Human neutrophils are the first line of cellular immunologic defense responding to a wide variety of stimuli, in many cases independently of a specific immune response. While the mechanisms of neutrophil activation by certain highly specific stimuli (e.g. bacterial peptides) have been extensively studied, a diversity of activation pathways can be predicted. We propose to use influenza A virus (IAV) as a tool to 1) understand how neutrophils may play a role in contaminant of virus, 2) elucidate an activation pathway which differs in important respects from that excited by other well studied stimuli and 3) understand how these interactions of IAV with the neutrophil result in the clinically important outcome of neutrophil deactivation and bacterial superinfection. Using intact IAV preparations we want to a) establish the biochemical basis of the atypical respiratory burst response elicited by IAV using cell free NADPH oxidase preparations, b) fully characterize the pertussis-toxin insensitive phospholipase C (PLC) stimulation caused by IAV in terms of time course, sensitivity to inhibitors and possible dependence on G proteins other than that related to chemoattractant receptors, c) explain the lack of Ca2+ influx accompanying this PLC activation by examining diacylglycerol, phosphatidic acid and inositol tetra phosphate production. We want also to establish whether purified IAV hemagglutinin (HA) preparations will elicit activation and/or deactivation of human neutrophils. We hope through this means to establish a) if certain activation responses can be linked (or separated) from subsequent deactivation and b) whether crosslinking of cell surface components by multimeric HA preparations elicit activation (and/or deactivation) while monomeric HA does not. These studies in aggregate will lay the groundwork for a more focused study of IAV-induced neutrophil deactivation than is currently possible.
