FEBS Letters | |
Identification of syntenin and other TNF‐inducible genes in human umbilical arterial endothelial cells by suppression subtractive hybridization | |
Schulze-Osthoff, Klaus1  Neuhaus, Thomas2  Totzke, Gudrun2  Grünewald, Elisabeth2  Vetter, Hans2  Stier, Sebastian2  Ko, Yon2  Sachinidis, Agapios2  Fronhoffs, Stefan2  | |
[1] Department of Immunology and Cell Biology, University of Münster, Münster, Germany;Medizinische Poliklinik, University of Bonn, Wilhelmstr. 35-37, D-53111 Bonn, Germany | |
关键词: Adhesion molecule; Endothelial cell; Syntenin; Nuclear factor-κB; Suppression subtractive hybridization; Tumor necrosis factor; HUAEC; human umbilical arterial endothelial cells; IL; interleukin; MCP-1; monocyte chemoattractant protein-1; NF-κB; nuclear factor-κB; IκB; inhibitor of NF-κB; PCR; polymerase chain reaction; RT; reverse transcription; SSH; suppression subtractive hybridization; TNF; tumor necrosis factor-α; VCAM-1; vascular cell adhesion molecule-1; | |
DOI : 10.1016/S0014-5793(00)01177-7 | |
学科分类:生物化学/生物物理 | |
来源: John Wiley & Sons Ltd. | |
【 摘 要 】
Endothelial cells play an important regulatory role in inflammatory responses by upregulating various proinflammatory gene products including cytokines and adhesion molecules. A highly potent mediator of this process is tumor necrosis factor-α (TNF). In the present study, the suppression subtractive hybridization (SSH) method was employed to identify rarely transcribed TNF-inducible genes in human umbilical arterial endothelial cells. Following mRNA isolation of non-stimulated and TNF-stimulated cells, cDNAs of both populations were prepared and subtracted by suppression PCR. Sequencing of the enriched cDNAs identified 12 genes differentially expressed including vascular cell adhesion molecule-1, monocyte chemoattractant protein-1, interleukin-8 and IκBα, an inhibitor of the transcription factor nuclear factor-κB. Interestingly, also syntenin, a PDZ motif-containing protein which binds to the cytoplasmic domain of syndecans, was identified by SSH. Time course studies using RT-PCR analysis confirmed that all genes were differentially expressed and rapidly induced by TNF. Our data reveal that SSH is a powerful technique of high sensitivity for the detection of differential gene expression in primary arterial endothelial cells.
【 授权许可】
Unknown
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