.): Sepsis and multiple organ dysfunction syndrome (MODS) are the leading causes of mortality in critically ill surgical patients. TNF and IL-1 are pro-inflammatory cytokines produced by endotoxin- stimulated macrophages and are known to be important mediators in sepsis. The PI proposes that this regulated production of pro-inflammatory cytokines is a cardinal feature of sepsis leading to MODS. This is the result of sequential exposure to multiple inflammatory insults over time. The PI hypothesizes that a tolerant macrophage phenotype is the relevant macrophage response to inflammatory stimuli in patients with sepsis or MODS. Using an in vitro mouse macrophage model, they have previously demonstrated profound modifications in LPS-activated macrophage functions following LPS pretreatment. These LPS-tolerant alterations include: 1) an inhibition of TNF secretion; 2) augmentation of IL-1 secretion; 3) impaired activation of ERK1 and 2 and T38 mitogen activated kinase (MAPK); and 4) altered regulation of NF-kB. The present studies are designed to systematically investigate: 1) the mechanisms by which LPS pretreatment induces LPS tolerance; 2) the specific defects responsible for the altered response to LPS; and 3) to characterize the signal transduction alterations present in monocytes of patients with trauma, sepsis, or burns. The PI will focus on LPS signal transduction pathways that can be stimulated in the absence of serum or CD14, and do not require protein tyrosine kinases, MAPK, or NF-kB activation. They propose to extend previous investigations using mouse macrophages, human monocytic cell lines, and monocytes from clinical trauma patients, burn patients, or patients with sepsis. The previous results suggest that relatively early LPS activation signaling events depend on upstream MAPK activation and act by changes in both pre- and post-translational transcriptional processing. The PI will use specific agonists and antagonists to delineate the precise signal transduction alterations present in LPS-tolerant cells compared to naive macrophages. They hope that using monocytes from patients will help them to determine when and how alterations in signal transduction arise and how they can develop approaches to counteract or modulate macrophage-regulated cytokine production.
