A Novel Paradigm In Stem Cell Trafficking: The Ratio of Peripheral Blood Sphingosine-1 Phosphate (S1P) to Bone Marrow Ceramide-1 Phosphate (C1P) Regulates Mobilization and Homing of Hematopoietic Stem Cells

Abstract 554

The stromal derived factor-1 (SDF-1)–CXCR4 axis plays an unquestioned role in developmental migration of hematopoietic stem cells (HSPCs) and their retention in the bone marrow (BM). However, changes in the SDF-1 gradient between BM and peripheral blood (PB) do not always support its having a crucial role as chemoattractant for mobilization or homing of HSPCs. As demonstrated by others (e.g., Bone Marrow Transplantation 2003; 31:651–654, and Transfus Apher Sci 2009;40:159) and us (Leukemia 2010;24:976–985) the plasma SDF-1 level does not correlate with mobilization of HSPCs. On the other hand, there is increasing doubt about an exclusive role for SDF-1 in homing of HSPCs in BM. This is based on evidence that i) CXCR4^−/− fetal liver HSPCs may home to BM in an SDF-1–independent manner (Immunity 1999;10:463-471), ii) homing of murine HSPCs made refractory to SDF-1 by incubation and co-injection with a CXCR4 receptor antagonist is normal or only mildly reduced (Science 2004;305:1000), and finally iii) HSPCs in which CXCR4 has been knocked down by means of an SDF-1 intrakine strategy also engraft in lethally irradiated recipients (Blood 2000;96:2074–,2080). All this strongly suggests the existence of other factors involved in the mobilization and homing of HSPCs. Moreover, while SDF-1 is a potent chemoattractant for HSPCs when employed at supraphysiological concentrations in vitro, as a peptide it is highly susceptible to degradation by proteases that are elevated, for example, in PB during stem cell mobilization or in the BM microenvironment after myeloablative conditioning for transplantation. Employing ELISA for detection in the present study, we observed insignificant changes in SDF-1 level both in PB during mobilization and in BM after myeloablative conditioning. We also found that mobilized PB (mPB) plasma as well as conditioned media (CM) from lethally irradiated mice chemoattract HSPCs in an SDF-1–independent manner as demonstrated by i) normal chemotaxis of AMD3100 pre-treated cells and ii) preservation of chemotactic activity of plasma and BM-derived CM following heat inactivation. However, the chemotactic activity of mPB plasma and BM CM was inhibited after stripping by activated charcoal. This suggested the involvement of small molecule bioactive lipids. It is known that sphingolipids, which are important components of cell membranes, give rise to two bioactive derivatives, sphingosine-1 phosphate (S1P) and ceramide-1 phosphate (C1P), with S1P already identified as a chemoattractant for HSPCs (Ann N Y Acad Sci. 200;1044:84–89). To our surprise, we found that C1P is also a strong chemoattractant for human and murine HSPCs. In addition, we observed that at physiological concentrations both these bioactive lipids i) activate phosphorylation of MAPKp42/44 and AKT in HSPCs, ii) induce expression of matrix metalloproteinases (MMPs), and iii) modulate adhesion to stroma and endothelium. Interestingly, by employing ELISA and/or mass spectophotometry we found that, while the S1P level increases in PB during mobilization, the C1P level increases in BM after myeloablative conditioning for transplantation. Based on these findings, we propose a new paradigm in which the S1P:C1P ratio plays a role in mobilization and homing of HSPCs. While S1P is a major chemoattractant that directs egress of HSPCs from BM into PB, C1P released from damaged cells in BM after myeloablative conditioning creates a homing gradient for circulating HSPCs. We also postulate that the S1P:C1P ratio plays a more universal role and is involved in regulating migration of other types of stem cells, such as circulating mesenchymal stem cells (MSCs), endothelial progenitor cells (EPCs), and very small embryonic-like (VSEL) stem cells. Accordingly, while S1P plays a role in egress of stem cells into PB, C1P released from damaged cells (e.g., in infarcted myocardium or brain tissue after stroke) chemoattracts circulating stem cells for potential repair.