Viral respiratory infections are the major cause of acute exacerbations, and the virus most often implicated is rhinovirus (RV). RV induced production of interleukin-8 (IL-8) and RANTES correlates closely with cellular inflammation, the severity of common cold symptoms, and changes in airway hyperresponsiveness. Together, these suggest that dysregulation of these chemokines in the lower airway, through effects on neutrophilic and lymphocytic inflammation, is a major factor in causing airway obstruction and asthma. Preliminary experiments conducted to establish mechanisms by which RV replication induces the production of chemokines indicate that RV double-stranded RNA (dsRNA) and activation of the double-stranded RNA-dependent protein kinase (PKR) are key events to increase epithelial cell generation of IL- 8 and RANTES. Based upon these findings, we hypothesize that dsRNA activation of PKR and RV 3C protease stimulate specific signaling cascades [nuclear factor kappa-B (NF-kappaB); mitogen- activated protein kinases (MAPK), CCAAT/enhancer-binding protein beta (C/EBPbeta)] that promote transcription of IL-8 and RANTES by epithelial cells. Furthermore, there is a loss in the regulation of this process in asthma, thus leading to dysregulated generation of these chemokines, and increased airway inflammation and asthma. To test these hypotheses, we propose to determine the effects of dsRNA on RANTES and IL-8 gene transcription, and selected transcription factors which regulate this process, in non-transformed bronchial epithelial (BE) cells. In addition, the effects of 3C protease on the cleavage of the repressor protein Oct-1 and upregulation of IL-8 transcription will be determined. Finally, we propose to evaluate differences in chemokine generation and regulation in BE cells derived from normal and asthmatic volunteers. Together, these studies will link specific events and proteins in the RV replication cycle to the generation of pro- inflammatory chemokines, and determine effects of asthma on these processes. By achieving these goals, the proposed studies will yield additional information about the pathogenesis of virus-induced asthma exacerbations in children, and thereby identify new targets for genetic analyses and more effective therapeutic strategies.
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