Phe508del CFTR Cl− Secretion Modulation

CF-HBE and CFBE cells were treated with VX-809 (3 μM, Selleckchem, Houston, TX) for 48 h to increase Phe508del CFTR Cl^- secretion. VX-809 is a key component of Orkambi, an FDA drug approved for use in CF patients to stimulate Phe508del CFTR Cl^- secretion by CF-HBE cells. To provide biological validation for our proteomics analysis, in one set of experiments, the same number of OMVs isolated from control or Tobramycin treated P. aeruginosa were added to the apical side of CF-HBE or CFBE cells for 1.5 h before measurements of Phe508del CFTR Cl^- secretion. OMVs were quantified using FM 4–64 fluorescent dye (Invitrogen) as described previously [25 (link)]. In a second validation experiment either P. aeruginosa (PA14-wt) or PA14, in which the aprA gene was deleted (PA14-ΔaprA), was added to the apical side of CF-HBE cells at a multiplicity of infection (MOI) of 30:1 for 6 hours. Phe508del CFTR Cl^- secretion was measured as described in detail previously [9 (link)].