214 Single-cell Functional Profiling of CFTR in Primary and Induced Pluripotent Stem Cell–derived Cystic Fibrosis-Relevant Epithelia

reversed by ivacaftor treatment.Additionally, ivacaftor treatment partially corrects airway inflammation and reduces concentrations of the airway gel-forming mucins Muc5b and Muc5ac in these rats.We used the hG551D rat model to understand how ivacaftor treatment alters the course of chronic lung infections with bacteria such as Pseudomonas aeruginosa.Methods: Six-month-old hG551D rats were treated with ivacaftor or methylcellulose vehicle before or after intratracheal inoculation with 10 6 CFUs of a P. aeruginosa mucoid clinical isolate PAM 57-15 in agarose-laden beads.Rats were euthanized 3 days after infection to study acute infection and 28 days after infection to study chronic infection.Lung tissue and bronchoalveolar lavage fluid (BALF) were collected at harvest.CFU enumeration from the lung homogenate was assessed to determine bacterial lung burden.Immune cells in the BALF were counted with differentiation.Mucins and cytokines were measured in the BALF using modified dot blot and enzyme-linked immunosorbent assay.Mucociliary transport was analyzed via micro-optical coherence tomography (OCT) imaging.Results: Rats treated with ivacaftor in the acute phase of infection had lower bacterial burden, a smaller percentage of neutrophils in the BALF, and less Muc5b and Muc5ac content in the BALF than the methylcellulose vehicle group.In this infection scheme, P. aeruginosa colonies recovered from the vehicle group were all mucoid, whereas 50% of the P. aeruginosa colonies recovered from rats treated with ivacaftor had reverted to a nonmucoid phenotype.In the second infection scheme, rats treated with ivacaftor before infection also had lower bacterial burden in the chronic phase than the vehicle group but no difference in neutrophil percentage or mucin content in the BALF.In the final scheme, hG551D rats that initiated modulator therapy after infection had established chronicity had no difference in bacterial burden, inflammation, or mucin from the vehicle group.Rats in the chronic infection experiment were also analyzed via μOCT, which found that mucociliary transport and mucus viscosity were no different in the rats treated with ivacaftor and the vehicle group.Conclusions: These data indicate that reductions in P. aeruginosa burden after initiation of highly effective modulator therapies such as ivacaftor depend on the phase of infection, with the most promising results acquired before the infection becomes chronic.These data also show that the beneficial effect of ivacaftor on mucociliary clearance is abrogated during chronic infection, suggesting that, to eradicate infection completely, additional therapeutic options will be needed.