title: Induction of plasminogen activator inhibitor type 1 and type 1 collagen expression in rat cardiac microvascular endothelial cells by interleukin-1 and its dependence on oxygen-centered free radicals.
abstract: BACKGROUND
  Ischemia with or without reperfusion induces the release of diverse products from monocytes, including cytokines such as interleukin-1 (IL-1). To determine whether these phenomena modulate fibrinolysis and potentially exacerbate impairment of the macrocirculation, microcirculation, or both, we characterized the effects of IL-1 on the expression of fibrinolytic system and matrix proteins in rat cardiac microvascular endothelial cells (CMECs).
  
  
  RESULTS
  Confluent CMECs were exposed to IL-1 in serum-free medium for 24 hours, and cell-conditioned medium was assayed for plasminogen activator inhibitor type 1 (PAI-1), the primary physiological inhibitor of plasminogen activators, and for type 1 collagen with Western blotting. IL-1 (2 ng/mL) specifically increased the accumulation of PAI-1 (4.4 +/- 0.6-fold; mean +/- SD; n = 9) without affecting tissue plasminogen activator (t-PA) or urokinase plasminogen activator (u-PA) levels, which remained unchanged. IL-1 increased the accumulation of collagen in conditioned media by 3.5 +/- 0.7-fold (n = 6). Conversely, the accumulation of both PAI-1 and collagen induced by IL-1 was inhibited with an IL-1 receptor antagonist (200 ng/mL; n = 6) and with cycloheximide (10 micrograms/mL; n = 6), implying that protein synthesis was a requirement for the effect. To determine whether the IL-1 effect was mediated by induction of oxygen-centered free radical production, known to be induced by IL-1, we exposed the cells to the hydroxyl radical scavenger tetramethylthiourea (10 mmol/L) and observed abolition of the IL-1-induced increase in the expression of PAI-1 and collagen (n = 6). Conversely, superoxides (generated with 10 mU/mL xanthine oxidase plus 0.6 mmol/L hypoxanthine, and 100 mumol/L hydrogen peroxide) induced the accumulation of PAI-1 and collagen (n = 6). IL-1 (1 microgram/kg body wt) and lipopolysaccharide (50 micrograms/kg body wt) administered in vivo increased PAI-1 protein in rat hearts as detected with Western blotting and PAI-1 immunostaining of rat heart microvessels, indicating the effects delineated in vitro were paralleled by effects in vivo.
  
  
  CONCLUSIONS
  These results indicate that IL-1-induced oxygen-centered free radicals stimulate elaboration of PAI-1 and collagen by CMECs. Accordingly, microvascularly mediated inhibition of fibrinolysis may predispose to the persistence of microvascular thrombi, thereby contributing to impaired microcirculatory function, the no-reflow phenomenon, and cardiac dysfunction after ischemia and reperfusion.
authors.affiliation: Department of Medicine, University of Vermont, College of Medicine, Burlington 05405, USA.
authors.forename: H
authors.initials: H
authors.lastname: Okada
authors.forename: J
authors.initials: J
authors.lastname: Woodcock-Mitchell
authors.forename: J
authors.initials: J
authors.lastname: Mitchell
authors.forename: T
authors.initials: T
authors.lastname: Sakamoto
authors.forename: K
authors.initials: K
authors.lastname: Marutsuka
authors.forename: B E
authors.initials: BE
authors.lastname: Sobel
authors.forename: S
authors.initials: S
authors.lastname: Fujii
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis*; D003331:Coronary Vessels / Q000187:drug effects* / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism; D005609:Free Radicals; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1 / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D017395:Plasminogen Activator Inhibitor 1 / Q000096:biosynthesis*; D011817:Rabbits; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012795:Sialoglycoproteins / Q000494:pharmacology
chemical_list: D005609:Free Radicals; C507764:Il1rn protein, mouse; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1; D008070:Lipopolysaccharides; D017395:Plasminogen Activator Inhibitor 1; D017382:Reactive Oxygen Species; D012795:Sialoglycoproteins; D003094:Collagen