MOUSE BONE-MARROW STROMAL CELL-LINE MC3T3-G2 PA6 WITH HEMATOPOIETIC-SUPPORTING ACTIVITY EXPRESSES HIGH-LEVELS OF STEM-CELL ANTIGEN SCA-1/

The murine clonal preadipose cell line, MC3T3-G2/PA6 (PA6), has the ab ility to support in vitro proliferation of hematopoietic stem cells de fined as colony-forming units in spleen (CFU-S). In order to ascertain the relationship between the hematopoietic-supporting activity of PA6 cells and their expression, we cultured a number of these cells for o ver 45 weeks and investigated the level at which they expressed severa l cell surface markers and membrane-bound growth factors. Besides expr essing stem cell factor (SCF) and macrophage colony-stimulating factor (M-CSF), PA6 cells were found by flow cytometry analysis to express h igh levels of stem cell antigen-1 (Sca-1). The expression level of Sca -l in PA6 cells correlated with the ability of the latter to support h ematopoiesis, whereas no such correlation was observed in the case of SCF and M-CSF expression, A cDNA clone encoding the protein recognized by anti-Sca-1 antibody was isolated from PA6 cells by expression clon ing, so that its nucleotide sequence encoded the protein identical to mouse alloantigen Ly-6A.2. Genetically engineered COS-7 cells, transfo rmed by the expression vector carrying the Ly-6A.2 gene, suppressed pr oliferation of murine lineage marker-negative (Lin(-)) bone marrow cel ls by themselves and synergistically augmented proliferation of these cells ill the presence of SCF. These results suggest that Ly-6A.2 regu lates the proliferation of hematopoietic progenitor cells, and is one of the molecules organizing the hematopoietic microenvironment provide d by stromal cells.