ISOLATION AND CHARACTERIZATION OF C-FOS-EXPRESSING MURINE BONE-MARROWSTROMAL CELL-LINES SUPPORTING MYELOID DIFFERENTIATION

We have previously reported that constitutive expression of c-fos onco gene allows long-term proliferation of primary mouse bone marrow strom al cells favoring the granulocytic differentiation of myeloid precurso rs in an in vitro assay. Retrovirus-mediated gene transfer of the huma n c-fos gene was used here for immortalizing nine mouse bone marrow ce ll lines which were studied in detail. However, due to low expression of the ectopic c-fos gene, none of them showed characteristics of tran sformation as assayed by dependence upon serum for growth, the inabili ty to form colonies in agar and contact inhibition. All of them displa yed a fibroblastoid phenotype, as deduced from morphological observati on and analysis of several differentiation markers, They mostly suppor ted the granulocytic differentiation of bone marrow myeloid precursors in a GM-assay, as did c-fos-expressing primary stromal cells. Their p otential for supporting myeloid progenitor proliferation was however s ignificantly lower than that of the whole adherent layer of the Dexter -type long-term bone marrow culture they derived from (STNT cells). Th ey showed significant variations with respect to their cytokine gene e xpression analyzed at the RNA level in keeping with the notion of stro mal cell heterogeneity in the bone marrow. Interestingly, none of them secreted GM-CSF, SCF or IL-3, which are cytokines reputed for their a bility to stimulate hematopoietic progenitors, and, strikingly, only t wo of them were able to produce detectable levels of GCSF in culture s upernatants despite the propensity of all of them to favor granulocyte differentiation. Finally, in a coculture assay, bone marrow cells wer e able to diminish the expression of several cytokine genes albeit at a much lower degree than in the original STNT cells.