The broad objective of this proposal is to determine the biochemical mechanisms responsible for the extensive epithelial cell injury occurring during the inflammatory phase of influenza viral infection. This knowledge will be exploited to develop methods to prevent or augment aspects of these mechanisms to minimize acute and longterm respiratory consequences associated with influenza infection. We propose experiments over the next five years to elucidate the biochemical and pathophysiological processes involved in influenza mediated airways injury. A multidisciplinary approach ranging from molecular biology to organ histology and physiology will address the interrelated specific aims: The first Specific Aim of this proposal will be to assess at the molecular and cellular levels the production and role of the xanthine: acceptor oxidoreductase system (XDH/XO) in the pathogenesis of influenza virus mediated epithelial injury and to correlate this with the morbidity and mortality associated with this virus. A hierarchy of methods ranging from in vitro, to in situ, to in vivo, will be employed to achieve this. The second Specific Aim of this proposal will be to investigate the regulation and role of iNOS generated NO generated reactive nitrogen species, will provide a further source of oxidant-mediated epithelial injury through the production of peroxynitrite. The third Specific Aim of this proposal will be to characterize the role of cytokine production in the modulation of XDH/XO and iNOS during influenza infection. Our preliminary results indicate that IFN-gamma, TNF, and Il-6 and Il-1 are increased over time in athe lungs of mice infected with influenza, corresponding to similar changes in XDH/XO and iNOS gene expression. With the use of molecular probes, we will examine the temporal relationship between cytokine appearance and XDH/XO (O2-) and iNOS (NO) activities, exploiting the use of monoclonal antibodies to focus on the role of particular cytokines.
