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Neutrophils are the primary cellular component of innate immunity. The ability of neutrophils to produce reactive oxygen species (ROS) is required for microbicidal activity, while excessive or inappropriate ROS generation contributes to tissue injury associated with ischemia-reperfusion injury and inflammatory diseases. Circulating neutrophils are typically poorly responsive to external stimuli. To allow appropriate responses to invading bacteria or other inflammatory stimuli, neutrophils undergo a phenotypic change, termed priming. Priming is defined as the ability of cytokines and chemotactic factors at non-stimulatory concentrations to induce a 10-20 fold increase in oxidative burst activity stimulated by a second exogenous stimulus. Although the molecular basis of priming remains to be established, two hypotheses are considered most likely candidates to explain this phenomenon. First, priming is associated with exocytosis of intracellular granules resulting in increased plasma membrane expression of receptors, signaling molecules, and the membrane components of the NADPH oxidase. Second, priming is associated with phosphorylation of cytosolic components of the NADPH oxidase, leading to enhanced assembly of this multisubunit enzyme upon subsequent stimulation. The p38 MAPK signal transduction pathway is required for neutrophil priming, and it participates in both granule exocytosis and NADPH oxidase component phosphorylation. Due to the inability to independently inhibit exocytosis and NADPH oxidase phosphorylation, studies directly testing the contribution of these two postulated mechanism to priming have not been performed. Additionally, whether p38 MAPK participation in neutrophil priming involves one or both of these potential mechanisms has not been determined.

My laboratory is currently working on the role of exocytosis in neutrophil priming by directly inhibiting the interaction of the SNARE proteins that mediate granule exocytosis. We are using a novel approach, generating TAT-fusion peptides mimicking the two coiled-coil domains of SNAP-23 and the coiled-coil SNARE domain of other SNARE proteins such as syntaxin 4 and VAMP-2, to inhibit neutrophil exocytosis. Our goal is to elucidate if granule exocytosis is necessary for neutrophil priming and also to determine if p38 MAPK activation contributes to priming mechanisms independently of exocytosis.