Pseudomonas Aeruginosa is a virulent opportunistic agent that in severely ill patients, causes an acute pneumonia that is associated with a high fatality rate. This application aims at in vivo and in vitro studies of basic mechanisms of Pseudomonas pneumonia. In the in vivo model, live Pseudomonas bacteria are instilled in the lungs of rats. Lung damage is quantified by measuring protein efflux from the airspace into the circulation, protein influx from the circulation to the airspace, production of infected pleural effusions, dissemination of systemic infection, and increased extravascular lung water. The in vitro model uses Madin-Darby canine kidney (MDCK) cells grown on filters. Live Pseudomonas are added to the apical surface of the MDCKS cells. The bacteria attach and kill the cells. The infection spreads centripetally. Cell damage is assessed morphologically and by the leakage of 14C-inulin across the permeabilized monolayer. The MDCK system, in common with in vivo pneumonia, shows dependence on exoenzyme S or a co-regulated protein for virulence. In this proposal, the in vitro and the in vivo systems will be combined to investigate host cell factors that determine susceptibility to Pseudomonas pneumonia. The four aims of the proposal are: (1) to determine how Pseudomonas attaches to cells, spreads from cell to cell and the role of cell invasion in infection, (2) to test the induction of intracellular signalling mechanisms by Pseudomonas; (3) to examine the role of glycolipids and glycoproteins on the mammalian cell surface as receptors for the bacteria, and determine if modification of these receptors alters bacteria-cell wall interaction; (4) to investigate how liposomes protect cells from Pseudomonas.
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