Multiple organ failure (MOF) appears to follow systemic inflammatory mediator activation related to trauma, burns, sepsis, acute pancreatitis, and ischemia/reperfusion (I/R) injury. MOF is the single greatest cause of mortality among surgical patients in the United States today and acute respiratory failure accounts for the majority of this. This proposal examines the pathogenesis of MOF-related injury to the pulmonary microvasculature following intestinal I/R injury in rats. While the specific mechanisms of cellular injury are not known, it appears that humoral and local mediators in the gut and liver regulate a systemic response which targets the lung. The proposed studies will examine the early and fundamental interactions of the complement system and key cytokines (IL-l and TNFalpha), with neutrophils and pulmonary microvascular endothelial cells. While mediators such as endotoxin, other cytokines, and PAF are involved in this injury, these appear to be either late or secondary events in the process of cellular injury. Because therapies directed at the initiating events ale more likely to be successful ultimately, the principal effort of the proposed studies is to specifically examine the critical early events in the process of pulmonary microvascular endothelial cell injury. The role of complement will be examined in Specific Aim l. In vivo complement inhibition using both conventional and novel strategies will be assessed for efficacy in preventing acute lung injury. In addition, in vitro assessment of the pulmonary microvascular cell response to complement activation will be done. A newly developed polymerase chain reaction (PCR)-based assay for specific mRNA in tissue will be used to quantitatively estimate cytokine and complement synthesis in lung, liver, and gut tissue. These data will be correlated with the appearance in plasma of complement fragments, TNFalpha and IL-1.
In Specific Aims 2 and 3, using specific antibodies and other antagonists, the relationship of TNFalpha, IL-1 to the development of acute pulmonary microvascular injury will be determined. Plasma from rats with intestinal I/R injury will be incubated with rat pulmonary microvascular endothelial cells and injury assessed using functional, biochemical, and morphometric means. Preliminary data show that I/R plasma contains factors which deplete endothelial cell ATP and that when combined with neutrophils, produce cytotoxicity. In vivo and in vitro assessment of neutrophil-mediated endothelial cell injury will be directed towards identifying the particular mechanisms of neutrophil-derived oxidant and protease induced injury. A new feature of this revised proposal is the examination of complement, TNFalpha, and IL-1 induced alterations in specific pulmonary microvascular endothelial cell adhesion molecules which regulate neutrophil interactions. It is anticipated that the proposed studies will provide information regarding the specific pathogenic mechanisms responsible for the development of acute microvascular lung injury following intestinal I/R injury. It is possible that these events will have broader relevance to the MOF syndrome.
