Barton, Matthias

Traupe, Tobias

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www.elsevier.com/locate/cardioresCardiovascular Research 61 (2004) 645–647EditorialProtease-activated receptors and EDHF: the icing on the cake?Tobias Traupe, Matthias Barton*Medizinische Poliklinik, Departement fu¨r Innere Medizin, Universita¨tsspital Zu¨rich, Ra¨mistrasse 100, CH-8091 Zu¨rich, SwitzerlandReceived 29 December 2003; accepted 7 January 2004See article by Kawabata et al. (pages 683–692) in thisissue.The regulation of vascular smooth muscle cell tone bythe vascular endothelium was initially described by Furch-gott and Zawadzki, suggesting the existence of an endo-thelium-derived relaxing factor (EDRF) acting throughcGMP, which was later identified as nitric oxide (NO)(see reference [1] for review). Since then, other actions ofNO such as control of cell growth, blood cell–endothelialcell interactions, immunomodulatory functions, and anti-aggregatory effects have been described [2]. In addition toNO and cyclooxygenase products, a third endothelium-derived principle, namely hyperpolarization of vascularsmooth muscle cells in response to an endothelium-derived hyperpolarizing factor (EDHF), has been identi-fied [3]. Production and bioactivity of endothelial factorsare disturbed in cardiovascular diseases, and endothelium-dependent dilator mechanisms are counteracted by vaso-constrictory prostanoids that contribute to abnormal vas-cular function [4].In this issue of Cardiovascular Research, Kawabata etal. report that activating peptides of the protease-activatedreceptor 2 (PAR2), namely trypsin and SLIGRL, causeendothelium-dependent relaxation by activating NO andEDHF pathways in rat mesenteric arteries in vitro. Theauthors show that gap junctions, but not epoxyeicosatrie-noic acids, hydrogen peroxide, or K+ are involved in therelease of EDHF in response to activation of PAR2.Moreover, vascular contractions due to an activating pep-tide of the protease-activated receptor 1 (TFLLR) wereobserved after inhibition of EDHF and NO or afterendothelium removal.0008-6363/$ - see front matter D 2004 European Society of Cardiology. Publishedoi:10.1016/j.cardiores.2004.01.008Abbreviations: EDHF, endothelium-derived hyperpolarizing factor; ET,endothelin; NO, nitric oxide; NOS, nitric oxide synthase; PAR, protease-activated receptor; SLIGRL, Ser-Leu-Ile-Gly-Arg-Leu-amide; TFLLR, Thr-Phe-Leu-Leu-Arg-amide; TRAP, thrombin receptor-activating peptides;VSMC, vascular smooth muscle cell* Corresponding author. Tel.: +41-1-255 5663; fax: +41-1-255 8747.E-mail address: barton@usz.ch (M. Barton).1. Vascular expression and function of protease-activated receptorsThe protease thrombin is generated during activation ofcoagulation and converts fibrinogen to soluble fibrin [5].In addition, thrombin signals directly to endothelial cells,vascular smooth muscle cells (VSMCs), platelets, fibro-blasts and other cell types by cleaving protease-activatedreceptors (PARs), which belong to the G-protein-coupledreceptor family [6]. The discovery of the first PAR, namelyPAR1, provided a new paradigm for receptor activation[7]. Since then, three other PARs have been cloned andshown to play a role in cellular responses to vascularinjury, inflammation and wound healing. Thrombin acts asan agonist on PAR1, PAR3, and PAR4 with differentpotencies [6] whereas PAR2 is activated by both trypsinand the thrombin-cleaved PAR1. Moreover, several throm-bin receptor-activating peptides (TRAPs) also activatePARs [6]. The binding of a PAR agonist to the receptoris irreversible once the receptor is cleaved, and it istherefore rather unlikely that these pathways are ‘‘routine-ly’’ involved in cellular signaling, particularly since thelevel of expression widely differs in various vascular celltypes. Interestingly, PARs are not only expressed inhealthy vascular cells but also in atherosclerotic plaquesand after vascular injury, indicating an important role inresponse to injury and inflammatory processes [5]. Al-though most of the published data suggest not only apathophysiological but also a physiological role for throm-bin receptors in regulation of cardiovascular function, micedeficient in PAR1 or PAR2 both exhibit normal cardiacfunction, blood pressure levels and vasomotor responses toacetylcholine and angiotensin II [8,9].2. PAR1 and vascular function: role of the endotheliumKawabata et al. investigated the vascular response to aPAR1-activating peptide (TFLLR) and found that thispeptide caused only a small NO-dependent relaxation inprecontracted rat aortic rings but a substantial contractiond by Elsevier B.V. All rights reserved.T. Traupe, M. Barton / Cardiovascular Research 61 (2004) 645–647646in quiescent rings that was further increased after nitricoxide synthase (NOS) inhibition. However, in the ratsuperior mesenteric artery PAR1 causes only a weakcontraction in quiescent rings that is substantially increasedby endothelium removal or inhibition of NO- and EDHF-dependent pathways. Additionally, the authors provideevidence suggesting that superoxide does not play a rolefor these responses by using superoxide dismutase (SOD).In contrast to the findings by Kawabata et al. other studiesdemonstrated that thrombin and TRAPs cause a substantialendothelium-dependent relaxation of the aorta and coro-nary artery from different species [10–14]; these effectsappear to be mainly mediated by cyclooxygenase productsand NO [11,14]. Interestingly, after removal of the endo-thelium thrombin caused contractile responses in coronaryarteries as observed in the present study [11,14], whichmay