Interactions among granulocyte-macrophage colony-stimulating factor, macrophage colony-stimulating factor, and IFN-gamma lead to enhanced proliferation of murine macrophage progenitor cells.

This report examines the actions of IFN-gamma on monocytopoiesis in murine liquid and semisolid bone marrow cultures. The proliferative response of bone marrow cells to macrophage CSF and granulocyte-macrophage CSF was assayed by measuring [3H]TdR uptake in a range of mouse strains. No interstrain difference in kinetics was observed for CSF-1 action, but GM-CSF acted significantly more rapidly on C57B1/6, Swiss, and to a lesser extent A/J mice than on BALB/c or CBA. IFN-gamma inhibited [3H]TdR incorporation elicited by CSF-1, and to a much lesser extent, GM-CSF. When the two CSF were added together, the effects were not additive; in fact, the response was the same as that seen with GM-CSF alone. When IFN-gamma was also added, the response was restored to the level seen with CSF-1 alone. In essence, the inhibitory actions of GM-CSF and IFN-gamma were mutually exclusive. The mechanism of these actions was investigated using colony assays. As expected, CSF-1 caused the formation of pure macrophage colonies, whereas GM-CSF stimulated production of macrophage, granulocyte, and mixed granulocyte macrophage colonies. When the two CSF were added in combination, the total colony count was greater than with either alone, but less than additive. The number of pure macrophage colonies was reduced to the number seen with GM-CSF alone. IFN-gamma reduced the number of colonies in the presence of CSF-1, but slightly increased the number with GM-CSF. In the presence of both CSF, IFN-gamma increased the colony count by around 25 to 40%, so that the numbers were greater than the combined total of CSF-1 plus GM-CSF added separately. Similar results were obtained in all mouse strains tested. The results suggest that the thymidine uptake data reflect changes in the number of progenitor cells responding rather than changes in cell cycle time. The results are discussed in terms of the possibility that coadministration of GM-CSF and CSF-1 could ameliorate the myelosuppressive actions of IFN-gamma in vivo, leading to more effective use of this agent as a biologic response modifier.

The effects of interleukin 3 (IL-3) on cells responsive to macrophage colony-stimulating factor (CSF-1) in liquid murine bone marrow culture.

We have examined variation between inbred mouse strains in the proliferative response of bone marrow cells in liquid culture to macrophage colony-stimulating factor (CSF-1) and interleukin 3 (IL-3). In all mouse strains, thymidine incorporation was stimulated by CSF-1 and, after an initial lag period, it reached a peak on day 5. In contrast, two mouse strains, A/J and Balb c, had much lower proliferative responses to IL-3 than did the other strains. In A/J there was no increase in thymidine incorporation above the initial baseline, although the fall in incorporation seen in the absence of any added growth factor was prevented. IL-3 also prevented the loss of CSF-1 responsiveness observed when A/J bone marrow cells were incubated in medium alone. The lag phase in the response to CSF-1 was progressively abolished following IL-3 pre-treatment. Thus, the data with A/J mice separate two distinct activities of IL-3 and show that proliferation is not required for the synergistic effect exerted by IL-3 on CSF-1-stimulated macrophage generation from bone marrow. In strains in which IL-3 alone was able to stimulate proliferation, its action was not additive with that of CSF-1, and addition of both factors together did not overcome the lag phase. This suggests that the two factors act on the same cell population, and that the known synergistic effect of IL-3 on macrophage colony formation in soft agar does not result from an increase in the initial rate of proliferation. The possibility that the combination of factors might alter the duration of the growth response in vivo is discussed.

Eicosanoids produced during interactions between Pseudomonas aeruginosa and alveolar macrophages are species-dependent.

Eicosanoid production during phagocytosis of pyogenic bacteria by rabbit alveolar macrophages was studied as a model of early events in the pathogenesis of pneumonia. Adherent alveolar macrophages, prelabelled with [3H]-arachidonic acid (AA), were incubated with live, opsonized Staphylococcus aureus or Pseudomonas aeruginosa (bacteria:macrophage ratio of 50:1) at 37 degrees C for 90 min. Supernatant eicosanoids were extracted and separated by reverse phase high performance liquid chromatography (RP-HPLC). While the amounts of labelled PGE2, TXB2, and PGD2 produced in response to the two organisms were equal, the amount of PGF2 alpha elicited by S. aureus amounted to three times that released during macrophage challenge with P. aeruginosa. Overall, preferential release of cyclooxygenase products occurred during phagocytosis of S. aureus. In contrast, eicosanoids identified presumptively as oxygenated metabolites of AA predominated in cultures challenged with opsonized P. aeruginosa. Live, non-opsonized P. aeruginosa elicited the same profile of eicosanoids, but in reduced amounts. Inhibitor studies indicated that these AA derivatives were not synthesized via the macrophage lipoxygenase pathway. Their production was dependent on the viability of P. aeruginosa. Macrophages challenged with opsonized, heat-killed P. aeruginosa resulted in production of an eicosanoid profile similar to that elicited by S. aureus. Secondary metabolism by P. aeruginosa of eicosanoids released from the macrophage did not contribute to the unique profile produced during the interaction of this organism with labelled macrophages. Our data indicate that during binding to macrophages, the primary human pathogen, P. aeruginosa, specifically modulates the profile of eicosanoids produced. This effect on inflammatory mediators may be of biological significance in the pathogenesis of pneumonia.