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Activation of chloride transport in CF airway epithelial cell lines and primary CF nasal epithelial cells by S-nitrosoglutathione
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk cellbiologi.
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2006 (Engelska)Ingår i: Respiratory research (Online), ISSN 1465-9921, E-ISSN 1465-993X, Vol. 7, s. 124-Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Background: It has been suggested that low mu M concentrations of S-nitrosoglutathione (GSNO), an endogenous bronchodilator, may promote maturation of the defective cystic fibrosis (CF) transmembrane conductance regulator ( CFTR). Because nitric oxide ( NO) and GSNO levels appear to be low in the CF airway, there is an interest in the possibility that GSNO replacement could be of therapeutic benefit in CF.

Methods: The effect of GSNO on chloride (Cl-) transport was investigated in primary nasal epithelial cells obtained from CF patients homozygous for the delF508 mutation, as well as in two CF cell lines (CFBE and CFSME), using both a fluorescent Cl- indicator and X-ray microanalysis. Maturation of delF508 CFTR was determined by immunoblotting.

Results: Treatment with 60 mu M GSNO for 4 hours increased cAMP-induced chloride efflux in nasal epithelial cells from 18 out of 21 CF patients, but did not significantly affect Cl- efflux in cells from healthy controls. This Cl- efflux was confirmed by measurements with a fluorescent Cl- indicator in the CFBE and CFSME cell lines. The effect of GSNO on Cl- efflux in CFBE cells could be inhibited both by a specific thiazolidinone CFTR inhibitor (CFTRinh-172) and by 4,4'-diisothiocyanatodihydrostilbene- 2,2'-disulfonic acid (H2DIDS). X-ray microanalysis showed that, following 4 hours incubation with 60 mu M GSNO, cAMP agonists caused a decrease in the cellular Cl- concentration in CFBE cells, corresponding to Cl- efflux. GSNO exposure resulted in an increase in the protein expression and maturation, as shown by immunoblot analysis. GSNO did not increase the cytosolic Ca2+ concentration in cultured airway epithelial cells.

Conclusion: Previous studies have suggested that treatment with GSNO promotes maturation of delF508-CFTR, consistent with our results in this study. Here we show that GSNO increases chloride efflux, both in the two CF cell lines and in primary nasal epithelial cells from delF508-CF patients. This effect is at least in part mediated by CFTR. GSNO may be a candidate for pharmacological treatment of the defective chloride transport in CF epithelial cells.

Ort, förlag, år, upplaga, sidor
2006. Vol. 7, s. 124-
Nationell ämneskategori
Medicin och hälsovetenskap
Identifikatorer
URN: urn:nbn:se:uu:diva-24748DOI: 10.1186/1465-9921-7-124ISI: 000241188400001PubMedID: 17022806OAI: oai:DiVA.org:uu-24748DiVA, id: diva2:52522
Tillgänglig från: 2007-02-07 Skapad: 2007-02-07 Senast uppdaterad: 2017-12-07Bibliografiskt granskad
Ingår i avhandling
1. Functional Aspects of Epithelia in Cystic Fibrosis and Asthma
Öppna denna publikation i ny flik eller fönster >>Functional Aspects of Epithelia in Cystic Fibrosis and Asthma
2008 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

The cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP activated chloride channel in the apical membrane of epithelial cells, is defective in patients with cystic fibrosis (CF). Research efforts are focused on chloride channel function in order to find a cure for the disease.

Genistein increased chloride transport in normal and delF508-CFTR cultured airway epithelial cells without cAMP stimulation. Prior pretreatment with phenylbutyrate did not affect the rate of the genistein-stimulated chloride efflux in these cells.

S-nitrosoglutathione is an endogenous bronchodilator, present in decreased amounts in the lungs of CF patients. We studied the effect of GSNO on chloride (Cl-) transport in primary nasal epithelial cells from CF patients homozygous for the delF508-CFTR mutation, as well as in two CF cell lines, using a fluorescent Cl- indicator and X-ray microanalysis. GSNO increased chloride efflux in the CF cell lines and in primary nasal epithelial cells from CF patients. This effect was partly mediated by CFTR. If the cells were exposed to GSNO in the presence of L-cysteine, Cl- transport was enhanced after 5 min, but not after 4 h. GSNO may be a candidate for pharmacological treatment of CF patients.

Chloride transport properties of cultured NCL-SG3 sweat gland cells were investigated. The CFTR protein was neither functional nor expressed in these cells. Ca2+-activated chloride conductance was confirmed and the putative Ca2+-activated chloride channel (CaCC) was further characterized in term of its pharmacological sensitivity.

Corticosteroids, the primary treatment for asthma, cause necrosis/apoptosis of airway epithelial cells. It was investigated whether a newer generation of drugs used in asthma, leukotriene receptor antagonists, had similar effects. Both montelukast and dexamethasone, but not beclomethasone or budesonide induced apoptosis/necrosis in superficial airway epithelial cells. Montelukast and corticosteroids also caused decreased expression of intercellular adhesion molecule -1 (ICAM-1) in epithelial but not endothelial cells.

Ort, förlag, år, upplaga, sidor
Uppsala: Acta Universitatis Upsaliensis, 2008. s. 91
Serie
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 361
Nyckelord
Cell biology, cystic fibrosis, CFTR, chloride transport, airway epithelium, sweat gland, asthma, corticosteroids, montelukast, Cellbiologi
Identifikatorer
urn:nbn:se:uu:diva-8905 (URN)978-91-554-7224-5 (ISBN)
Disputation
2008-06-05, B7:113a, BMC, Husargatan 3, Uppsala, 13:15
Opponent
Handledare
Tillgänglig från: 2008-05-15 Skapad: 2008-05-15 Senast uppdaterad: 2013-09-20Bibliografiskt granskad

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