Polymorphisms in the 5′-UTR of the tissue factor gene are associated with altered expression in human endothelial cells

Abstract

Background: Enhanced tissue factor (TF) expression mediates many disease processes. Recently, four completely concordant polymorphisms were detected in the 5′-UTR of the TF gene. Three were single base changes and one was an 18-bp insertion/deletion at-1208. Objectives: This study was undertaken to determine if the I-allele or the D-allele would associate with elevated TF expression in human umbilical vein endothelial cells (HUVEC). Methods: HUVEC were genotyped by polymerase chain reaction for 18-bp insert status. TF expression was induced by interleukin (IL)-1 or phorbol 12-myristate 13-acetate (PMA). Total TF activity was determined by a one-stage clotting assay and surface TF activity by a chromogenic assay. Protein binding differences between the I- and D-alleles were examined by gel shift assays. Results: IL-1- or PMA-induced total TF activity in D-allele HUVEC was increased 2.0-2.5-fold above that seen in II HUVEC. Surface clotting activity in D-allele cells was 1.3-1.7-fold greater than in II-allele cultures. Experiments with consensus site mutation oligos suggested that the 18-bp insert creates GATA and CCAAT-enhancer binding protein (C/EBP) transcription factor recognition sites. Conclusions: The D-allele is associated with enhanced TF activity in HUVEC. The differences in TF expression between the alleles may be due to variant transcription factor binding in the -1208 region. Further studies are warranted to investigate whether the D-allele is associated with increased incidence of pathological processes that involve TF. © 2004 International Society on Thrombosis and Haemostasis.