This FIRST award proposal investigates alterations in intracellular signal transduction as molecular mechanisms in sepsis and endotoxemia. Gram- negative sepsis is a leading cause of death worldwide. Treatment remains supportive. New and more effective therapies will require defining the pathogenesis of sepsis at molecular levels within the cell. The central hypothesis of this proposal is that specific intracellular signal transduction pathways are altered in macrophages upon repetitive endotoxin (LPS) stimulation. The activation states of these pathways ultimately protect or harm the host by exerting translational control of cytokine gene expression. The specific aims of this proposal focus on distinct LPS-signaling pathways which diverge at inositol phosphate turnover. The pathway LPS chooses to take is proposed to determine the cellular LPS response. Using molecular biology tools applied to cell culture and animal models of LPS presensitization (tolerance), we propose to confirm or exclude the following: 1) Altered inositol phosphate signaling limits LPS responses; 2) LPS signaling is linked to translational control of cytokine gene expression through phosphorylation of specific cytoplasmic proteins; 3) Upon repetitive LPS stimulation, cell signaling shifts away from these pathways allowing the cell and host to regulate its response while confronting the bacterial threat. In a clinically relevant model of experimental sepsis, we propose to pharmacologically redirect cellular signaling and provide a protective survival benefit. Preliminary data suggest the central hypothesis is correct, and moreover, point to an unexpected link between LPS and insulin as extracellular ligands: co-utilization of signaling pathways coupling ligand binding with the translational control of cellular protein synthesis.