Few physiologically relevant model systems are available to investigate host-pathogen interactions in chronic obstructive pulmonary disease (COPD) at the site of the small airways. Non-typeable Haemophilus influenzae (NTHi) is an aetiological pathogen in 25% of COPD hospitalisations, with emerging biofilm properties.

This research aims to model host inflammatory responses and structural changes of a small airways epithelium (SABCi) grown at air-liquid-interface (ALI) in response to co-culture with two COPD lung-derived and biofilm-forming NTHi strains, ST14 and ST408, over 96-hours. We hypothesised that co-culture with NTHi would lead to biofilm formation and drive characteristic COPD phenotypes in the SABCi ALI model.

The SABCi ALI NTHi biofilm co-culture model demonstrated no evidence of de-differentiation, increased cytotoxicity, tight junction barrier degradation or increased permeability measured by TEER, LDH and FITC-Dextran assays (p>0.05), respectively.

ST14 elicited significant IL-6 and IL-8 responses (p<0.05) and TNF? responses (p<0.0001) by ELISA whilst ST408 elicited significant IL-8 and TNF? responses (p<0.05) demonstrating time- and strain-dependent host innate immune responses to NTHi biofilms.

Fluorescence in-situ hybridisation and fluorescence imaging with SYPRO Ruby and TOTO-1 stains identified apical aggregations of NTHi embedded within eDNA- and protein-rich aggregates consistent with biofilm EPS matrices. This study began to explore the immunohistological changes occurring including epithelial ciliation (?-tubulin) and mucus secretion (MUC5A/B).

NTHi biofilms may drive inflammation of COPD small airways disease and provide novel therapeutic targets.