Heparin induces the mobilization of human multipotent circulating mesoangioblasts from the heart

Abstract

Rationale: We previously identified circulating mesoangioblasts (cMABs), a subset of mesenchymal stem cells that co-express KDR and Nkx2.5, from the patients undergoing cardiac surgery with cardiopulmonary bypass. cMABs are capable of differentiating into endothelial cells, smooth muscle cells and cardiomyocytes. More recently, we demonstrated that hepatocyte growth factor (HGF) induces cMAB mobilization in animal model. Objective: We therefore tested the hypothesis that heparin induces cMAB mobilization in humans by increasing the serum levels of HGF. We also explored the niche of cMABs. Methods and results: Patients undergoing cardiac catheterization were treated with an increasing dose of heparin at 100U/kg (n=7), 200 U/kg (n=11) and 300 U/kg (n=11). Serum HGF levels were determined by ELISA. Mononuclear cells (MNCs) were isolated from peripheral blood by Ficoll density gradient centrifugation and cultured on a fibronectin-coated dish, and the number of outgrowing colonies was counted. Consistent with previous studies, heparin dose-dependently increased serum HGF levels (28.1±3.0, 34.4±1.9, and 42.8±0.9 ng/ml, respectively, compared to 1.3±0.2 ng/ml in control subjects). Moreover, the number of cMAB colonies was also increased by heparin in a dose-dependent manner (0.27±0.26, 0.33±0.11, and 0.75±0.22 colonies/MNCs (x106), respectively). No cMAB colony outgrowth was observed in control subjects. Time course analysis of serum HGF levels and the extent of cMAB mobilization revealed that HGF concentration positively correlated with the number of cMAB colonies. The characteristics of cMABs mobilized by heparin were essentially the same with those of cMABs mobilized during cardiac surgery: they were negative for hematopoietic marker CD45 and positive for mesenchymal marker CD73, the HGF-receptor c-Met, endothelial marker KDR, cardiac mesoderm marker Nkx2.5, and pluripotency markers (Oct3/4, KLF4, and cMyc). To explore the niche of cMABs, we obtained blood from aortic sinus (Ao), right atrium (RA) and coronary sinus (CS), and the number of outgrowing cMAB colonies was evaluated. The highest number of outgrowing colonies was observed in CS-derived blood. Conclusions: Heparin induces the mobilization of heart-derived multipotent mesoangioblasts in human circulation presumably by increasing the serum levels of HGF.