Biofilm-Induced Type 2 Innate Immunity in a Cystic Fibrosis Model of Pseudomonas aeruginosa.

Kenny Bielen,Tom K Raju,Samir Kumar-Singh,Jan Boddaert,Philippe G Jorens,Bart ‘S Jongers,Christine Lammens,Herman Goossens,Surbhi Malhotra-Kumar

doi:10.3389/fcimb.2017.00274

Kenny Bielen, Tom K Raju + Show 7 more

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https://doi.org/10.3389/fcimb.2017.00274

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Biofilm-producing strains of Pseudomonas aeruginosa are a major cause of morbidity and mortality in cystic fibrosis (CF) patients. In these patients, increased levels of IL-17 as well as of IL-5 and IL-13 along with arginase (Arg)-positive macrophages have been observed in bronchoalveolar lavage fluid. While IL-17 is a strong proinflammatory cytokine associated with host defense against bacterial and fungal infections and is also elevated in several autoimmune diseases, IL-5/IL-13 and Arg1-positive M2 macrophages are part of the anti-inflammatory type 2 (Th2) immunity. To study whether increased IL-5 and IL-13 levels are related to biofilm formation, which is frequently observed in CF patients colonized by P. aeruginosa, we utilized an agarose bead-embedded P. aeruginosa rat model commonly employed in in vivo biofilm studies. We showed that “sterile” agarose bead instillation in rat notably increased lung transcript levels of IL-5 and IL-13 at two post-instillation study-points, day 1 and day 3. Concurrently, increased infiltration of type 2 innate cells such as eosinophils and Arg1 positive M2 activated macrophages (Arg1+CD68+) was also observed both at day 1 and day 3 while the proportion of M1 activated macrophages (iNOS+CD68+) at these time-points decreased. In contrast, P. aeruginosa-loaded beads caused a drastic elevation of proinflammatory Th1 (IFNγ, TNFα, IL-12a) and antibacterial Th17 (IL-17a, IL-17f, IL-22, IL-23a) cytokines along with a high influx of neutrophils and M1 macrophages, while Th2 cytokines (IL-5 and IL-13) drastically declined at day 1 post-infection. Interestingly, at day 3 post-infection, both Th1 and Th17 cytokines sharply declined and corroborated with decreased M1 and increased M2 macrophages. These data suggest that while IL-17 is linked to episodes of acute exacerbations of infection in CF patients, the increased Th2 cytokines and M2 macrophages observed in these patients are largely due to the biofilm matrix. The data presented here has important implications for clinical management of CF patients.

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Pseudomonas aeruginosa is the major cause of pulmonary infection in cystic fibrosis (CF) patients (Gibson et al, 2003)
Analyzing the lung transcript levels for the Th17 cytokine family, we showed increased expression of IL-17a (157-fold, P < 0.001) and IL-22 (112-fold, P < 0.01) at d1 post-bead instillation that were sustained at the d3 time-point (IL-17a, 158-fold, P < 0.001; IL-22, 65-fold, P < 0.01; Figure 1B)
Lung transcript levels of Th2 cytokines in sterile agar beads-instilled animals were markedly elevated with increased expression of IL-5 and IL-13 at d1 post-bead instillation (IL-5, 39-fold, P < 0.01; IL-13, 102-fold, P < 0.001) that remained elevated at d3 (IL-5, 30-fold, P < 0.05; IL-13, 26fold, P < 0.001)

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Pseudomonas aeruginosa is the major cause of pulmonary infection in cystic fibrosis (CF) patients (Gibson et al, 2003). Chronic P. aeruginosa colonization is associated with increased morbidity and mortality in CF patients especially during episodes of acute exacerbations (Bhatt, 2013). This heightened vulnerability and persistence of bacterial colonization is caused by increased mucous secretion in the alveolar spaces that provide an ideal environment to form biofilms (Sadikot et al, 2005; Moreau-Marquis et al, 2008). Suppressive therapy reduces P. aeruginosa lung burden, but does not impact lung biofilm burden (Fernandez-Barat et al, 2016) To mimic this distinct pathology observed in CF and other chronic obstructive lung diseases such as bronchiectasis, agarose beads are employed to establish a chronic lung infection model. The bioelectric characteristics in the tracheal airways of mice and humans diverge significantly and CFTR genetic models continue to demonstrate substantial cAMP-inducible changes in chloride permeability despite the absence of a functioning CFTR (Grubb et al, 1994; Liu et al, 2006)

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