Background: Aberrant epithelial repair is a key event in the airway remodelling which characterises obliterative bronchiolitis (OB) in the transplanted lung. The potential for airway epithelium from lung transplant recipients to undergo epithelial to mesenchymal cell transition (EMT) was assessed in culture and in vivo in lung allograft tissue. Methods: Change in epithelial and mesenchymal marker expression was assessed after stimulation with transforming growth factor b 1 (TGF-b 1 ) alone or in combination with tumour necrosis factor a (TNFa) and compared with untreated controls. The ability of cells to deposit extracellular matrix, secrete matrix metalloproteinases (MMPs) and invade collagen was investigated. Immunolocalisation of epithelial and mesenchymal markers was compared in airway tissue from stable recipients and those with OB. Results: Untreated cells maintained epithelial morphology and phenotype. TGF-b 1 reduced expression of epithelial markers, increased expression of vimentin and fibronectin, promoted collagen I and fibronectin deposition and increased MMP-9 production. Co-treatment with TNFa dramatically accentuated phenotypic and some functional features of EMT. Airway epithelial biopsies from recipients with OB demonstrated significantly increased staining for mesenchymal markers and significantly reduced Ecadherin staining compared with stable recipients.Conclusions: These observations demonstrate the ability of human airway epithelium to undergo EMT and suggest this phenomenon may be a potential link between inflammatory injury and TGF-b 1 -driven airway remodelling in the development of OB.

Background: Obliterative bronchiolitis in chronic rejection of lung allografts is characterised by airway epithelial damage and fibrosis. The process whereby normal epithelium is lost and replaced by fibroblastic scar tissue is poorly understood, but recent findings suggest that epithelial cells can become fibroblasts through epithelial-mesenchymal transition (EMT). It is hypothesised that EMT occurs in lung allografts and plays a potential role in airway remodelling. Methods: Sixteen stable lung transplant recipients underwent bronchoscopy with bronchoalveolar lavage (BAL), endobronchial biopsies, and bronchial brushings. Biopsy sections were stained for the fibroblast marker S100A4. Brushings were cultured on collagen, stained with anti-S100A4, and examined for further EMT markers including matrix metalloproteinase (MMP) zymographic activity and epithelial invasion through collagen coated filters. Results: A median 15% (0-48%) of the biopsy epithelium stained for S100A4 in stable lung transplant recipients and MMP-7 co-localisation was observed. In non-stimulated epithelial cultures from lung allografts, S100A4 staining was identified with MMP-2 and MMP-9 production and zymographic activity. MMP total protein and activity was increased following stimulation with transforming growth factor (TGF)-b 1 . Non-stimulated transplant epithelial cells were invasive and penetration of collagen coated filters increased following TGF-b 1 stimulation. Conclusions: This study provides evidence of EMT markers in lung allografts of patients without loss of lung function. The EMT process may represent a final common pathway following injury in more common diseases characterised by airway remodelling.

We have shown that elevated levels of pepsin, a biomarker of gastric aspiration, are consistently identified in the BAL of lung allografts. The highest levels were seen in patients with > or = grade A2 acute rejection. This provides further evidence supporting the possible role of aspiration in the development of overall allograft injury.

BackgroundWe conducted a placebo-controlled trial of azithromycin therapy in bronchiolitis obliterans syndrome (BOS) post lung transplantation.MethodsWe compared azithromycin (250 mg alternate days, 12 weeks) with placebo. Primary outcome was FEV1 change at 12 weeks.Results48 patients were randomised; (25 azithromycin, 23 placebo). It was established, post randomisation that two did not have BOS. 46 patients were analysed as intention to treat (ITT) with 33 ‘Completers’. ITT analysis included placebo patients treated with open-label azithromycin after study withdrawal.OutcomeThe ITT analysis (n=46, 177 observations) estimated mean difference in FEV1 between treatments (azithromycin minus placebo) was 0.035 L, with a 95% CI of −0.112 L to 0.182 L (p=0.6). Five withdrawals, who were identified at the end of the study as having been randomised to placebo (four with rapid loss in FEV1, one withdrawn consent) had received rescue open-label azithromycin, with improvement in subsequent FEV1 at 12 weeks. Study Completers showed an estimated mean difference in FEV1 between treatment groups (azithromycin minus placebo) of 0.278 L, with 95% CI for the mean difference: 0.170 L to 0.386 L (p=<0.001). Nine of 23 ITT patients in the azithromycin group had ≥10% gain in FEV1 from baseline. No patients in the placebo group had ≥10% gain in FEV1 from baseline while on placebo (p=0.002). Seven serious adverse events, three azithromycin, four in the placebo group, were deemed unrelated to study medication.ConclusionsAzithromycin therapy improves FEV1 in patients with BOS and appears superior to placebo. This study strengthens evidence for clinical practice of initiating azithromycin therapy in BOS.Trial registration numberEU-CTR, 2006-000485-36/GB.

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