cells (EPCs) from peripheral blood mononuclear cells improves neovascularization after critical ischemia, yet the lineage origin and defining characteristics of EPCs remain unclear. This study examined whether EPC phenotype and neovascularization capacity depend on monocytic origin. Monocytic CD14+ cells were isolated, cultured on fibronectin in endothelial medium with VEGF, and expanded ex vivo to generate EPCs. EPCs derived from both CD14+ and CD14− mononuclear cells displayed equivalent endothelial marker expression and clonal expansion, and when transplanted into hind‑limb ischemic mice, both populations increased relative blood flow from 0.27 to ~0.65–0.66; in contrast, freshly isolated CD14+ cells or their macrophage/dendritic derivatives did not improve perfusion, showing that nonmonocytic CD14− cells can generate functional EPCs that uniquely enhance vascular repair.

Abstract

Background— Transplantation of ex vivo expanded circulating endothelial progenitor cells (EPCs) from peripheral blood mononuclear cells improves the neovascularization after critical ischemia. However, the origin of the endothelial progenitor lineage and its characteristics have not yet been clearly defined. Therefore, we investigated whether the phenotype and functional capacity of EPCs to improve neovascularization depend on their monocytic origin. Methods and Results— Monocytic CD14 + cells were isolated from mononuclear cells and incubated on fibronectin-coated dishes in endothelial medium in the presence of vascular endothelial growth factor. After 4 days of cultivation, adherent cells deriving from CD14 + or CD14 − mononuclear cells showed equal expression of endothelial marker proteins and capacity for clonal expansion as determined by measuring endothelial colony-forming units. In addition, transplanted EPCs (5×10 5 cells) deriving from CD14 + or CD14 − cells were incorporated into vascular structures of nude mice after hind-limb ischemia and significantly improved neovascularization from 0.27±0.12 (no cells) to 0.66±0.12 and 0.65±0.17, respectively ( P <0.001; laser Doppler-derived relative blood flow). In contrast, no functional improvement of neovascularization was detected when freshly isolated CD14 + mononuclear cells without ex vivo expansion were used (0.33±0.17). Moreover, macrophages or dendritic cells differentiated from isolated CD14 + cells were significantly less effective in improving neovascularization than EPCs cultivated from the same starting population ( P <0.01). Conclusions— These data demonstrate that EPCs can be generated from nonmonocytic CD14 − peripheral blood mononuclear cells and exhibit a unique functional activity to improve neovascularization after hind-limb ischemia.