Terminal sialylation is altered in airway cells with impaired CFTR-mediated chloride transport

Case Western Reserve University, Cleveland, Ohio, United States
AJP Lung Cellular and Molecular Physiology (Impact Factor: 4.08). 04/2001; 280(3):L482-92.
Source: PubMed


Reduced terminal sialylation at the surface of airway epithelial cells from patients with cystic fibrosis may predispose them to bacterial infection. To determine whether a lack of chloride transport or misprocessing of mutant cystic fibrosis transmembrane conductance regulator (CFTR) is critical for the alterations in glycosylation, we studied a normal human tracheal epithelial cell line (9/HTEo(-)) transfected with the regulatory (R) domain of CFTR, which blocks CFTR-mediated chloride transport; DeltaF508 CFTR, which is misprocessed, wild-type CFTR; or empty vector. Reduced cAMP-stimulated chloride transport is seen in the R domain and DeltaF508 transfectants. These two cell lines had consistent, significantly reduced binding of elderberry bark lectin, which recognizes terminal sialic acid in the alpha-2,6 configuration. Binding of other lectins, including Maakia amurensis lectin, which recognizes sialic acid in the alpha-2,3 configuration, was comparable in all cell lines. Because the cell surface change occurred in R domain-transfected cells, which continue to express wild-type CFTR, it cannot be related entirely to misprocessed or overexpressed CFTR. It is associated most closely with reduced CFTR activity.

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    • "Our prior studies, in two separate cell model systems, have shown that there is increase in available asialoGM1 (aGM1), which binds to P. aeruginosa pilin and flagellin and serves as a major ligand for this organism, on the CF member of the cell pair [17-19]. In these same cell pairs, there is an increased response of IL-8, IL-6, and granulocyte macrophage colony stimulating factor (GM-CSF) to a laboratory strain of P. aeruginosa, PAO1, in the CF member of the pair [6]. "
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    ABSTRACT: In many model systems, cystic fibrosis (CF) phenotype airway epithelial cells in culture respond to P. aeruginosa with greater interleukin (IL)-8 and IL-6 secretion than matched controls. In order to test whether this excess inflammatory response results from the reported increased adherence of P. aeruginosa to the CF cells, we compared the inflammatory response of matched pairs of CF and non CF airway epithelial cell lines to the binding of GFP-PAO1, a strain of pseudomonas labeled with green fluorescent protein. There was no clear relation between GFP-PAO1 binding and cytokine production in response to PAO1. Treatment with exogenous aGM1 resulted in greater GFP-PAO1 binding to the normal phenotype compared to CF phenotype cells, but cytokine production remained greater from the CF cell lines. When cells were treated with neuraminidase, PAO1 adherence was equalized between CF and nonCF phenotype cell lines, but IL-8 production in response to inflammatory stimuli was still greater in CF phenotype cells. The polarized cell lines 16HBEo-Sense (normal phenotype) and Antisense (CF phenotype) cells were used to test the effect of disrupting tight junctions, which allows access of PAO1 to basolateral binding sites in both cell lines. IL-8 production increased from CF, but not normal, cells. These data indicate that increased bacterial binding to CF phenotype cells cannot by itself account for excess cytokine production in CF airway epithelial cells, encourage investigation of alternative hypotheses, and signal caution for therapeutic strategies proposed for CF that include disruption of tight junctions in the face of pseudomonas infection.
    Full-text · Article · Feb 2005 · Respiratory research
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    • "Eine mögliche Erklärung der veränderten terminalen Glycosylierung bei der Cystischen Fibrose ist eine Veränderung des Golgi pH-Wertes bzw. des Trans-Golgi-Netzwerkes (Kube et al., 2001; Scanlin und Glick, 1999; Tatterson et al., 2001). Der Golgi pH- Wert liegt in der Regel unter dem des Zytoplasmas zwischen pH 6,2 und pH 6,4 (al-Awqati, 1995; Barasch und al-Awqati, 1993). "
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    ABSTRACT: Hamburg, Univ., Diss., 2004. Computerdatei im Fernzugriff.
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    ABSTRACT: Structural differences have been reported in the glycosylation patterns of cystic fibrosis glycoproteins. Although the gene mutated in cystic fibrosis (CFTR) has been cloned and characterized as a chloride channel, its relationship to the highly viscous mucus and structural glycoprotein and mucin abnormalities in cystic fibrosis still remains to be defined. We have evaluated O-glycan biosynthesis in CHO and BHK cells that express CFTR and DeltaF508 CFTR as in vitro models, and utilized the cftr knockout mouse as an in vivo model of CFTR dysfunction. Activities of glycosyltransferases and sulfotransferases synthesizing mucin type O-glycan chains were determined in these models. Differences in transferase activity levels were found between tissues and cell types and during mouse development. No specific patterns of activities were associated with the lack of CFTR or with DeltaF508CFTR expression. This suggests that it is not the presence or absence of normal CFTR, or the presence of mutant CFTR alone, but rather cell specific additional factors or pathophysiological consequences that determine the changes in mucin glycosylation in cystic fibrosis.
    Full-text · Article · Oct 2001 · Glycoconjugate Journal
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