In vivo production and function of IL-12 p40 homodimers.
The bioactivity of IL-12 is mediated by heterodimers of disulfide-linked p35 and p40 protein subunits. Homodimeric p40 competes with heterodimer for binding to the high affinity IL-12R and inhibits IL-12 bioactivity in vitro. However, the production and significance of p40 homodimer as a cytokine antagonist in vivo have not been determined. In these studies, we observed increased amounts of both IL-12 p40 monomer and homodimer in the serum of C57BL/6 mice following injection of 300 microg of Salmonella enteritidis LPS. Homodimer constituted between 20 and 40% of the total circulating p40 in endotoxemic sera, as confirmed by both Sephacryl S-100 gel filtration and p40-specific immunoprecipitation analyses. Similar relative amounts of homodimer and monomer were observed in endotoxemic BALB/c, C57BL/6, IFN-gamma-deficient C57BL/6 mice and C57BL/6 mice previously infected with bacille Calmette-Guérin. To determine whether IL-12 p40 homodimer was capable of antagonizing IL-12-dependent IFN-gamma responses in vivo, we pretreated C57BL/6 mice with purified rIL-12 p40 homodimer before i.p. challenge with endotoxin. Mice treated with 40 to 80 microg of p40 homodimer generated 80 to 82% less circulating IFN-gamma during acute endotoxemia than saline controls (p < 0.01). We conclude that p40 homodimer is produced in vivo, functions as a cytokine antagonist in the context of the mouse model of acute endotoxemia, and may represent a novel form of self-regulating cytokine response.
Heinzel
F P
FP
Division of Geographic Medicine, Case Western Reserve University School of Medicine, and Veterans Affairs Medical Center, Cleveland, OH 44106, USA.
Hujer
A M
AM
Ahmed
F N
FN
Rerko
R M
RM
D000818:Animals; D019446:Endotoxemia / Q000378:metabolism; D005260:Female; D007158:Immunologic Techniques; D007371:Interferon-gamma / Q000096:biosynthesis; D018664:Interleukin-12 / Q000037:antagonists & inhibitors / Q000737:chemistry*; D008070:Lipopolysaccharides; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008970:Molecular Weight; D011994:Recombinant Proteins; D012477:Salmonella enteritidis; D013329:Structure-Activity Relationship
D008070:Lipopolysaccharides; D011994:Recombinant Proteins; D018664:Interleukin-12; D007371:Interferon-gamma