Plasmin Induces Endothelium-Dependent Nitric Oxide–Mediated Relaxation in the Porcine Coronary Artery

Abstract

Objective-Plasmin is a key enzyme in fibrinolysis. We attempted to determine the possible role of plasmin in the regulation of vascular tone, while also investigating the mechanism of plasmin-induced vasorelaxation. Methods and Results-In porcine coronary artery, plasmin induced an endothelium-dependent relaxation. This relaxing effect was mostly abolished by a proteinase inhibitor, a plasmin inhibitor, or a nitric oxide (NO) synthase inhibitor. The preceding stimulation with plasmin significantly inhibited the subsequent relaxation induced by thrombin but not that induced by proteinase-activated receptor-1-activating peptide. The relaxation induced by trypsin and substance P remained unaffected by the preceding plasmin stimulation. The pretreatment with plasmin, thrombin, or trypsin significantly attenuated the plasmin-induced relaxation. In porcine coronary artery endothelial cells ( Key Words: plasmin Ⅲ proteinase-activated receptor Ⅲ vasorelaxation Ⅲ nitric oxide Ⅲ endothelium P roteinases involved in the blood coagulation cascade not only play an important role in hemostasis but also exert various vascular effects such as the regulation of the vascular tone, tissue remodeling, and angiogenesis. 1,2 Proteinase-activated receptors (PARs) play a major role in mediating such cellular effects of proteinases. [3][4][5][6] PARs belong to a unique family of G protein-coupled receptor. 3 Four members of PARs have been cloned. PAR1, PAR3, and PAR4 serve as receptors for thrombin, whereas PAR1, PAR2, and PAR4 serve as receptors for trypsin. 3 The activation of PAR is initiated by the proteolytic cleavage at the specific site of the extracellular domain. 3 Under physiological conditions, thrombin and trypsin mainly have been reported to induce an endothelium-dependent vasorelaxation in various type of blood vessel. [5][6][7] We have previously reported that thrombin and trypsin induced a transient elevation of cytosolic Ca 2ϩ concentration ([Ca 2ϩ ] i ) in vascular and aortic valve endothelial cells 8 -10 and an endothelium-dependent relaxation in the porcine coronary artery. 8,9,10,11 Plasmin, as a key enzyme in fibrinolysis, has been reported to induce the cell migration in Chinese hamster ovary cells, 12 the cell proliferation in the murine thoracic aorta smooth muscle cells, 13 and an increase in the expression of Cyr61, a growth factor-like gene, in fibroblast. 14 These reports suggested PAR1 to mediate the cellular effects of plasmin. However, the roles of plasmin in the regulation of vascular tone still remain to be determined. Plasmin has been reported to cleave PAR1 at the C-terminal side of the tethered ligand region, thus inactivating the responsiveness of PAR1. 15,16 Plasmin has been shown to cleave the extracellular region of PAR2 at the N-terminal side of the trypsin cleavage site as well as the trypsin site, 16 although it was also shown to attenuate the subsequent PAR2-mediated [Ca 2ϩ ] i elevation in the rat brain capillary endothelial cell. 17 Therefore, the receptor(s) that mediate the cellular...

Plasmin as a proinflammatory cell activator

“…Further direct effects of plasmin on endothelial cells include activation of PLC, stimulation of the arachidonic acid cascade, including prostacyclin biosynthesis [67], as well as induction of integrin ␣ V ␤ 3 -dependent stress fiber formation [51], cytosolic Ca 2ϩ increase, and NO production, which were independent of any PAR1 activation [68]. Moreover, plasmin was reported to inhibit the thrombin-induced signaling via PAR1, implying the proteolytic inactivation of the thrombin receptor [68]. At high concentrations, plasmin induces detachment of the endothelial cell monolayer and when the stimulation persists, initiation of apoptosis [9,69].…”

Plasmin-Dependent Modulation of the Blood–Brain Barrier: A Major Consideration during tPA-Induced Thrombolysis?

“…In fact, zymographies (either in situ casein zymography 8,26,54,73 or plasminogen-gelatin zymography 51 ) or amidolytic activity assays 54 of the rodent brain, whether performed using naïve 8,54,73 or ischemic tissue, 26,51,54,73 cannot demonstrate any tPA or plasmin activity without inclusion of exogenous plasminogen in these assays. This is suggestive of either negligible levels or a very tight control of active endogenous plasmin in the vascular and extravascular compartments 123,125,142 No Non-specific (streptokinase 142 )…”

“…For example, plasmin initiates in vitro a PAR-1-independent increase in cytosolic Ca 2 þ and nitric oxide production in human and porcine coronary ECs, leading to vasorelaxation and implying plasmin, like tPA, 65,66 as a modulator of vascular tone. 123 In the brain, PAR-1, a substrate for plasmin (see Plasmin Activation of Astrocytes below), could not be detected in rat BECs utilizing in situ hybridization 124 but was later found together with PAR-3 (and PAR-2) on these cells in vitro by RT-PCR. 125 Whether PAR-1 on BECs is expressed in a polar distribution across the apical and basolateral membranes, an important determinant for possible activation by brain-derived proteases, has not been determined 125 (the susceptibility of BECs to blood-born proteases, such as thrombin or plasmin, suggests at least an apical expression).…”

“…When describing the effect of plasmin on astrocytes, an emphasis should be placed on PAR-1, a G-protein coupled receptor most potently activated by thrombin, which can also be cleaved by plasmin. 135,136 However, the action of plasmin on PAR-1 is complex as plasmin can activate 103,136,137 or inactivate 123,125,135,137 the receptor, cleaving the N-terminal exodomain of PAR-1 at the thrombin cleavage site (Arg 41 -Ser 42 ), 137 but also doing so inappropriately in other positions, 135 creating a number of different tethered ligands.…”

Plasmin: A Modulator of Immune Function

“…43 Once dissociated, S100A10 is rapidly ubiquitinated and degraded. 42,43 Notably, plasmin was shown to activate phospholipase A2 in endothelial cells, resulting in the release of arachidonic acid and subsequent production of prostacyclin 44 as well as to enhance NO-mediated vasorelaxation 45 and chemotactic MCP-1 release, 9 consistent with proinflammatory effects of plasmin.…”