title: Potent cardioprotection from ischemia-reperfusion injury by a two-domain fusion protein comprising annexin V and Kunitz protease inhibitor.
abstract: BACKGROUND
  Considerable evidence suggests that coagulation proteases (tissue factor [TF]/activated factor VII [FVIIa]/FXa/thrombin) and their target protease activated receptors (PAR-1/PAR-2) play important roles in myocardial ischemia-reperfusion (I-R) injury. We hypothesized that localized inhibition of TF/FVIIa on the membrane surfaces of ischemic cells could effectively block coagulation cascade and subsequent PAR-1/PAR-2 cell signaling, thereby protecting the myocardium from I-R injury.
  
  
  OBJECTIVE
  We recently developed an annexin V-Kunitz inhibitor fusion protein (ANV-6L15) that could specifically bind to anionic phospholipids on the membrane surfaces of apoptotic cells and efficiently inhibit the membrane-anchored TF/FVIIa. In this study, we investigated the cardioprotective effect of ANV-6L15 in a rat cardiac I-R model in comparison with that of hirudin.
  
  
  METHODS
  Left coronary artery occlusion was maintained for 45 min followed by 4 h of reperfusion in anesthetized Sprague-Dawley rats. One minute before or 2 min after coronary ligation, rats received an intravenous bolus injection of ANV-6L15 (2.5-250 μg kg(-1) ), vehicle, or hirudin via bolus injection and continuous infusion.
  
  
  CONCLUSIONS
  ANV-6L15 dose-dependently reduced infarct size by up to 87% and decreased plasma levels of cardiac troponin I, tumor necrosis factor-α, and soluble intercellular adhesion molecule-1, by up to 97%, 96%, and 66%, respectively, with little impact on the coagulation parameters. ANV-6L15 also ameliorated hemodynamic derangements, attenuated neutrophil infiltration and reduced Terminal deoxynucleotidyl transferase dUTP nick end labeling-positive apoptotic cardiomyocytes. Hirudin was less efficacious even at supraclinical dose. ANV-6L15 confers exceptionally potent cardioprotection and is a promising drug candidate for the prevention of myocardial I-R injury.
authors.affiliation: Division of Thoracic & Cardiovascular Surgery, Chang Gung Memorial Hospital-Keelung, Keelung, Taiwan. yehccl@cgmh.org.tw
authors.forename: C-H
authors.initials: CH
authors.lastname: Yeh
authors.forename: T-P
authors.initials: TP
authors.lastname: Chen
authors.forename: Y-C
authors.initials: YC
authors.lastname: Wang
authors.forename: S-W
authors.initials: SW
authors.lastname: Fang
authors.forename: T-C
authors.initials: TC
authors.lastname: Wun
mesh_terms: D000818:Animals; D017304:Annexin A5 / Q000737:chemistry*; D017209:Apoptosis; D007611:Aprotinin / Q000737:chemistry*; D002316:Cardiotonic Agents / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D006439:Hemodynamics; D020287:In Situ Nick-End Labeling; D008297:Male; D015428:Myocardial Reperfusion Injury / Q000517:prevention & control*; D009504:Neutrophils / Q000378:metabolism; D010743:Phospholipids / Q000737:chemistry; D017434:Protein Structure, Tertiary; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry*; D019210:Troponin I / Q000097:blood; D014409:Tumor Necrosis Factor-alpha / Q000097:blood
chemical_list: C587037:ANV-6L15 protein; D017304:Annexin A5; D002316:Cardiotonic Agents; D010743:Phospholipids; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D019210:Troponin I; D014409:Tumor Necrosis Factor-alpha; C098973:recombinant Kunitz protease inhibitor; D007611:Aprotinin
keywords: annexin V; apoptosis; cardioprotective agents; ischemia-reperfusion injury; proteinase-activated; receptors