Precursors of macrophages in embryonic rat lungs fail to exhibit granulocyte-forming potential.
BACKGROUND
Mesenchyme-like macrophage (M) precursors called angular cells are present in rat lungs on the thirteenth day of gestation and by then can differentiate into outright macrophages. Based on studies of bone marrow-derived cells, it is widely believed that the macrophage line necessarily proceeds from a colony-forming unit with dual granulocyte-macrophage potential (CFU-GM). In embryos this seems doubtful since macrophages are already scattered throughout the body before the first granulocytes appear. We examined the question in organ cultured 14 day prenatal rat lungs after having shown earlier that the macrophage population developed in explants is increased by exposure to M- and GM-colony-stimulating factors (CSFs) but is unaffected by multi (IL-3)- or granulocyte (G)-CSF. Reportedly retinoic acid (RA) shifts CFU-GM strongly towards granulocytic differentiation and inhibits mitosis of unipotential macrophage precursors but not differentiated cells. Transforming growth factor beta 1 (TGF) inhibits multipotential blood progenitors but allows proliferation of committed precursors, and TGF together with GM-CSF induces granulocytopoiesis from CFU-GM.


METHODS
Lung pairs were grown on a serum-containing medium or one supplemented either by RA, TGF, or TGF/GM-CSF to form a control and three experimental groups. A fourth experiment compared responses to M-CSF exposure and M-CSF/TGF. Macrophage population growth was estimated by measuring the areas of coronas formed by macrophages emerged from the explants. F-actin was stained with fluorescein-labeled phalloidin.


RESULTS
In all experiments macrophages were produced unmixed with granulocytes. By +8 days they had largely emerged to form coronas about the lungs. In cultures exposed to RA, macrophages were less intensely stained for actin and slower to emerge than controls. At +8 days, however, coronal areas were not significantly different from controls, as was also true for the TGF group. In contrast, coronal areas of cultures grown with TGF/GM-CSF were much larger. At +17 days, mean coronal area of TGF cultures was about half that of controls (P < 0.05), whereas mean coronal area of the TGF/GM-CSF group was 5.4 times greater (P < 0.001). Macrophages from control and TGF-exposed cultures responded to M-CSF by an increase in coronal area which was greater among cultures given M-CSF alone than those given TGF + M-CSF (both P < 0.005).


CONCLUSIONS
Macrophage precursors in embryonic lungs are distinct from CFU-GM.
Sorokin
S P
SP
Department of Anatomy and Neurobiology, Boston University School of Medicine, Massachusetts 02118.
McNelly
N A
NA
Hoyt
R F
RF
Svoboda
K K
KK
D000818:Animals; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006098:Granulocytes / Q000166:cytology* / Q000187:drug effects; D008168:Lung / Q000166:cytology / Q000187:drug effects / Q000196:embryology* / Q000276:immunology; D016173:Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology* / Q000187:drug effects; D008856:Microscopy, Fluorescence; D009924:Organ Culture Techniques; D051381:Rats; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014212:Tretinoin / Q000494:pharmacology
D016212:Transforming Growth Factor beta; D014212:Tretinoin; D016173:Macrophage Colony-Stimulating Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor