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Involvement of CSF‐1 in generating a stroma‐independent hematopoietic stem cell line

The hematopoietic stem cell line, Myl‐D7, is maintained by a self‐renewing stem cell population that spontaneously generates myeloid, lymphoid, and erythroid progeny. MS‐5 stromal cells are necessary for the growth of Myl‐D7 cells. One component of the Myl‐D7 cells proliferation activity released by MS‐5 stromal cells was enriched by Q sepharose fractionation and shown to be colony stimulating factor‐1 (CSF‐1) by Western blotting, BAC1.2F5 cell bioassay and inhibition of Myl‐D7 proliferation by CSF‐1 antibody. The requirement of Myl‐D7 cells for CSF‐1 was also demonstrated independently by selecting for rare, stroma‐independent Myl‐D7 mutant clones able to grow without stroma and additional factors. Eighty‐nine stroma‐independent mutant clones were obtained and belonged to two classes. The majority of mutants did not secrete any growth promoting activity. The second, rarer class of mutants releases a factor that stimulates proliferation/survival for up to several months and approximately half of the secretors express high levels of CSF‐1 mRNA. Wild type Myl‐D7 grown with supernatants from the secretor cells retained the stem cell phenotype. These data suggest that CSF‐1 may act as a key factor in stroma‐regulated hematopoiesis and cell–cell interaction. J. Cell. Physiol. 206: 556–562, 2006. © 2005 Wiley‐Liss, Inc.