Introduction: The zinc finger protein A20 is responsible for the termination of NF-κB signalling through immune receptors including Toll-like receptors (TLRs). A20 deficient mice suffer from persistent activation of NF-κB by TLRs and tumour necrosis factor receptor, resulting in mortality among neonates and multiorgan inflammation in adults [1]. A20 has been shown to be a negative regulator of TLR2 and TLR4 induced inflammatory cytokine (IL-8) release in the airway epithelium [2] and, more recently, was found to attentuate allergic asthma in mice [3]. Cystic Fibrosis (CF) is characterised by chronically inflammed lung disease, which is the primary cause of mortality amoung sufferers. We hypothesise that in the CF epithelium, defective A20 action leads to enhanced pro-inflammatory signalling through the NF-κB pathway. Methods: Non-CF (HBE) and CF (CFBE) bronchial epithelial cell lines were grown to 80% confluency on collagen-coated plates prior to stimulation with 50µg/ml PA-lipopolysaccharide (LPS; Sigma). A20 and NF-κB (P50 and P65) levels were measured by flow cytometry, 0-12h after stimulation. Nuclear extracts for NF-κB staining were obtained using a DNA CycleTest kit (BD Biosciences). IL-8 release was measured by ELISA (Peprotech) using supernatants from cells treated with PA-LPS for 24h. Statistics were calculated by ANOVA, with n=3-6 for all experiments. Results: IL-8 release was significantly higher in CFBE cells (p<0.05). Despite increased intracellular A20 expression 4h (P<0.001) after PA-LPS exposure, expression in CFBE cells fell below basal levels by 12h exposure, while HBE cells showed sustained A20 expression. Basal NF-κB (both P50 and P65) expression in CFBE cells was significantly greater than in HBE cells (P<0.0001). P50 levels remained unchanged in HBE cells following PA-LPS stimulation, while CFBE cells showed significant increases (2 and 4h after stimulation, P<0.001). P65 expression increased significantly in both HBE and CFBE cells with exposure to PA-LPS (P<0.05-0.001). However, these increases were more pronounced in CFBE cells. Notably, P65 expression remained significantly up-regulated at 12h (when A20 expression was inhibited) in CFBE cells and not in HBE cells, while P50 expression returned to basal levels in both cell types. Conclusions: In CF epithelium, A20 signalling is significantly inhibited causing increased NF-κB activation, contributing to a state of chronic inflammation.