Alterations in transcription factor binding at the IL-2 promoter region inanergized human CD4(+) T lymphocytes

Background. The mechanisms responsible for the induction of clonal anergy a re not well understood. We have utilized an in vitro model of human T cell anergy to explore the perturbations in cell signaling at the level of inter leukin (IL)-2 gene transcription and to define the contribution of other cy tokines to this effect. Methods. An in vitro model of clonal anergy was established by using CD4(+) T lymphocytes from healthy human donors. Cells were anergized by prestimul ation with an anti-CD3 monoclonal antibody (mAb) followed by restimulation 72 hr later with anti-CD3 mAb with or without anti-CD28. Results. CD4(+) T cells, anergized with anti-CD3 monoclonal antibody (OKT3) prestimulation, displayed a marked reduction in proliferation (P=0.0036) a nd IL-2 production (P <0.0001). Co-incubation with IL-10 reduced cellular p roliferation in OKT3/CD28 pretreated cells by 19% (P=NS) and reduced IL-2 p roduction by 40% (P=0.0024). Anergized T cells demonstrated a reduced bindi ng activity of the AP-1 complex to the IL-2 promoter. Supershift experiment s and Western blots confirmed that the binding of c-Fos, JunB, and JunD, bu t not of FosB, was reduced in anergized cells. At the sis-inducible element (SIE)-binding region of the c-Fos promoter, Stat3 binding was reduced. Conclusions. T cell anergy, induced by prestimulation with OKT3, is charact erized by reduced proliferation and a profound decrease in IL-2 production. Anergy can be prevented by co-incubation with anti-CD28 and partially re-e stablished by IL-10. Anergy is accompanied by a reduction in AP-1 binding t o the IL-2 promoter, with selective reduction in binding of c-Fos, JunB, an d JunD. Defective binding for Stat3 at the c-Fos promoter suggests an invol vement of the Jak-Stat pathway.