PAR1 antagonist mediated antithrombotic activity in extracorporeal arterio-venous shunt in the rat

Abstract

The activation of platelets by thrombin is mediated at least in part by cleavage of protease-activated receptors (PARs) [ [1] Vu T.K. Hung D.T. Wheaton V.I. Coughlin S.R. Molecular cloning of a functional thrombin receptor reveals a novel proteolytic mechanism of receptor activation. Cell. 1991; 64: 1057-1068 Abstract Full Text PDF PubMed Scopus (2641) Google Scholar ]. Four distinct PARs have been cloned to date, with PAR1, PAR3 and PAR4 being activated by thrombin, whereas PAR2 is sensitive to trypsin [ [2] Ossovskaya V.S. Bunnett N.W. Protease-activated receptors: contribution to physiology and disease. Physiol Rev. 2004; 84: 579-621 Crossref PubMed Scopus (908) Google Scholar ]. Human platelets express PAR1 and PAR4 and activation of either is sufficient to trigger platelet aggregation and secretion [ 1 Vu T.K. Hung D.T. Wheaton V.I. Coughlin S.R. Molecular cloning of a functional thrombin receptor reveals a novel proteolytic mechanism of receptor activation. Cell. 1991; 64: 1057-1068 Abstract Full Text PDF PubMed Scopus (2641) Google Scholar , 3 Kahn M.L. Nakanishi-Matsui M. Shapiro M.J. Ishihara H. Coughlin S.R. Protease-activated receptors 1 and 4 mediate activation of human platelets by thrombin. J Clin Invest. 1999; 103: 879-887 Crossref PubMed Scopus (680) Google Scholar ]. Studies with selective neutralizing antibodies have indicated that PAR1 mediates activation of human platelets at low thrombin concentrations and that, in the absence of PAR1 function, PAR4 can mediate platelet activation but only at high thrombin concentrations [ [4] Coughlin S.R. Thrombin signalling and protease-activated receptors. Nature. 2000; 407: 258-264 Crossref PubMed Scopus (2083) Google Scholar ]. Curiously, PAR1 appears not to play a role in rat platelets. PAR1-activating peptides do not activate rat platelet aggregation [ 5 Kinlough-Rathbone R.L. Rand M.L. Packham M.A. Rabbit and rat platelets do not respond to thrombin receptor peptides that activate human platelets. Blood. 1993; 82: 103-106 PubMed Google Scholar , 6 Derian C.K. Santulli R.J. Tomko K.A. Haertlein B.J. Andrande-Gordon P. Species differences in platelet responses to thrombin and SFLLRN receptor-mediated calcium mobilization and aggregation, and regulation by protein kinases. Thromb Res. 1995; 78: 505-519 Abstract Full Text PDF PubMed Scopus (65) Google Scholar ]. In contrast to this, PAR3 is not expressed by human or guinea-pig platelets, but by mouse and rat platelets [ [7] Hishihara H. Connolly A.J. Zeng D. Kahn M.L. Zheng Y.W. Timmons C. et al. Protease-activated receptor 3 is a second thrombin receptor in humans. Nature. 1997; 386: 502-506 Crossref PubMed Scopus (794) Google Scholar ]. Recently, Leger et al. [ [8] Leger A.J. Jacques S.L. Badar J. Kaneider N.C. Derian C.K. Andrade-Gordon P. et al. Blocking the protease-activated receptor 1-4 heterodimer in platelet-mediated thrombosis. Circulation. 2006; 113: 1244-1254 Crossref PubMed Scopus (187) Google Scholar ] elegantly showed that in human platelets PAR1 and PAR4 form a stable heterodimer that enables thrombin to act as a bivalent functional agonist. Collectively, all these studies reveal the complexity of the PAR-mediated platelet activation in different animal species. Thus, the aim of the present study was to investigate the putative in vivo role of PAR1 in a thrombosis model conducted in anesthetized rats. We have investigated the effects of selective PAR4 antagonist, YD-3 [ [9] Wu C.C. Hwang T.L. Liao C.H. Kuo S.C. Lee F.Y. Lee C.Y. et al. Selective inhibition of protease-activated receptor 4-dependent platelet activation by YD-3. Thromb Haemost. 2002; 87: 1026-1033 PubMed Google Scholar ] but also relatively selective PAR1 antagonists ER-121958 and SCH-203099 [ 10 Ahn H.S. Foster C. Boykow G. Stamford A. Manna M. Graziano M. Inhibition of cellular action of thrombin by N3-cyclopropyl-7-[[4-(1-methylethyl)phenyl]methyl]-7H-pyrrolo[3,2-f]quinazoline-1,3-diamine (SCH 79797), a nonpeptide thrombin receptor antagonist. Biochem Pharmacol. 2000; 60: 1425-1434 Crossref PubMed Scopus (138) Google Scholar , 11 Ahn H.S. Chackalamannil S. Nonpeptide thrombin receptor antagonists. Drugs Future. 2001; 26: 1065-1085 Crossref Scopus (13) Google Scholar , 12 Chackalamannil S. Xia Y. Thrombin receptor (PAR-1) antagonists as novel antithrombotic agents. Expert Opin Ther Patents. 2006; 16: 493-505 Crossref PubMed Scopus (27) Google Scholar ] and conventional antithrombotic agents on an extracorporeal arteriovenous shunt model in the rat.