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 ability to support in vitro proliferation of hematopoietic stem cells defined 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 over 45 weeks and investigated the level at which they expressed several cell surface markers and membrane-bound growth factors. Besides expressing stem cell factor (SCF) and macrophage colony-stimulating factor (M-CSF), PA6 cells were found by flow cytometry analysis to express high levels of stem cell antigen-1 (Sca-1). The expression level of Sca-1 in PA6 cells correlated with the ability of the latter to support hematopoiesis, 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 cloning, so that its nucleotide sequence encoded the protein identical to mouse alloantigen Ly-6A.2. Genetically engineered COS-7 cells, transformed by the expression vector carrying the Ly-6A.2 gene, suppressed proliferation of murine lineage marker-negative (Lin) bone marrow cells by themselves and synergistically augmented proliferation of these cells in the presence of SCF. These results suggest that Ly-6A.2 regulates the proliferation of hematopoietic progenitor cells, and is one of the molecules organizing the hematopoietic microenvironment provided by stromal cells.
Satoh
M
M
Tokyo Research Laboratories, Kyowa Hakko Kogyo, Co., Ltd., Machida-shi, Japan.
Mioh
H
H
Shiotsu
Y
Y
Ogawa
Y
Y
Tamaoki
T
T
D000818:Animals; D000950:Antigens, Ly / Q000032:analysis* / Q000235:genetics; D001483:Base Sequence; D001853:Bone Marrow / Q000378:metabolism*; D019556:COS Cells; D002455:Cell Division; D002460:Cell Line; D003001:Cloning, Molecular; D018920:Coculture Techniques; D005434:Flow Cytometry; D015870:Gene Expression; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000032:analysis* / Q000235:genetics; D051379:Mice; D008811:Mice, Inbred DBA; D008969:Molecular Sequence Data; D019089:Stem Cell Factor / Q000494:pharmacology; D017154:Stromal Cells / Q000378:metabolism*; D014162:Transfection
D000950:Antigens, Ly; C074650:Ly6a protein, mouse; D008565:Membrane Proteins; D019089:Stem Cell Factor