Nitric Oxide (NO) has been identified as an endothelial cell derived-relaxing factor and the pathway responsible for its synthesis in human cells has been elucidated. Further, antagonists of this pathway have been shown to increase blood pressure in animals challenged with tumor necrosis factor and endotoxin, important mediators of septic shock. In these investigations, the NO pathway is being examined in normal volunteers given endotoxin both intravenously and intrabronchially. We are looking for evidence of nitric oxide synthase (NOS) induction during systemic and local inflammatory responses to clarify the role of this pathway in human disease. The initial phase of this project began in normal volunteers given either sodium nitroprusside or endotoxin (Vandivier, R.W. et al., American Journal of Respiratory Critical Care Medicine, 153:A54, 1996). Methods were developed to measure exhaled NO, and serum and urine nitrite/nitrate and cyclic guanosine monophosphate (cGMP) concentrations. We found that sodium nitroprusside produced a fall in mean arterial pressure (MAP) and measurable increases in exhaled NO and urinary nitrite/nitrate and cGMR In contrast, endotoxin produced a fall in MAP, but no measurable changes in the activity of the NO path way. Current studies are examining the role of the NO pathway in pulmonary inflammation after intrabronchial administration of endotoxin. This administration produces a localized inflammatory response characterized by neutrophil recruitment and cytokine production. These experiments are being conducted in conjunction with in vitro studies examining the role of NO in modulating the interaction between human neutrophils and bronchial epithelial cells. Methods are being developed for NOS Western blot, ribonuclease protection assays, and NOS enzyme assays for use on clinical specimens.
