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EBV-transformed B-cells from an individual homozygously mutated (G329A) in FUT7 do not roll on E-selectin
Linköping University, Department of Biomedicine and Surgery, Clinical Chemistry. Linköping University, Faculty of Health Sciences.
Institute of Laboratory Medicine, Department of Clinical Chemistry and Transfursion Medicine, Sahlgrenska University Hospital, Göteborg, Sweden.
Institute of Laboratory Medicine, Department of Clinical Chemistry and Transfursion Medicine, Sahlgrenska University Hospital, Göteborg, Sweden.
Linköping University, Department of Biomedicine and Surgery, Clinical Chemistry. Linköping University, Faculty of Health Sciences.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

The α1,3 fucosyltransferase VII (Fuc-TVII) is involved in the biosynthesis of E- and P-selectin ligands such as sialyl Lewis x on human leukocytes. Recently, individuals were characterized carrying a missense mutation (G329A; Arg110-Gln) in the FUT7 gene encoding this enzyme. The mutated FUT7 construct was not able to produce an active Fuc-TVII enzyme in transfection studies and polymorphonuclear granulocytes from an individual carrying the mutation homozygously showed severely reduced expression of sialyl Lewis x. In the present study we have established Epstein-Barr virus transformed B-celllines from this individual (SIGN) and from an individual not carrying the mutation (IWO). The cell lines were confirmed to be of B-cell origin by flow cytometry analysis. IWO cells interacted with E-selectin in an in vitro flow chamber analysis whereas SIGN cell did not. However, when SIGN cell were transfected with wild type FUT7 cDNA interaction with E-selectin could be restored. Cell surface expression of the sLex related epitopes recognized by antibodies CSLEX-1, KM-93 and HECA-452 was elevated on IWO cells compared to SIGN cells, consistent with a role of these antigens in E-selectin recognition. Despite high expression of PSGL-1 neither of the cell lines interacted with P-selectin under flow. These cell lines may be useful in the further characterization ofEselectin ligands and the obtained results encourage further studies on the consequences of the FUT7-G329A mutation in vivo.

National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-81861OAI: oai:DiVA.org:liu-81861DiVA: diva2:556111
Funder
Swedish Research Council, 0002Swedish Research Council, 8266
Available from: 2012-09-24 Created: 2012-09-24 Last updated: 2012-09-24Bibliographically approved
In thesis
1. Carbohydrate dependent adhesion of leukocytes and the role of fucosyltransferase VII
Open this publication in new window or tab >>Carbohydrate dependent adhesion of leukocytes and the role of fucosyltransferase VII
2003 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Galectins, E-and P-selectins are carbohydrate-binding proteins that are up regulated at inflammatory lesions. Selectins expressed ou the activated endothelium mediate transient binding to leukocyte ligands that require sequential action of α 2,3-sialyltransferases and a 1,3-fucosyltransferases. ln leukocytes α1,3 fucosyltransferase VII adds fucose to α 2,3- sialylated lactosarnine acceptors in the final step of the biosynthesis of the selectin binding epitope sialyl Lewis x.

The finding of low sialyl Lewis x expression in polymorphonuclear leukocytes from a patient with ulcerative colitis led to the discovery of a missense mutation (G329A) in the human gene coding for α1,3 fucosyltransferase VII, FUT7. Studies including enzymatic and immunochemical analysis oftransfected cell lines and isolated polymorphonuclear leukocytes from patients confirmed that this mutation impair the ability of α1,3 fucosyltransferase VII to synthesize sialyl Lewis x. The frequency of the mutation were measured in a mixed Swedish population and found to be -1 %. One individual carrying the FUT7-G329A mutation homozygously was identified. This individual suffered from ulcer disease, noninsulin-dependent diabetes, osteoporosis, spondyloarthrosis, and Sjögren's syndrome, but the relationship between disease and the mutation is not established.

Studies using an in vitro flow chamber assay showed that transfected B-lymphoma cells and Epstein-Barr virus transformed B cells only transcribing FUT7-G329A were not able to interact with E-selectin during flow whereas polymorphonuclear leukocytes from the FUT7-G329A homozygous individual interacted well with both P- and E selectin during flow. The mRNA level of the fucosyltransferase IV coding gene, FUT4, was found to be elevated in the homozygous individual, which resulted in elevated levels ofthe CD65s epitope. However, transfected B-lymphoma cells and Epstein- Barr virus transformed cells did not show a similar elevation of FUT4 mRNA. In in vitro studies galectin-1 and- 3 were observed to be able to recruit polymorphonuclear leukocytes during flow. This thesis gives further insight into the molecular mechanisms leading to carbohydrate dependent dynamic adhesion between polymorphonuclear leukocytes and lectins during inflammation.

Place, publisher, year, edition, pages
Linköping: Linköpings universitet, 2003. 41 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 762
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-25551 (URN)9999 (Local ID)91-7373-204-4 (ISBN)9999 (Archive number)9999 (OAI)
Public defence
2003-03-28, Berzeliussalen, Universitetssjukhuset, 13:00 (Swedish)
Opponent
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2012-09-24Bibliographically approved

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Bengtson, PerPåhlsson, Peter

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