title: Cystic fibrosis transmembrane conductance regulator-independent phagosomal acidification in macrophages.
abstract: It was reported recently that the cystic fibrosis transmembrane conductance regulator (CFTR) is required for acidification of phagosomes in alveolar macrophages (Di, A., Brown, M. E., Deriy, L. V., Li, C., Szeto, F. L., Chen, Y., Huang, P., Tong, J., Naren, A. P., Bindokas, V., Palfrey, H. C., and Nelson, D. J. (2006) Nat. Cell Biol. 8, 933-944). Here we determined whether the CFTR chloride channel is a generalized pathway for chloride entry into phagosomes in macrophages and whether mutations in CFTR could contribute to alveolar macrophage dysfunction. The pH of mature phagolysosomes in macrophages was measured by fluorescence ratio imaging using a zymosan conjugate containing Oregon Green(R) 488 and tetramethylrhodamine. Acidification of phagolysosomes in J774A.1 macrophages (pH approximately 5.1 at 45 min), murine alveolar macrophages (pH approximately 5.3), and human alveolar macrophages (pH approximately 5.3) was insensitive to CFTR inhibition by the thiazolidinone CFTR(inh)-172. Acidification of phagolysosomes in alveolar macrophages isolated from mice homozygous for DeltaF508-CFTR, the most common mutation in cystic fibrosis, was not different compared with that in alveolar macrophages isolated from wild-type mice. We also measured the kinetics of phagosomal acidification in J774A.1 and murine alveolar macrophages using a zymosan conjugate containing fluorescein and tetramethylrhodamine. Phagosomal acidification began within 3 min of zymosan binding and was complete within approximately 15 min of internalization. The rate of phagosomal acidification in J774A.1 cells was not slowed by CFTR(inh)-172 and was not different in alveolar macrophages from wild-type versus DeltaF508-CFTR mice. Our data indicate that phagolysosomal acidification in macrophages is not dependent on CFTR channel activity and do not support a proposed mechanism for cystic fibrosis lung disease involving defective phagosomal acidification and bacterial killing in alveolar macrophages.
authors.affiliation: Department of Medicine, University of California, San Francisco, California 94143-0521, USA.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Haggie
authors.forename: A S
authors.initials: AS
authors.lastname: Verkman
mesh_terms: D000818:Animals; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology; D002264:Carboxylic Acids / Q000138:chemical synthesis / Q000378:metabolism; D002460:Cell Line; D018118:Chloride Channels / Q000502:physiology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000502:physiology*; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000378:metabolism; D006575:Heterocyclic Compounds, 3-Ring / Q000737:chemistry; D006720:Homozygote; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008247:Lysosomes / Q000502:physiology; D008264:Macrophages / Q000502:physiology*; D016676:Macrophages, Alveolar / Q000378:metabolism / Q000502:physiology; D017737:Macrophages, Peritoneal / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D008815:Mice, Inbred Strains; D008856:Microscopy, Fluorescence; D009154:Mutation; D010588:Phagosomes / Q000502:physiology*; D012235:Rhodamines; D013997:Time Factors; D015054:Zymosan / Q000737:chemistry
chemical_list: D002264:Carboxylic Acids; D018118:Chloride Channels; D005456:Fluorescent Dyes; D006575:Heterocyclic Compounds, 3-Ring; C436864:Oregon Green 488 carboxylic acid; D012235:Rhodamines; C412715:tetramethylrosamine; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D015054:Zymosan