AbstractEffects of colony‐stimulating factors M‐CSF, GM‐CSF, G‐CSF, and IL‐3 were assessed on cells of macrophage lineage present in organ cultured 14‐day prenatal rat lungs. Treatment groups were compared between one another and against control lungs grown on standard medium containing 40 fetal bovine serum without added factors, where a monoculture of macrophages rapidly develops from precursors present at explantation, leading to appearance of a large mature population on the pleural surface outside the lungs. Studies were carried out in living cultures and by light and electron microscopy using peroxidase‐coupled isolectin B4ofGriffonia simplicifoliato identify macrophages and their precursors. In the first experiment, 14‐day prenatal lung explants (14+0 days) containingmacrophage precursorsbut not matured cells were exposed to individual CSFs for 7 days in an attempt to determine whether precursors are committed irrevocably to the macrophage line or can be altered by exposure to factors promoting significant granulocyte development. In succeeding experiments, 4‐ and 7‐day‐old cultures (14+4, 14+7 days) containingmatured macrophageswere targeted to see whether macrophage survival can be extended beyond expectations in controls and whether mitotic activity is stimulated. Recombinant CSFs were used at dosage levels known to promote colony formation in vitro (200–1,000 CFU/ml).Cultures exposed from prenatal day 14to M‐, GM‐, G‐CSF, or IL‐3 yielded a monoculture of macrophages without exception. Populations developed in the presence of M‐ or GM‐CSF were much larger than in controls or cultures grown with the other blood factors. GM‐CSF‐exposed cultures produced by far the largest macrophages, among them many multinucleate giant cells. Macrophages developed in the presence of G‐CSF were also significantly larger than controls.Growth of the mature macrophage populationwas greatly stimulated by exposure to M‐CSF or GM‐CSF but not by IL‐3 or G‐CSF. Mitotic figures were noted in the coronas of emerged cells surrounding stimulated cultures, compared to none in the controls. Ultrastructurally, macrophages stimulated by M‐CSF retained a mature appearance like macrophages in control, IL‐3, and G‐CSF treatment groups, whereas many in the GM‐CSF group became less differentiated. As tolong‐term survival, a single 14‐day explant was grown for 8 days on standard medium (the equivalent date for birth), then placed in a soft agar medium containing M‐CSF. Supplemented irregularly by M‐CSF and GM‐CSF, the culture remained viable until fixed on the 137th “postnatal” day and retained a small population of macrophages. Conclusions: (1) the macrophage lineage from embryonic rat lungs can be manipulated in culture; (2) macrophage precursors in these lungs seem committed to the macrophage line; (3) replication of both immature and mature macrophages is stimulated by M‐CSF and GM‐CSF; (4) with M‐CSF, however, retention of mature characteristics and longevity are favored, whereas with GM‐CSF maturity
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