Gram-negative sepsis continues to cause substantial morbidity and mortality. Much of the pathophysiology is believed to be caused by toxic bacterial cell wall molecules, including lipopolysaccharides (LPS), that cause direct toxicity and/or induce a secondary pro-inflammatory response. Our preliminary data indicate that LPS is released from dying bacteria that blood in complexes that contain three conserved outer membrane proteins (OMPs), and that these OMPs may have biological activity themselves. The three bacterial membrane proteins have been identified as outer membrane protein A (OmpA), peptidoyglycan associated lipoprotein (PAL), and murein lipoprotein (MLP). The overall objective of the proposal is to evaluate the role(s) of OmpA, PAL, and MLP in Gram-negative sepsis, with the ultimate goal of developing a strategy of treating sepsis with cross-reacting anti-OMP antibodies. Our first specific aim is to expand studies suggesting that there is a form of PAL released from the bacterial surface that floats at low density together with human lipoproteins. Our third specific aim is to study monoclonal and polyclonal IgG directed to epitopes on OmpA, PAL, and MLP for their ability to bind and neutralize the biological effects of each outer membrane protein and to promote opsinophagocyosis of LPS/OMP complexes. Our final specific aim is to study the protective efficacy of monoclonal and polyclonal immunoglobulin G directed to OmpA, PAL, and MLP in two models of Gram-negative infection. The studies should advance our fundamental understanding of the pathophysiology of Gram-negative sepsis. Each of the OMPs are highly conserved amongst clinical strains of Gram-negative bacteria. Therefore, a successful outcome of the project would provide a direct and immediate approach to developing anti-OMP antibodies for human use for the treatment of sepsis.
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