Minute numbers of contaminant CD8+ T cells or CD11b+CD11c+ NK cells are the source of IFN-gamma in IL-12/IL-18-stimulated mouse macrophage populations.
Macrophages were reported to be strong producers of interferon gamma (IFN-gamma) after stimulation by interleukin 12 (IL-12) plus IL-18, which gave rise to a novel concept of auto-crine macrophage activation. Here, we show that peritoneal exudate and bone marrow-derived mouse macrophages generated by conventional techniques contain small quantities of CD11b(+)CD11c(+)CD31(+)DX5(+)NK1.1(+) natural killer (NK) cells or CD3(+)CD8(+)TCRbeta(+) T cells, respectively. Intracellular cytokine staining, purification of macrophages by sorting, and the analysis of macrophages from alymphoid RAG2(-/-)gamma-chain(-/-) mice revealed that the high amount of IFN-gamma protein in the supernatants of unseparated IL-12/IL-18-stimulated macrophage populations originates exclusively from the contaminating lymphoid cells. Notably, IL-12/IL-18 still induced IFN-gamma mRNA in highly purified macrophages from wild-type mice and in macrophages from RAG2(-/-)gamma-chain(-/-) mice, whereas nuclear translocation of signal transducer and activator of transcription 4 (STAT4) and production of IFN-gamma protein were no longer detectable. These results question the concept of autocrine macrophage activation by secreted IFN-gamma, suggest differences in the expression of IFN-gamma mRNA and protein between macrophages and lymphoid cells, and illustrate that the limited purity of most myeloid cell populations (</= 98%) might lead to false conclusions.
Schleicher
Ulrike
U
Institute of Clinical Microbiology, Immunologu and Hygiene, University of Erlangen, Germany.
Hesse
Andrea
A
Bogdan
Christian
C
D000818:Animals; D039481:CD11b Antigen / Q000032:analysis*; D039521:CD11c Antigen / Q000032:analysis*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D005260:Female; D005434:Flow Cytometry; D007371:Interferon-gamma / Q000235:genetics* / Q000378:metabolism; D018664:Interleukin-12 / Q000494:pharmacology*; D020382:Interleukin-18 / Q000494:pharmacology*; D007694:Killer Cells, Natural / Q000166:cytology / Q000276:immunology*; D017737:Macrophages, Peritoneal / Q000166:cytology / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D013601:T-Lymphocytes / Q000276:immunology*
D039481:CD11b Antigen; D039521:CD11c Antigen; D020382:Interleukin-18; D018664:Interleukin-12; D007371:Interferon-gamma