Abstract
This study was designed to assess the presence of endogenous granulocyte-macrophage colony-stimulating factor (GM-CSF) within adherent layers of human Dexter-type cultures and to investigate the effect on granulomonopoiesis of adding exogenous GM-CSF to the culture medium. The presence of GM-CSF was demonstrated using a bioassay, in which adherent layers from normal bone marrows gave rise to endogenous granulocyte-macrophage colony-forming units (CFU-GM) that were specifically inhibited by increasing amounts of an anti-GM-CSF neutralizing antibody. Using an immunoassay, the estimated amounts of GM-CSF were less than or equal to 40 pg per flask in adherent layers, while remaining undetectable in supernatants. The addition of 10 ng or purified recombinant GM-CSF per milliliter of culture medium increased slightly the CFU-GM output over a 5-week culture period. The addition of 50 ng/mL decreased significantly the CFU-GM output after 5 weeks of culture. This decrease was associated with major modifications of the adherent layer cell composition. Large round or ovoid macrophages were generated at the expense of the interdigitated and elongated stromal cells and the extracellular fibronectin network was no longer observed. These studies suggest that GM-CSF production by accessory cells (stromal cells and/or monocytes) is almost equal to its consumption by hematopoietic cells, a situation similar to that found in long-term cultures of murine marrows. They also show that the maintenance of granulomonopoiesis is decreased by adding more than 10 ng/mL of exogenous GM-CSF to the culture medium, which is related to the induction of adherent macrophages, the disappearance of the major smooth-muscle-like stromal cell component of the adherent layer, and that of the fibronectin extracellular matrix.
