A novel bone marrow frozen section assay for studying hematopoietic interactions in situ: the role of stromal bone marrow macrophages in erythroblast binding

Bone marrow macrophages are found in intimate contact with erythroblasts. Exact mechanisms and functions of this interaction are unclear. New insights into erythroblast binding were obtained using a newly developed bone marrow frozen section assay. This modified Woodruff and Stamper assay has some important advantages compared to other adhesion assays. Erythroblasts specifically adhered to bone marrow macrophages forming clusters, as appear in vivo. Selective depletion of bone marrow macrophages by intravenous injection of dichloromethyl-enediphosphonate resulted in a release of immature erythroid cells to the peripheral blood. Furthermore no erythroblasts adhered to bone marrow sections without macrophages. Evaluating the binding of erythroblasts to bone marrow macrophages we found that this binding is temperature and cation dependent. The receptor for erythroblasts present on macrophages recognizes a sialyated protein as ligand on erythroblasts, since neuraminidase treatment of erythroblasts resulted in a decrease in binding. A possible candidate for the erythroblast receptor on macrophages is the ED2 antigen. ED2 is a differentiation antigen present on resident macrophages that has some biochemical features characteristic of an adhesion molecule. Erythroblast binding to bone marrow was inhibited using a monoclonal antibody directed against ED2.