Reduced neutrophil elastase inhibitor elafin and elevated TGFβ1 are linked to inflammatory response in sputum of CF patients with Pseudomonas aeruginosa

Research questionPulmonary disease progression in patients with cystic fibrosis (CF) is characterized by inflammation and fibrosis, and aggravated by Pseudomonas aeruginosa (Pa). We investigated the impact of Pa specifically 1) on protease/antiprotease balance and 2) inflammation, as well as 3) the link of both parameters to clinical parameters of CF-patients.MethodsTGFβ1, IL1β, IL8, neutrophil elastase (NE) and elastase inhibitor elafin were measured (ELISA assays), and gene expression of the NF-ĸB pathway was assessed (RT-PCR) in the sputum of 60 CF-patients with a minimum age of 5 years. Spirometry was assessed according to ATS guidelines.Results1) NE was markedly increased in Pa-positive sputum, whereas elafin was significantly decreased. 2) Increased IL1β/IL8 were associated with both Pa infection and reduced FEV1, as well as sputum TGFβ1 was elevated in Pa-infected CF-patients and linked to an impaired lung function. 3) Moreover, gene expression of NF-ĸB signaling components was increased in sputum of Pa-infected patients; these findings were positively correlated with IL8.ConclusionOur study links Pa infection to an imbalance of NE and NE-inhibitor elafin and increased inflammatory mediators. Moreover, our data demonstrate an association between high TGFβ1 sputum levels and a progress in chronic lung inflammation and pulmonary fibrosis in CF. Controlling the excessive airway inflammation by inhibition of NE and TGFβ1 might be promising therapeutic strategies in future CF therapy and a possible complement to CFTR-modulators.

Treatment with a neutrophil elastase inhibitor and ofloxacin reduces P. aeruginosa burden in a mouse model of chronic suppurative otitis media

Chronic suppurative otitis media (CSOM) is a widespread, debilitating problem with poorly understood immunology. Here, we assess the host response to middle ear infection over the course of a month post-infection in a mouse model of CSOM and in human subjects with the disease. Using multiparameter flow cytometry and a binomial generalized linear machine learning model, we identified Ly6G, a surface marker of mature neutrophils, as the most informative factor of host response driving disease in the CSOM mouse model. Consistent with this, neutrophils were the most abundant cell type in infected mice and Ly6G expression tracked with the course of infection. Moreover, neutrophil-specific immunomodulatory treatment using the neutrophil elastase inhibitor GW 311616A significantly reduces bacterial burden relative to ofloxacin-only treated animals in this model. The levels of dsDNA in middle ear effusion samples are elevated in both humans and mice with CSOM and decreased during treatment, suggesting that dsDNA may serve as a molecular biomarker of treatment response. Together these data strongly implicate neutrophils in the ineffective immune response to P. aeruginosa infection in CSOM and suggest that immunomodulatory strategies may benefit drug-tolerant infections for chronic biofilm-mediated disease.