IgA increases NF-kB in CALU-3 cells*1Implications for cystic fibrosis airway inflammation

Abstract Rationale Cystic fibrosis (CF) airways are plagued by neutrophil-predominant inflammation and infection. CF epithelial cells have increased activity of the pro-inflammatory transcription factor, NF-κB, resulting in increased production of the neutrophil chemotactic factor, IL-8. IgA is an important protein that helps protect the airway epithelium from attachment by infectious pathogens, and is found in increased concentrations in CF serum and sputum. We asked whether IgA itself might contribute to the pro-inflammatory process by increasing NF-κB activity and therefore IL-8 production in CALU-3 airway epithelial cells that endogenously express both the CFTR (wild-type) and the receptor for IgA, the polymeric immunoglobulin receptor (pIgR). Methods CALU-3 cells were stimulated with buffer or pIgA, and processed for DNA binding activity of NF-κB (p65 subunit) by electromobility gel shift assay (EMSA). Increased NF-κB activity was also assessed by western blotting for the inhibitory molecule, IκB, which is degraded prior to translocation of NF-κB to the nucleus. The effect of IgA on IL-8 mRNA production was determined by RNAse protection assay. Results IgA increased (1) NF-κB DNA binding by EMSA, (2) IκB degradation on western blots, and (3) IL-8 mRNA production in CALU-3 cells compared to media alone. Conclusions IgA increases NF-κB activity in CALU-3 cells. Although IgA should contribute to protective immune defense, this protein may inadvertently exacerbate CF lung disease. Increased levels of IgA during infectious exacerbations may further increase NF-κB activity, IL-8 production and neutrophil recruitment resulting in further epithelial damage. However, the effect of IgA on cells with mutant CFTR needs to be determined.