The opportunistic human pathogen Pseudomonas aeruginosa (PA) causes persistent lung infections in susceptible individuals. People with pre-existing lung conditions such as cystic fibrosis or COPD are at high risk of developing chronic infection that is recalcitrant to antibiotic treatment and progressively destroys lung function. Despite ongoing extensive research, studies of host-pathogen interactions in PA are still limited by the lack of suitable long-term infection models.
In this work we set up a new system for modelling chronic infection with PA for extended periods of time using both normal bronchial epithelial (NHBE) cells as well as a lung epithelial cell line (Calu-3). We cultivated cells on transwell membranes at the air liquid interface to create a polarized cell layer and infected these with the laboratory strain PAO1 or well characterized clinical isolates from the BACTOME database that are adapted to the human host environment and exhibit decreased virulence.
After infection from the apical side, bacterial load, cell viability and pro-inflammatory cytokine secretion were determined between 6 hours and 5 days after infection. In contrast to cells infected with the cytotoxic control strain PAO1, clinical isolate-infected cells remained viable despite high bacterial loads (108 CFU/ml) and exhibited a functional immune response up to 5 days post-infection. Concurrently, H&E staining of infected Calu-3 cells revealed an intact polarized monolayer throughout infection.
We propose this 3D human cell model to be used for future investigations into host-pathogen interactions and compound screenings against PA. The described method is robust and reproducible, can be performed in a basic laboratory setting, and allows for detailed monitoring of cellular and bacterial parameters, such as viability, cytokine secretion and bacterial load.