Activation of ephrin A proteins influences hematopoietic stem cell adhesion and trafficking patterns

Abstract

To determine if Eph receptors and ephrins can modulate the homing of hematopoietic cells in a murine bone marrow transplantation model. EphA and ephrin A gene expression by mouse hematopoietic stem cells and the progenitor cell line FDCP-1 was determined by real-time reverse transcription polymerase chain reaction and flow cytometry. The effect of ephrin A activation on adhesion of hematopoietic progenitors was determined by in vitro adhesion assays in which cells were exposed to fibronectin or vascular cell adhesion molecule-1 (VCAM-1) and an increasing gradient of immobilized EphA3-Fc. Adhesion to fibronectin and VCAM-1 was further investigated using soluble preclustered EphA3-Fc. We used soluble unclustered EphA3-Fc as an antagonist to block endogenous EphA-ephrin A interactions in vivo. The effect of injecting soluble EphA3-Fc on the mobilization of hematopoietic progenitor cells was examined. We determined the effect on short-term homing by pretreating bone marrow cells with EphA3-Fc or the control IgG before infusion into lethally irradiated mice. Preclustered and immobilized EphA3-Fc increased adhesion of progenitor cells and FDCP-1 to fibronectin and VCAM-1 (1.6- to 2-fold higher adhesion; p < 0.05) relative to control (0 μ/cm(2) EphA3-Fc extracellular molecule alone). Injection of the antagonist soluble EphA3-Fc increased progenitor cell and colony-forming unit-spleen cells in the peripheral blood (42% greater colony-forming unit in culture; p < 0.05, 3.8-fold higher colony-forming unit-spleen) relative to control. Treating bone marrow cells with EphA3-Fc resulted in a reduction by 31% in donor stem cells homing to the bone marrow and accumulation of donor cells in recipient spleens (50% greater than control) and greater recovery of donor stem cells from the peripheral blood.