期刊
CIRCULATION RESEARCH
卷 104, 期 5, 页码 589-U59出版社
LIPPINCOTT WILLIAMS & WILKINS
DOI: 10.1161/CIRCRESAHA.108.183905
关键词
cardiovascular research; endothelial cells; tissue factor; tumor necrosis factor; vascular biology
资金
- Deutsche Forschungsgemeinschaft [GRK 865, SFB-TR19]
- NIH [DK065752]
Tumor necrosis factor (TNF)-alpha-stimulated human umbilical vein endothelial cells express 2 naturally occurring forms of tissue factor (TF), the primary initiator of blood coagulation: the soluble alternatively spliced isoform and the full-length TF isoform. The regulatory pathways enabling this phenomenon are completely unknown. Cdc2-like kinases and DNA topoisomerase I regulate alternative splicing via phosphorylation of serine/arginine-rich proteins. In this study, we examined effects of serine/arginine-rich protein kinases on TF splicing following stimulation with TNF-alpha. Human endothelial cells were pretreated with specific inhibitors or small interfering RNAs against Cdc2-like kinases and DNA topoisomerase I before stimulation with TNF-alpha. TF levels were determined by semiquantitative RT-PCR, real-time PCR, and Western blotting. Cellular procoagulant activity was analyzed in a chromogenic TF activity assay. All 4 known Cdc2-like kinases forms were expressed in human endothelial cells. Selective inhibition of Cdc2-like kinases and DNA topoisomerase I elicited distinct changes in TF biosynthesis in TNF-alpha-stimulated endothelial cells, which impacted endothelial procoagulant activity. This study is the first to demonstrate that serine/arginine-rich protein kinases modulate splicing of TF pre-mRNA in human endothelial cells and, consequently, endothelial procoagulant activity under inflammatory conditions. (Circ Res. 2009; 104: 589-599.)
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