title: Extracellular Ca(2+) suppresses endotoxin-inducible tissue factor activation in monocytic THP-1 cells.
abstract: BACKGROUND
  Monocytic tissue factor (TF), an initiator of extrinsic blood coagulation, is often activated under various inflammatory conditions including endotoxemia. This activation could be a contributing factor to the manifestation of disseminated intravascular coagulation following septic shock.
  
  
  OBJECTIVE
  We herein determine if extracellular Ca(2+) ([Ca(2+)](ex)) regulates bacterial endotoxin (LPS)-inducible monocytic TF activation.
  
  
  METHODS
  We have employed a model monocytic cell line (THP-1) to explore the mode of action of [Ca(2+)](ex) on the modulation of LPS-induced TF activation. TF activity was measured by a single stage clotting assay, while TF expression as well as LPS recognition and its receptor expression were studied in immunofluorescent approaches.
  
  
  RESULTS
  LPS-induced TF activation was inversely correlated to [Ca(2+)](ex). Upon exposure of THP-1 cells to LPS (1.5 microg ml(-1)) for 6 h in the Hanks' medium without CaCl(2), TF was activated by nearly 10-fold. TF activation appreciably decreased with the increasing [Ca(2+)](ex). No more than 3.5-fold TF activation was detected at 5 mM [Ca(2+)](ex). Consistent with the significantly lower degree of TF activation, LPS-induced TF expression at 5 mM [Ca(2+)](ex) was 60 per cent less than that without [Ca(2+)](ex). FACScan analysis showed that LPS recognition was significantly blocked at 5 mM [Ca(2+)](ex) which however had no effect on the expression of CD14 and CD11b, the proposed major LPS receptors. Moreover, LPS binding in vitro was significantly inhibited by 5 mM CaCl(2).
  
  
  CONCLUSIONS
  Our results demonstrate that [Ca(2+)](ex) blocked LPS recognition without affecting its receptor expression on THP-1 monocytes. This insensitivity to LPS thereby resulted in the depressed inducible monocytic TF expression and activation.
authors.affiliation: Department of Surgery, Wayne State University, School of Medicine, Detroit, MI 48201, USA. ad5742@wayne.edu
authors.forename: A J
authors.initials: AJ
authors.lastname: Chu
authors.forename: M J
authors.initials: MJ
authors.lastname: Fox
authors.forename: J K
authors.initials: JK
authors.lastname: Prasad
mesh_terms: D015703:Antigens, CD / Q000378:metabolism; D001777:Blood Coagulation; D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D016650:Fluorescein-5-isothiocyanate; D005455:Fluorescent Antibody Technique; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009000:Monocytes / Q000378:metabolism*; D013925:Thromboplastin / Q000378:metabolism*
chemical_list: D015703:Antigens, CD; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D013925:Thromboplastin; D016650:Fluorescein-5-isothiocyanate; D002118:Calcium