CFU-C proliferation and granulopoiesis in suspension cultures of bone marrow,cord blood and adult blood

Summary. Normal bone marrow, cord blood and adult blood cells were separated on ficoll-hypaque density gradients and cultured in MCoy 5A medium with 15% human serum in 25 cm² flasks for up to 14 days to characterize their growth. The system permits the study of granulopoiesis from the generation in culture of new CFU-C to the emergence of neutrophils and monocyte-macrophage cells at the end of the culture period. Bone marrow, cord blood and adult blood, showed 2-3-fold increases in CFU-C, but characteristically only small numbers of CFU-C were detected in adult blood, and only small numbers of granulocytes were produced. The total nucleated cell count only rose consistently in bone marrow cultures. Adherent cell layers formed in all cultures with confluent or near confluent growth of macrophages after 14 days. Some macrophages were multinucleated giant cells; these cells were found most commonly in adult blood cultures. Exclusively in bone marrow cultures macroscopic fibroblast colonies and smaller fibroblast clusters of about 20-50 cells occurred. CFU-C did not proliferate if they were deprived of adherent cells, but some growth did occur when bone marrow depleted of adherent cells was cultured on preformed marrow or fibroblast-free peripheral blood adherent layers. These allogeneic adherent cells were, however, less stimulatory than the autologous cells. It is concluded that CFU-C progenitors are detectable in blood and bone marrow, while fibroblast colony forming cells do not circulate. It appears that macrophages in the absence of fibroblasts will support some CFU-C generation and granulopoiesis in suspension culture. This system is a useful model for examining the cellular and humoral regulation of granulopoiesis in vitro and further studies are in progress.