title: In situ assays demonstrate that interferon-gamma suppresses infection-stimulated hepatic fibrin deposition by promoting fibrinolysis.
abstract: BACKGROUND
  Inflammatory cytokines potently impact hemostatic pathways during infection, but the tissue-specific regulation of coagulation and fibrinolysis complicates studies of the underlying mechanisms.
  
  
  RESULTS
  Here, we describe assays that quantitatively measuring prothrombinase (PTase), protein C-ase (PCase) and plasminogen activator (PA) activities in situ, thereby facilitating studies of tissue-specific hemostasis. Using these assays, we investigate the mechanisms regulating hepatic fibrin deposition during murine toxoplasmosis and the means by which interferon-gamma (IFN-gamma) suppresses infection-stimulated fibrin deposition. We demonstrate that Toxoplasma infection upregulates hepatic PTase, PCase, and PA activity. Wild type and gene-targeted IFN-gamma-deficient mice exhibit similar levels of infection-stimulated PTase activity. By contrast, IFN-gamma-deficiency is associated with increased PCase activity and reduced PA activity during infection. Parallel analyses of hepatic gene expression reveal that IFN-gamma-deficiency is associated with increased expression of thrombomodulin (TM), a key component of the PCase, increased expression of thrombin-activatable fibrinolysis inhibitor (TAFI), a PC substrate, and reduced expression of urokinase PA (u-PA).
  
  
  CONCLUSIONS
  These findings suggest that IFN-gamma suppresses infection-stimulated hepatic fibrin deposition by suppressing TM-mediated activation of TAFI, thereby destabilizing fibrin deposits, and concomitantly increasing hepatic u-PA activity, thereby promoting fibrinolysis. We anticipate that further application of these in situ assays will improve our understanding of tissue-specific hemostasis, its regulation by cytokines, and its dysregulation during coagulopathy.
authors.affiliation: Trudeau Institute, 154 Algonquin Avenue, Saranac Lake, NY 12983, USA.
authors.forename: I K
authors.initials: IK
authors.lastname: Mullarky
authors.forename: F M
authors.initials: FM
authors.lastname: Szaba
authors.forename: C G
authors.initials: CG
authors.lastname: Winchel
authors.forename: M A
authors.initials: MA
authors.lastname: Parent
authors.forename: L W
authors.initials: LW
authors.lastname: Kummer
authors.forename: N
authors.initials: N
authors.lastname: Mackman
authors.forename: L L
authors.initials: LL
authors.lastname: Johnson
authors.forename: S T
authors.initials: ST
authors.lastname: Smiley
mesh_terms: D000818:Animals; D025901:Carboxypeptidase B2 / Q000378:metabolism; D005337:Fibrin / Q000378:metabolism*; D005342:Fibrinolysis; D006487:Hemostasis; D007239:Infections / Q000378:metabolism*; D007371:Interferon-gamma / Q000172:deficiency / Q000502:physiology*; D008099:Liver / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D010960:Plasminogen Activators / Q000032:analysis; D018180:Thrombomodulin / Q000032:analysis; D013925:Thromboplastin / Q000032:analysis; D014124:Toxoplasmosis, Animal; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
chemical_list: D018180:Thrombomodulin; D007371:Interferon-gamma; D005337:Fibrin; D013925:Thromboplastin; D025901:Carboxypeptidase B2; D010960:Plasminogen Activators; D014568:Urokinase-Type Plasminogen Activator