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Protection of vascular barrier integrity by activated protein C in murine models depends on protease-activated receptor-1.

ABSTRACT: Protease activated receptor-1 (PAR1) mediates barrier protective signalling of activated protein C (APC) in human endothelial cells in vitro and may contribute to APC's beneficial effects in patients with severe sepsis. Mouse models are of key importance for translational research but species differences may limit conclusions for the human system. We analysed whether mouse APC can cleave, activate and induce signalling through murine PAR1 and tested in newly established mouse models if long-term infusion of APC prevents from vascular leakage. Cell surface immunoassays demonstrated efficient cleavage of endogenous murine endothelial PAR1 by either murine or human APC. Pharmacological concentrations of APC of either species had powerful barrier protective effects on cultured murine endothelial cells that required PAR1 cleavage. Vascular endothelial growth factor-mediated hyperpermeability in the skin was reduced by either endogenously generated as well as directly infused recombinant mouse APC in wild-type mice. However APC did not significantly alter the vascular barrier function in PAR1-deficient mice. In endotoxin-challenged mice, infused APC significantly prevented from pulmonary fluid accumulation in the wild-type mice but not in mice lacking PAR1. Our results directly show that murine APC cleaves and signals through PAR1 in mouse endothelial cells. APC reduces vascular permeability in mouse models and PAR1 plays a major role in mediating these effects. Our data in vitro and in vivo support the paradigm that PAR1 contributes to protective effects of APC on vascular barrier integrity in sepsis.

SUBMITTER: Schuepbach RA

PROVIDER: S-EPMC2680237 | biostudies-literature | 2009 Apr

REPOSITORIES: biostudies-literature

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Protection of vascular barrier integrity by activated protein C in murine models depends on protease-activated receptor-1.

Schuepbach Reto A RA Feistritzer Clemens C Fernández José A JA Griffin John H JH Riewald Matthias M

Thrombosis and haemostasis 20090401 4

Protease activated receptor-1 (PAR1) mediates barrier protective signalling of activated protein C (APC) in human endothelial cells in vitro and may contribute to APC's beneficial effects in patients with severe sepsis. Mouse models are of key importance for translational research but species differences may limit conclusions for the human system. We analysed whether mouse APC can cleave, activate and induce signalling through murine PAR1 and tested in newly established mouse models if long-term ...[more]

PMID: 19350118

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Project description:Activation of the protease-activated receptor 2 (PAR2) or the transient receptor potential vanilloid type 1 (TRPV1) channels expressed in cardiac sensory afferents containing calcitonin gene-related peptide (CGRP) and/or substance P (SP) has been proposed to play a protective role in myocardial ischemia-reperfusion (I/R) injury. However, the interaction between PAR2 and TRPV1 is largely unknown. Using gene-targeted TRPV1-null mutant (TRPV1(-/-)) or wild-type (WT) mice, we test the hypothesis that TRPV1 contributes to PAR2-mediated cardiac protection via increasing the release of CGRP and SP. Immunofluorescence labeling showed that TRPV1 coexpressed with PAR2, PKC-epsilon, or PKAc in cardiomyocytes, cardiac blood vessels, and perivascular nerves in WT but not TRPV1(-/-) hearts. WT or TRPV1(-/-) hearts were Langendorff perfused with the selective PAR2 agonist, SLIGRL, in the presence or absence of various antagonists, followed by 35 min of global ischemia and 40 min of reperfusion (I/R). The recovery rate of coronary flow, the maximum rate of left ventricular pressure development, left ventricular end-diastolic pressure, and left ventricular developed pressure were evaluated after I/R. SLIGRL improved the recovery of hemodynamic parameters, decreased lactate dehydrogenase release, and reduced the infarct size in both WT and TRPV1(-/-) hearts (P < 0.05). The protection of SLIGRL was significantly surpassed for WT compared with TRPV1(-/-) hearts (P < 0.05). CGRP(8-37), a selective CGRP receptor antagonist, RP67580, a selective neurokinin-1 receptor antagonist, PKC-epsilon V1-2, a selective PKC-epsilon inhibitor, or H-89, a selective PKA inhibitor, abolished SLIGRL protection by inhibiting the recovery of the rate of coronary flow, maximum rate of left ventricular pressure development, and left ventricular developed pressure, and increasing left ventricular end-diastolic pressure in WT but not TRPV1(-/-) hearts. Radioimmunoassay showed that SLIGRL increased the release of CGRP and SP in WT but not TRPV1(-/-) hearts (P < 0.05), which were prevented by PKC-epsilon V1-2 and H-89. Thus our data show that PAR2 activation improves cardiac recovery after I/R injury in WT and TRPV1(-/-) hearts, with a greater effect in the former, suggesting that PAR2-mediated protection is TRPV1 dependent and independent, and that dysfunctional TRPV1 impairs PAR2 action. PAR2 activation of the PKC-epsilon or PKA pathway stimulates or sensitizes TRPV1 in WT hearts, leading to the release of CGRP and SP that contribute, at least in part, to PAR2-induced cardiac protection against I/R injury.

Dataset Information

Protection of vascular barrier integrity by activated protein C in murine models depends on protease-activated receptor-1.

Publications

Protection of vascular barrier integrity by activated protein C in murine models depends on protease-activated receptor-1.

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