Papel do Receptor Ativado por Protease (PAR)2 no recrutamento de neutrófilos induzido pelo Fator Ativador de Plaquetas (PAF)

Abstract

At the lung inflammation microenvironment containing platelet-activating factor (PAF), proteases and protease-activated receptor (PAR)2 are also abundantly present. Despite this evidence, a cooperation between PAF and (PAR)2 in lung inflammation has not yet been investigated. Thus, the aim of this study was to evaluate the possible cooperation between PAFR and PAR2 mediating lung inflammation in mice. Initially, male BALB/c mice received intranasal instillation of C-PAF or PAR2 agonist. In some protocols, mice were pre-treated with intraperitoneal injection with the PAF receptor antagonist, PAR2 receptor antagonist, or protease inhibitor. Bronchoalveolar lavage or lung tissue were collected for quantification of neutrophil numbers, cytokine concentration, histopathological analysis, and measurement of myeloperoxidase and N-Acetyl-BD-glucosaminidase enzymes. In addition, analysis of leukocyte rolling and adhesion was performed by intravital microscopy. For cell culture studies, RAW 264.7 cell lines were pre-incubated with PAR2 antagonist 1h before C-PAF, followed by analysis of intracellular Ca2+ signaling, immunofluorescence for PAR2/NF-κB (p65), (PAR)2/PAFR co-immunoprecipitation, as well as for analysis of PAR2 mRNA expression by real-time PCR. Our results demonstrate that CXCL1 and CXCL2 production, as well as C-PAF-mediated neutrophil recruitment, at least in part, are dependent on PAR2 activation in mouse lungs. Furthermore, we showed that in macrophages, PAR2 antagonism impairs intracellular Ca2+ signaling and C-PAF-induced NF-ҠB (p65) activation. Therefore, together, these results evidence the interaction between PAR2 and PAFR in lung inflammation and in the activation of NF-κB (p65) and intracellular mobilization of Ca2+ in macrophages.