ENDOTOXIN-NEUTRALIZING PROTEIN PROTECTS AGAINST ENDOTOXIN-INDUCED ENDOTHELIAL BARRIER DYSFUNCTION

Bacterial lipopolysaccharide induces tyrosine phosphorylation of paxil lin, actin reorganization, and opening of the transendothelial paracel lular pathway through which macromoles flux. In this study, lipid A wa s shown to be the bioactive portion of the lipopolysaccharide molecule responsible for changes in endothelial barrier function. We then stud ied whether endotoxin-neutralizing protein, a recombinant peptide that is derived from Limulus antilipopolysaccharide factor and targets lip id A, could block the effects of lipopolysaccharide on protein tyrosin e phosphorylation, actin organization, and movement of C-14-bovine ser um albumin across bovine pulmonary artery endothelial cell monolayers. In the presence of serum, a 6-h exposure to lipopolysaccharide (10 ng /ml) increased transendothelial C-14-albumin flux compared to the simu ltaneous media control. Coadministration of endotoxin-neutralizing pro tein (greater than or equal to 10 ng/ml) with lipopolysaccharide (10 n g/ml) protected against lipopolysaccharide-induced barrier dysfunction . This protection was dose dependent, conferring total protection at e ndotoxin-neutralizing protein/lipopolysaccharide ratios of greater tha n or equal to 10:1. Similarly, endotoxin-neutralizing protein was capa ble of blocking the lipopolysaccharide-induced endothelial cell respon ses that are prerequisite to barrier dysfunction, including tyrosine p hosphorylation of paxillin and actin depolymerization. Finally, endoto xin-neutralizing protein cross-protected against lipopolysaccharide de rived from diverse gram-negative bacteria. Thus, endotoxin-neutralizin g protein offers a novel therapeutic intervention for the vascular end othelial dysfunction of gram-negative sepsis and its attendant endotox emia.