WS08.04Analyzing the impact of elexacaftor/tezacaftor/ivacaftor treatment and Pseudomonas infection on gene expression in cystic fibrosis primary bronchial epithelial cultures

Abstract

Cystic Fibrosis (CF) is caused by mutations in the CFTR gene which encodes a chloride and bicarbonate ion channel. Ablation or impairment of CFTR function affects multiple organs however, the major pathological consequence affecting patient health is recurring bacterial lung infections, resulting in respiratory failure. P. aeruginosa is one of the predominant bacteria which causes chronic infection in 60-80% CF adults. Although highly effective CFTR modulator therapies are now available, they do not fully eliminate lung infections and their impact on bacterial infections in airway cell culture is not fully understood.

Objective: Using our recently established model of diseased airway epithelium, we analyzed the effect of bacterial infection and CFTR modulators on gene expression in primary human bronchial epithelial (HBE) cultures.

Methods: Fully differentiated HBE cultures obtained from 6 homozygous F508del donors were infected with P. aeruginosa strain PA01. Tobramycin was added 12 h post infection and cultures were treated with CFTR modulators VX-445, VX-661 and VX-770 for 48 h. RNA isolated from HBE ± bacterial infection and ± ETI was used for bulk RNAseq.

Results: We successfully established a 72 h bacteria-epithelia co-culture model in airway cells to mimic persistent bacterial infection. In Pseudomonas infected HBE, RNAseq demonstrated significant changes in expression of genes involved in key biological pathways such as innate immune response, inflammation, hypoxia, cytoskeleton organization. Interestingly, HBE treated with ETI without any bacterial infection also exhibited more than 400 differentially expressed genes. Consistent with our previous report, we observed significant reduction in bacterial load in ETI-treated HBE cells.

Conclusion: Our study underscores the importance of incorporating live bacteria to faithfully replicate the CF lung for investigating how bacterial infections and CFTR modulators impact gene expression in airway epithelia.