Regulation Of Hematopoietic Cell Adhesion And Migration By Cytokine Signals Cooperated With Sphingosine Kinase

The homing of hematopoietic stem cells (HSC) is referred as a process that circulating hematopoietic stem cells lodge in the bone marrow microenvironment where self-renewal, proliferation and differentiation will occur. It is hypothesized that homing is a multistep process, consisting of adhesion of the HSCs to marrow endothelial cells, followed by transendothelial migration directed by chemoattractants, and finally anchoring within the extravascular bone marrow spaces. However, the molecular mechanisms that govern this process are poorly understood. Clarifying of homing mechanisms will be very helpful for us to understand the hematopoietic development, hematopoiesis regulation and for the application of hematopoietic stem cells.The adhesion and migration of hematopoietic stem cells, which are regulated by various cytokines derived from bone marrow microenvironments, are critical homing steps. SCF and SDF-1 a are the major cytokines secreted by bone marrow stromal cells and play important roles in the regulation of hematopoietic stem cell homing. In this study, we evaluated the stimulating effect of SCF on the integrin-mediated adhesion of hematopoietic cells to Fn and chemoattract effects of SDF-1 a on the cord blood-derived CD34+ cells. And we also investigated the cooperation between the cytokine signals and sphingosine kinase in the regulation of hematopoietic cell adhesion and migration.The adhesive ability of hematopoietic cells depends on the density and activation of cell surface adhesion molecules. Interaction of VLA-4, VLA-5 integrin receptors with Fn of cell matrix plays important roles in selective homing of hematopoietic stem cells into bone marrow. We analyzed the expression of VLA-4, VLA-5 integrins and the SCF receptor (c-kit) by cord blood-derived CD34+ cells and Mo7e cells. Both these cells express VLA-4, VLA-5 integrins and the SCF receptor (c-kit). So these cells were chosen as a model system to study the functions of SCF on adhesion of hematopoietic cells. We also established a cell adhesion assay in which the adherentcells were determined by quantifying the cellular LDH. SCF enhances the adhesion of Mo7e cells and cord blood -derived CD34+ cells to Fn in a concentration-dependent manner. Anti-pi, a4, a5 (subunits of VLA-4, VLA-5) antibodies block the adhesion of Mo7e cells to Fn induced by SCF. These results indicate that SCF enhances the adhesion of Mo7e cells to Fn via VLA-4 and VLA-5 integrins. The functions of SCF are mediated through the activation of its high affinity tyrosine kinase receptor, c-kit proto-oncogene. The activated c-kit receptor recruits a number of signaling proteins that led to activation of PI3K and ERK/MAPK. Treatment of Mo7e cells with Ly294002, wortmannin (WT), the specific inhibitors of PI3K, result in blocking the SCF-induced adhesion to Fn. It is indicated that PI-3 kinase pathway is involved in the SCF-induced homing of hematopoietic stem cells.The chemoattract migration has been recognized as a key event in the homing of hematopoietic stem and progenitor cells. Signals through the chemokine stromal derived factor -1 a (SDF-1 a ) and its receptor CXCR4 regulates the migration of several types of cells including hematopoietic stem cells and lymphocytes. Cord blood-derived CD34+cells express CXCR4, the receptor for SDF-1 a . Recombinant human SDF-1 a promotes the migration of cord blood-derived CD34+ cells in a concentration dependent manner. CD34+ cells are a heterogenous group of hematopoietic stem/progenitor cells including CFU-mix, CFU-GM,BFU-E. SDF-1 a induces the migration of these colony forming cells. The SDF-1 a -induced migration of cord blood-derived CD34+ cells was blocked by Ly294002 and WT, specific inhibitors of the PI3K. It is suggested that PI3K might be involved in SDF-1 a -induced migration of hematopoietic cells.Sphingosine kinase (SPK) is a key enzyme catalyzing the formation of sphingosine 1 phosphate (SIP), a lipid messenger that is implicated in the regulation of a wide variety of important cellular events through intracellular as we...