Signaling mechanisms inducing hyporesponsiveness of phagocytes during systemic inflammation.

The inflammatory responsiveness of phagocytes to exogenous and endogenous stimuli is tightly regulated. This regulation plays an important role in systemic inflammatory response syndromes (SIRSs). In SIRSs, phagocytes initially develop a hyperinflammatory response, followed by a secondary state of hyporesponsiveness, a so-called "tolerance." This hyporesponsiveness can be induced by endotoxin stimulation of Toll-like receptor 4 (TLR4), resulting in an ameliorated response after subsequent restimulation. This modification of inflammatory response patterns has been described as innate immune memory. Interestingly, tolerance can also be triggered by endogenous TLR4 ligands, such as the alarmins myeloid-related protein 8 (MRP8, S100A8) and MRP14 (S100A9), under sterile conditions. However, signaling pathways that trigger hyporesponsiveness of phagocytes in clinically relevant diseases are only barely understood. Through our work, we have now identified 2 main signaling cascades that are activated during MRP-induced tolerance of phagocytes. We demonstrate that the phosphatidylinositol 3-kinase/AKT/GSK-3ß pathway interferes with NF-κB-driven gene expression and that inhibition of GSK-3ß mimics tolerance in vivo. Moreover, we identified interleukin-10-triggered activation of transcription factors STAT3 and BCL-3 as master regulators of MRP-induced tolerance. Accordingly, patients with dominant-negative STAT3 mutations show no tolerance development. In a clinically relevant condition of systemic sterile stress, cardiopulmonary bypass surgery, we confirmed the initial induction of MRP expression and the tolerance induction of monocytes associated with nuclear translocation of STAT3 and BCL-3 as relevant mechanisms. Our data indicate that the use of pharmacological JAK-STAT inhibitors may be promising targets for future therapeutic approaches to prevent complications associated with secondary hyporesponsiveness during SIRS.

Peroxisome Proliferator-Activated Receptor-γ Modulates the Response of Macrophages to Lipopolysaccharide and Glucocorticoids.

Although glucocorticoids (GC) represent the most frequently used immunosuppressive drugs, their effects are still not well understood. In our previous studies, we have shown that treatment of monocytes with GC does not cause a global suppression of monocytic effector functions, but rather induces differentiation of a specific anti-inflammatory phenotype. The anti-inflammatory role of peroxisome proliferator-activated receptor (PPAR)-γ has been extensively studied during recent years. However, a relationship between GC treatment and PPAR-γ expression in macrophages has not been investigated so far. Studies using PPAR-γ-deficient mice have frequently provided controversial results. A potential reason is the use of primary cells, which commonly represent inhomogeneous populations burdened with side effects and influenced by bystander cells. To overcome this constraint, we established ER-Hoxb8-immortalized bone marrow-derived macrophages from Ppargfl/fl and LysM-Cre Ppargfl/fl mice in this study. In contrast to primary macrophages, the ER-Hoxb8 system allows the generation of a homogeneous and well-defined population of resting macrophages. We could show that the loss of PPAR-γ resulted in delayed kinetic of differentiation of monocytes into macrophages as assessed by reduced F4/80, but increased Ly6C expression in early phases of differentiation. As expected, PPAR-γ-deficient macrophages displayed an increased pro-inflammatory phenotype upon long-term LPS stimulation characterized by an elevated production of pro-inflammatory cytokines TNF-α, IL1-ß, IL-6, IL-12 and a reduced production of anti-inflammatory cytokine IL-10 compared to PPAR-γ WT cells. Moreover, PPAR-γ-deficient macrophages showed impaired phagocytosis. GC treatment of macrophages led to the upregulation of PPAR-γ expression. However, there were no differences in GC-induced suppression of cytokines between both cell types, implicating a PPAR-γ-independent mechanism. Intriguingly, GC treatment resulted in an increased in vitro migration only in PPAR-γ-deficient macrophages. Performing a newly developed in vivo cell-tracking experiment, we could confirm that GC induces an increased recruitment of PPAR-γ KO, but not PPAR-γ WT macrophages to the site of inflammation. Our findings suggest a specific effect of PPAR-γ on GC-induced migration in macrophages. In conclusion, we could demonstrate that PPAR-γ exerts anti-inflammatory activities and shapes macrophage functions. Moreover, we identified a molecular link between GC and PPAR-γ and could show for the first time that PPAR-γ modulates GC-induced migration in macrophages.