The inflammatory pathogenesis in airways of patients with cystic fibrosis (CF) is still unresolved. We demonstrate here that in in situ human DeltaF508 homozygous CF bronchial tissues, submucosal gland cells exhibit an absence of inhibitor factor kappaBalpha (IkappaBalpha) and high levels of chemokine interleukin-8 (IL-8) expression. These results were confirmed by cultured human CF bronchial gland cells in which a lack of cytosolic IkappaBalpha and high levels of constitutively activated nuclear factor kappaB (NFkappaB) associated with an up-regulation of IL-8 production (13-fold increase) were found when compared to non-CF (control) disease bronchial gland cells. We also demonstrated that the isoflavone genistein, a well known CFTR mutant Cl(-) channel stimulator, significantly reduces the endogenous and Pseudomonas aeruginosa lipopolysaccharide-induced IL-8 production in cultured CF bronchial gland cells by increasing cytosolic IkappaBalpha protein levels. Overall, results show that genistein is a potent inhibitor of the activated NFkappaB identified in CF gland cells. This strong inhibition of constitutively activated NFkappaB and the resulting down-regulation of IL-8 production by genistein in the CF gland cells highlights the key role played by cytosolic IkappaBalpha in the regulation of inflammatory processes in CF human airway cells.

Increasing evidence suggests that in airways from cystic fibrosis (CF) patients, inflammation may precede bacterial infection and be related to an endogenous dysregulation of proinflammatory cytokines in airway epithelial cells. Several investigators have reported that, in CF airway fluids, elevated NaCl concentrations may also contribute to the diseased state by inhibiting the bactericidal properties of airway fluid. Because many proinflammatory cytokines are transcriptionally regulated by the NF-κB, we investigated whether an elevated extracellular NaCl content in airway fluids significantly impaired the regulation of the NF-κB/IκBα complex and the chemokine IL-8 production in primary non-CF and CF human bronchial gland epithelial cells. Exposure of non-CF gland cells to hypotonic (85 mM) NaCl solution, compared with isotonic (115 mM) NaCl and hypertonic (170 mM) NaCl solutions, resulted in a significant decrease in IL-8 production that was paralleled by a strong inhibition of activated NF-κB associated with an increased cytosolic expression of IκBα and a decrease in the IκB kinase α protein level. In CF gland cells, we demonstrated that, compared with the high IL-8 in an hypertonic solution, the release of IL-8 was significantly reduced 2-fold in an isotonic solution and 5-fold in a hypotonic solution. Strikingly, exposure of CF bronchial gland cells to either hypotonic or isotonic milieu did not result in a marked inhibition of the activated NF-κB/IκBα system. This is the first demonstration that primary human CF bronchial gland cells exhibit abnormally high IL-8 production through constitutively activated NF-κB and high IκB kinase α level, whatever the hypo-, iso-, and hypertonic NaCl milieu.

In many airway diseases, the airway epithelium is severely damaged and has to regenerate rapidly to restore its function. The regeneration process involves chronological steps of epithelial cell migration, proliferation, stratification, and differentiation. The present study has used an in vivo humanized airway xenograft model in nude mice that mimics the regeneration dynamics of human airway epithelium after severe injury, and human-specific molecular tools, to study the expression profiles of epithelial matrix metalloproteinases (MMPs)-7 and -9, of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1), and of the pro-inflammatory cytokine interleukin-8 (IL-8) during the different steps of human airway epithelium regeneration. It was found that during the cell migration and proliferation steps, airway epithelial cells expressed IL-8 at a high level, whereas airway epithelial pseudo-stratification and surface airway epithelial differentiation were associated with increased expression of MMPs and a progressive decrease in IL-8. Interestingly, immunohistochemical analysis revealed exclusive expression of MMPs at the apical part of the well-differentiated regenerated airway epithelium, and incubation of the regenerating epithelial cells with MMP inhibitors led to abnormal epithelial differentiation. These data provide new insight into the temporal expression of MMPs and IL-8 during the regeneration of airway epithelium and demonstrate the involvement of these factors during the different steps that lead to restoration of a well-differentiated and functional airway epithelium.

In response to bacterial infection, airway epithelium releases inflammatory mediators including cytokines and chemokines that lead to immune cell efflux and could stimulate the adaptive T cell immune response. The aim of our study was to analyze, in a double chamber culture, the chemokine changes in response to Staphylococcus aureus and their consequences for T cells. Our data show that S. aureus stimulates basolateral and apical release of IL-8 and eotaxin by airway epithelial cells. We also observed increased chemokine receptor expression on CD8+ and CD4+ T cells and enhanced chemotaxis of CD4+ T cells toward apical supernatant. Our data strongly suggest that S. aureus interaction with airway epithelium contributes to specific migration of T cells to inflamed sites.

Several recent reports have suggested that airway inflammation may precede infection and relate to an endogenous dysregulation of pro-inflammatory cytokines in cystic fibrosis (CF) airways. Evidence suggests that activation of the nuclear factor kappa B (NFkappaB), which regulates the inflammatory gene transcription, depends on the degradation of the inhibitory factor IkappaBalpha. We show that, in in situ human DeltaF508 CF bronchial tissues, inhibitor factor IkappaBalpha is not present in gland cells, although endogenous levels of chemokine IL-8 are high. These data are confirmed by studying cultured CF human bronchial gland cells, in which a lack of cytosolic IkappaBalpha and high levels of activated NFkappaB, concomitant with IL-8 overproduction (a 13-fold increase) are found when compared to non-CF bronchial gland cells. Interestingly, treatment of CF gland cells with the isoflavone genistein, a well known CFTR mutant Cl(-) channel stimulator, results in a significant decrease ( P< 0.001) in IL-8 production down to levels released by non-CF gland cells. The addition of genistein also reverses the effects of lipopolysaccharide (LPS) Pseudomonas-aeruginosa-induced nuclear translocation of NFkappaB by increasing IkappaBalpha protein level (65%) in CF gland cells. Our data indicate that the induction of IkappaBalpha protein in CF airway glandular epithelial cells may be a novel mechanism by which IL-8-mediated lung inflammatory events are markedly reduced in CF patients, at least at the airway glandular level.