DIFFERENTIATION OF CANINE BONE-MARROW CELLS WITH HEMATOPOIETIC CHARACTERISTICS FROM AN ADHERENT STROMAL CELL PRECURSOR

Stromal cell lines were established from canine long-term marrow cultu res, cloned by limiting dilution, and maintained in stromal cell-condi tioned medium. These cells grew adherent, maintained stable growth rat e and morphology under standard conditions (in 20-30% conditioned medi um; confluency, 70-90%), and supported hemopoiesis in long-term marrow cultures. In the presence of exogenous recombinant canine stem cell f actor (rcSCF), round cells developed from the adherent layer, detached , and remained in culture as viable floating cells. Round floating cel ls also appeared when cultures were grown to >90% confluency without r cSCF. Round cells were smaller than adherent cells, expressed CD34, sh owed basophilic plasma, and stained positive for c-kit, MHC-class II m arkers, and myeloid markers. In standard assays for colony formation, the detached cells produced granulocyte-macrophage colony-forming unit s (CFU-GM), fibroblast colony-forming units (CFU-F), and less well-def ined colony-forming units. In addition, on allogeneic feeder cells in long-term cultures, these cells generated hemopoietic colonies. Striki ngly, the differentiation was reversible: when nonadherent cells were resuspended at lower density in serum-containing medium, they reattach ed and grew to confluence when, once again, round cells detached. Deta ched cells from this secondary cycle produced mainly CFU-F and few CFU -GM when placed in clonal assays. These results suggest that some fibr oblast-like stromal cells have the potential to differentiate into cel ls with hemopoietic characteristics. These observations provide eviden ce for the existence of a quiescent precursor of hemopoietic progenito rs in the bone marrow stroma of the adult dog.