title: CFTR regulation of intracellular pH and ceramides is required for lung endothelial cell apoptosis.
abstract: The functional significance of the expression of cystic fibrosis transmembrane regulator (CFTR) on endothelial cells has not yet been elucidated. Since CFTR has been implicated in the regulation of intracellular sphingolipid levels, which are important regulators of endothelial cell apoptosis in response to various insults, we investigated the role of CFTR in the apoptotic responses of lung endothelial cells. CFTR was detected as a functional chloride channel in primary lung endothelial cells isolated from both pulmonary arteries (human or mouse) and bronchial arteries (sheep). Both specific CFTR inhibition with 2-(phenylamino) benzoic acid diphenylamine-2-carboxylic acid, 5-[(4-carboxyphenyl)methylene]-2-thioxo-3-[(3-trifluoromethyl)phenyl-4-thiazolidinone (CFTR(inh)-172), or 5-nitro-2-(3-phenylpropylamino)benzoic acid and CFTR knockdown significantly attenuated endothelial cell apoptosis induced by staurosporine or H(2)O(2). CFTR(inh)-172 treatment prevented the increases in the ceramide:sphingosine-1 phosphate ratio induced by H(2)O(2) in lung endothelial cells. Replenishing endogenous ceramides via sphingomyelinase supplementation restored the susceptibility of CFTR-inhibited lung endothelial cells to H(2)O(2)-induced apoptosis. Similarly, the anti-apoptotic phenotype of CFTR-inhibited cells was reversed by lowering the intracellular pH, and was reproduced by alkalinization before H(2)O(2) challenge. TUNEL staining and active caspase-3 immunohistochemistry indicated that cellular apoptosis was decreased in lung explants from patients with cystic fibrosis compared with those with smoking-induced chronic obstructive lung disease, especially in the alveolar tissue and vascular endothelium. In conclusion, CFTR function is required for stress-induced apoptosis in lung endothelial cells by maintaining adequate intracellular acidification and ceramide activation. These results may have implications in the pathogenesis of cystic fibrosis, where aberrant endothelial cell death may dysregulate lung vascular homeostasis, contributing to abnormal angiogenesis and chronic inflammation.
authors.affiliation: Section of Pulmonology and Critical Care, Department of Pediatrics, Indiana University, Indianapolis, Indiana 46202-5120, USA.
authors.forename: Julie
authors.initials: J
authors.lastname: Noe
authors.forename: Daniela
authors.initials: D
authors.lastname: Petrusca
authors.forename: Natalia
authors.initials: N
authors.lastname: Rush
authors.forename: Ping
authors.initials: P
authors.lastname: Deng
authors.forename: Mary
authors.initials: M
authors.lastname: VanDemark
authors.forename: Evgeny
authors.initials: E
authors.lastname: Berdyshev
authors.forename: Yuan
authors.initials: Y
authors.lastname: Gu
authors.forename: Patricia
authors.initials: P
authors.lastname: Smith
authors.forename: Kelly
authors.initials: K
authors.lastname: Schweitzer
authors.forename: Joseph
authors.initials: J
authors.lastname: Pilewsky
authors.forename: Viswanathan
authors.initials: V
authors.lastname: Natarajan
authors.forename: Zao
authors.initials: Z
authors.lastname: Xu
authors.forename: Alexander G
authors.initials: AG
authors.lastname: Obukhov
authors.forename: Irina
authors.initials: I
authors.lastname: Petrache
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002518:Ceramides / Q000378:metabolism*; D003550:Cystic Fibrosis / Q000378:metabolism / Q000473:pathology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008168:Lung / Q000166:cytology / Q000378:metabolism; D051379:Mice; D016877:Oxidants / Q000378:metabolism; D018408:Patch-Clamp Techniques; D013107:Sphingolipids / Q000378:metabolism
chemical_list: C505032:CFTR protein, human; D002518:Ceramides; D016877:Oxidants; D013107:Sphingolipids; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D006861:Hydrogen Peroxide