The morbidity and mortality from severe meningococcal disease remain high despite effective microbicidal therapy and improved intensive care. Meningococcal endotoxin, lipooligosaccharide (LOS) and the propensity of the organism to shed large amounts of LOS-containing membrane vesicles have been implicated in the genesis of the dramatic and devastating clinical disease. Our integrated program examines fundamental aspects of severe meningococcal disease, using Neisseria meningitidis serogroup B (NMB) as the representative organism, because there is no effective human vaccine, and polymorphonuclear leukocytes (PMNs) and endothelial cells (EC) as the host targets, because innate host defenses are critical early in intravascular infection. The Program's hypothesis is that the LOS of NMB endotoxin, either as membrane blebs or intact bacteria, elicits quantitatively and/or qualitatively different responses that does purified LOS or endotoxin from other gram-negative organisms. Within this framework, the focus in Project 3 is host cellular response and studies will be directed at the following specific aims: 1. To characterize features of the PMN response which mediate the killing and/or clearing of ingested NMB detoxification of LOS. Studies examine the impact of priming of PMN on the intracellular fate of ingested NMB, the subcellular distribution or phosphorylation state of p47phox, the ability of LOS exposure to subvert or enhance ingestion, degranulation, intracellular microbicidal events, and/or degradation of LOS, intracellular microbicidal microbicidal events, and/or degradation of LOS. 2. To characterize features of the EC response which mediate and/or clearing of internalized NMB and detoxification of LOS. Studies assess the structural features of LOS which influence in EC: the fate of internalized NMB and detoxification NMB and detoxification of LOS, the LOS-stimulated pro-coagulant activity, the release of pro-inflammatory cytokines, expression of adhesion molecules, and the integrity of EC monolayers. 3. To define host determinants which figure in the extracellular responses to invasive NMB. Studies examine the role of terminal components of complement, alone and in the simultaneous presence of PMN-derived anti-bacterial proteins (e.g. bactericidal permeability increasing protein), in direct killing as well as opsonophagocytosis of NMB, the ability of LOS/NMB-adherent PMNs and their products on the intracellular fate of NMB and detoxification of LOS. Taken together these studies complement and extend those in Projects 1 and 2 testing how the structural determinants of NMB LOS and its presentation mediate responses of PMNs and EC. We are confident that studies in this model system will advance understanding of innate host responses, beneficial and pathophysiologic, in other clinically relevant situations.
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