期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 284, 期 12, 页码 7681-7686出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M809161200
关键词
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资金
- National Institutes of Health [EB00415, DK35124, HL73856, HL59198, EY13574, DK72517, DK81355]
- Cystic Fibrosis Foundation (Research Development Program and Drug Discovery grants)
- American Heart Association [0765070Y]
The mechanisms remain uncertain by which mutations in CFTR cause lung disease in cystic fibrosis (CF). Teichgraber et al. recently reported increased ceramide in CF lungs, which was proposed to result from defective lysosomal acidification in airway epithelial cells and consequent impairment of pH-dependent ceramide-metabolizing enzymes (Teichgraber, V., Ulrich, M., Endlich, N., Reithmuller, J., Wilker, B., Conceicao Ce Olivereira-Munding, C., van Heeckeren, A. M., Barr, M. L., von Kurthy, G., Schmid, K. W., Weller, M., Tummler, B., Lang, F., Grassme, H., Doring, G., and Gulbins, E. (2008) Nat. Med. 14, 382-391). Here, we measured lysosomal pH in several CFTR-expressing and -deficient cell lines, freshly isolated airway epithelial cells from non-CF and CF mice and humans, and well-differentiated primary cultures of human non-CF and CF airway epithelial cells. Lysosomal pH was measured by ratio imaging using a fluorescent pH indicator consisting of 40-kDa dextran conjugated to Oregon Green 488 and tetramethylrhodamine. In all cell types, lysosomal pH was similar to 4.5, unaffected by the thiazolidinone CFTR inhibitor CFTRinh-172, and increased to similar to 6.5 following bafilomycin inhibition of the vacuolar proton pump. Lysosomal pH did not differ significantly in airway epithelial cells from non-CF versus CF humans or mice. Our results provide direct evidence against the conclusions of Teichgraber et al. that lysosomal acidification is CFTR-dependent, impaired in CF, or responsible for ceramide accumulation. As such, alternative mechanisms are needed to explain increased ceramide in CF airways. Non-CFTR mechanisms, such as ClC-type chloride channels, are likely involved in maintaining electroneutrality during organellar acidification.
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