be due to PAR1-linked increases in vascular smoothmuscle cell Ca2 + flux [15,16]. Additionally, contractileresponses after PAR1 activation have been described evenin the presence of an intact endothelium in human umbil-ical and placental arteries [17], in line with Kawabata et al.Taken together, these findings suggest that PAR1-mediatedrelease of endothelium-derived contracting factors(EDCFs) may at least in part be responsible for theobserved vascular contractions.So what is the nature of these EDCFs? It has been shownthat the endothelium produces endothelin-1 (ET-1), prosta-glandin H2/thromboxane A2, and superoxide anion undercertain conditions in response to other vasoactive substances[1,18]. Although the authors demonstrated that superoxideanion is not involved in the contractile responses observed,a possible role for prostaglandin H2/thromboxane A2 or ET-1 was not investigated. Indeed, this has been demonstratedfor PAR1 in the renal artery [19] and for PAR2 as discussedbelow [20,21]. It is noteworthy that thromboxane A2 and itsreceptor have been shown to mediate thrombin-inducedcontractions in endothelium-denuded human arteries [22].Moreover, thrombin also facilitates production and releaseof ET-1 [23,24], which could also explain the PAR1-mediated contractions observed by Kawabata et al. The factthat PAR1 is activated at low thrombin concentrationssuggests a pathophysiological role for this receptor basedon the observations made in these experimental studies. Thedata discussed above indicate functional differences ofPAR1 on endothelial cells and VSMCs depending on thevascular bed investigated. Furthermore, it has been shownthat agonists of PAR1 cause a biphasic vascular response,including both relaxation and contraction [19].3. A link between PAR2 and EDHF?Trypsin and PAR2-activating peptides can induce endo-thelium-dependent relaxation in rat [25] and rabbit aorta[20] as well as in other arteries [26–29], which is mainlydue to NO. Kawabata et al. also evaluated the role of PAR2for endothelium-dependent vasoreactivity in isolated ratmesenteric artery and aorta by using SLIGRL and trypsin.The results of these experiments show that these agonistslead to a substantial NO-dependent relaxation in rat aortaand cause complete endothelium-dependent relaxation in ratmesenteric arteries that is only in part inhibited by NG-nitro-L-arginine methyl ester (L-NAME, a nonselective inhibitorof NOS) and indomethacin (a nonselective inhibitor ofcyclooxygenase). Thus, the authors also present evidencethat another endothelium-dependent principle is involved inPAR2-mediated signaling in the vascular wall of resistancearteries. Kabawata et al. demonstrate that the vasodilatoryeffect in response to PAR2 activation involves EDHF andgap-junction signaling. These observations could beexplained by the caliber of resistance arteries, in line withprevious data showing that the contribution of EDHF forendothelium-dependent relaxation increases with decreasingvessel diameter [30]. Similar observations were made inmice with atherosclerosis [31], and EDHF may contribute toabnormal endothelial vasoreactivity in patients with athero-sclerosis [32]. Kawabata et al. elegantly demonstrate a rolefor EDHF in the PAR2-mediated vasorelaxation in themesenteric artery by blocking small-, intermediate- andlarge-conductance Ca2 +-sensitive K+ channels. The authorsalso show involvement of gap junctions since the relaxantresponse was blocked by carbenoxolone, an inhibitor of gapjunctional intercellular communication. The findings ofKawabata et al. are in agreement with a recent studysuggesting a role for EDHF in PAR2-mediated vasodilationof coronary arteries [33] that may also be functional in othervascular beds such as the gastric artery [34].PAR2-activation did not cause contractions in rat mesen-teric artery rings as reported by Kawabata et al., althoughPAR2-mediated vasoconstriction has been observed in dif-ferent vascular beds and appears to be both endotheliumdependent [20,21] as well as endothelium independent [35].This endothelium-dependent vasoconstrictor response is pos-sibly due to a yet unidentified contractile factor (EDCF) fromhuman endothelial cells and may be mediated via a novelPAR2 receptor subtype, at least in human umbilical veins[21].The findings by Kawabata et al. demonstrate a distinctrole for EDHF and gap junction signaling in PAR2-mediated vascular responses in mesenteric resistance arter-ies. Whether these mechanisms are important for maintain-ing vascular integrity in diseases such as atherosclerosis[32], arterial thrombosis, or restenosis remains to bedetermined.AcknowledgementsThe authors work is supported by the Swiss NationalFoundation (SCORE 32.58421.99 and 32-58426.99/1), theKurt und Senta Hermann Stiftung, and the Hanne Lieber-mann Stiftung, Zu¨rich.T. Traupe, M. Barton / Cardiovascular Research 61 (2004) 645–647 647References[1] Traupe T, Ortmann J, Mu¨nter K, Barton M. Endothelial therapy ofatherosclerosis and its risk factors. Curr Vasc Pharmacol 2003;9:111–21.[2] Harrison DG. Cellular and molecular mechanisms of endothelial celldysfunction. J Clin Invest 1997;9:2153–7.[3] Busse R, Edwards G, Feletou M, Fleming I, Vanhoutte PM, WestonAH. EDHF: bringing the concepts together. Trends Pharmacol Sci2002;8:374–80.[4] Luscher TF, Barton M. Biology of the endothelium. Clin Cardiol1997;8(Suppl. II):II-3–10.[5] Patterson C, Stouffer GA, Madamanchi N, Runge MS. 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Protease-activated receptors and EDHF: the icing on the cake?

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