CFTR is a negative regulator of NFkappaB mediated innate immune response.
BACKGROUND
Dysfunctional CFTR in the airways is associated with elevated levels of NFkappaB mediated IL-8 signaling leading to neutrophil chemotaxis and chronic lung inflammation in cystic fibrosis. The mechanism(s) by which CFTR mediates inflammatory signaling is under debate.


RESULTS
We tested the hypothesis that wt-CFTR down-regulates NFkappaB mediated IL-8 secretion. We transiently co-expressed wt-CFTR and IL-8 or NFkappaB promoters driving luciferase expression in HEK293 cells. Wt-CFTR expression in HEK293 cells suppresses both basal and IL1beta induced IL-8, and NFkappaB promoter activities as compared to the control cells transfected with empty vector (p<0.05). We also confirmed these results using CFBE41o- cells and observed that cells stably transduced with wt-CFTR secrete significantly lower amounts of IL-8 chemokine as compared to non-transfected control cells. To test the hypothesis that CFTR must be localized to cell surface lipid rafts in polarized airway epithelial cells in order to mediate the inflammatory response, we treated CFBE41o- cells that had been stably transduced with wt-CFTR with methyl-beta-cyclodextrin (CD). At baseline, CD significantly (p<0.05) induced IL-8 and NFkappaB reporter activities as compared to control cells suggesting a negative regulation of NFkappaB mediated IL-8 signaling by CFTR in cholesterol-rich lipid rafts. Untreated cells exposed to the CFTR channel blocker CFTR-172 inhibitor developed a similar increase in IL-8 and NFkappaB reporter activities suggesting that not only must CFTR be present on the cell surface but it must be functional. We verified these results in vivo by comparing survival, body weight and pro-inflammatory cytokine response to P. aeruginosa LPS in CFTR knock out (CFKO) mice as compared to wild type controls. There was a significant (p<0.05) decrease in survival and body weight, an elevation in IL-1beta in whole lung extract (p<0.01), as well as a significant increase in phosphorylated IkappaB, an inducer of NFkappaB mediated signaling in the CFKO mice.


CONCLUSIONS
Our data suggest that CFTR is a negative regulator of NFkappaB mediated innate immune response and its localization to lipid rafts is involved in control of inflammation.
Vij
Neeraj
N
Department of Pediatric Respiratory Sciences, The Johns Hopkins School of Medicine, Baltimore, Maryland, United States of America. nvij1@jhmi.edu
Mazur
Steven
S
Zeitlin
Pamela L
PL
D000818:Animals; D002460:Cell Line; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007113:Immunity, Innate / Q000502:physiology*; D016209:Interleukin-8 / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011401:Promoter Regions, Genetic
D016209:Interleukin-8; D016328:NF-kappa B; D019005:Cystic Fibrosis Transmembrane Conductance Regulator