It is generally agreed that bacterial lipopolysaccharide (LPS) is responsible for the initiation of the pathophysiologic changes which occur in patients with gram-negative septicemia. The high mortality rate in this patient group contributes to one of the leading causes of death in the United States. The long-term objectives of this proposal are to determine the molecular mechanisms of the interactions of LPS with host cells and how these interactions relate to the development of LPS-induced injury. The major objective of this research is to determine the biochemical properties of potentially injurious mediators produced by LPS-stimulated macrophages (MPhi) and to evaluate the pathogenic significance of these mediators in LPS-induced injury. This will be accomplished by establishing a number of in vitro systems to perform quantitative, biochemical studies. The proposed studies will utilize explanted rabbit hepatic macrophages (H-MPhi), murine peritoneal MPhi and MPhi cell lines to study a number of selected properties of LPS-stimulated MPhi. Specifically we will determine the biochemical properties, mechanism of action and pathogenic significance of a membrane bound procoagulant activity (PCA) identified in LPS stimulated H-MPhi and of a factor in the supernatants of LPS-stimulated MPhi which suppresses steroidogenesis in explanted rabbit adrenocortical cells. These studies will include purification of the H-MPhi PCA and of the MPhi factor responsible for inhibiting steroidogenesis. Studies will also be performed to characterize the molecular basis of LPS binding and uptake by MPhi. These experiments will include isolation and characterization of LPS receptors from MPhi membranes and ultrastructural analysis at the E.M. level using immunocytochemistry of LPS/MPhi interactions.
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