title: Anti-human protein S antibody induces tissue factor expression through a direct interaction with platelet phosphofructokinase.
abstract: BACKGROUND
  Autoantibodies including anti-human protein S antibody (anti-hPS Ab) and anti-human protein C antibody (anti-hPC Ab) can be detected in patients with autoimmune diseases with hypercoagulability. The objective of the present study was to determine the effects and molecular pathways of these autoantibodies on tissue factor (TF) expression in human coronary artery endothelial cells (HCAECs).
  
  
  METHODS
  HCAECs were treated with anti-hPS Ab or anti-hPC Ab for 3 hours. TF expression was measured by real-time PCR and Western blot. TF-mediated procoagulant activity was determined by a commercial kit. MAPK phosphorylation was analyzed by Bio-Plex luminex immunoassay and Western blot. The potential proteins interacting with anti-hPS Ab were studied by immunoprecipitation, mass spectrometry and in vitro pull-down assay.
  
  
  RESULTS
  Anti-hPS Ab, but not anti-hPC Ab, specifically induced TF expression and TF-mediated procoagulant activity in HCAECs in a concentration-dependent manner. This effect was confirmed in human umbilical endothelial cells (HUVECs). ERK1/2 phosphorylation was induced by anti-hPS Ab treatment, while inhibition of ERK1/2 by U0216 partially blocked anti-hPS Ab-induced TF upregulation (P<0.05). In addition, anti-hPS Ab specifically cross-interacted with platelet phosphofructokinase (PFKP) in HCAECs. Anti-hPS Ab was able to directly inhibit PFKP activities in HCAECs. Furthermore, silencing of PFKP by PFKP shRNA resulted in TF upregulation in HCAECs, while activation of PFKP by fructose-6-phosphate partially blocked the effect of anti-hPS Ab on TF upregulation (P<0.05).
  
  
  CONCLUSIONS
  Anti-hPS Ab induces TF expression through a direct interaction with PFKP and ERK1/2 activation in HCAECs. Anti-hPS Ab may directly contribute to vascular thrombosis in the patient with autoimmune disorders.
authors: Molecular Surgeon Research Center, Division of Surgical Research, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX, USA. Electronic address: jchen@bcm.tmc.edu.
authors: Changyi
authors: C
authors: Chen
authors: Molecular Surgeon Research Center, Division of Surgical Research, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX, USA.
authors: Dan
authors: D
authors: Liao
authors: Molecular Surgeon Research Center, Division of Surgical Research, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX, USA.
authors: Jing
authors: J
authors: Wang
authors: Molecular Surgeon Research Center, Division of Surgical Research, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX, USA.
authors: Zhengdong
authors: Z
authors: Liang
authors: Molecular Surgeon Research Center, Division of Surgical Research, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX, USA.
authors: Qizhi
authors: Q
authors: Yao
mesh_terms: D001323:Autoantibodies / Q000276:immunology*; D002460:Cell Line; D003331:Coronary Vessels / Q000166:cytology*; D042783:Endothelial Cells / Q000276:immunology* / Q000378:metabolism; D005786:Gene Expression Regulation; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D020935:MAP Kinase Signaling System; D025502:Phosphofructokinase-1, Type C / Q000276:immunology*; D017293:Protein S / Q000276:immunology*; D013925:Thromboplastin / Q000235:genetics*
chemical_list: D001323:Autoantibodies; D017293:Protein S; D013925:Thromboplastin; D025502:Phosphofructokinase-1, Type C
keywords: Anti-human protein S antibody; Autoantibody; ERK1/2; Endothelial cell; Platelet phosphofructokinase; Tissue factor