PAR2 promotes M1 macrophage polarization and inflammation via FOXO1 pathway

Abstract

Macrophages polarization plays essential but different roles in most diseases such as atherosclerosis, adipose tissue inflammation, and insulin resistance. Our previous study revealed that protease-activated receptor 2 (PAR2), a G-protein coupled receptor influenced macrophage function, but little is known regarding the regulation of macrophage polarization process and its potential mechanisms. In the present study, bone marrow-derived macrophages (BMDM) isolated from C57/BL6 mice and cultured with L929-conditional medium and murine macrophage cell line RAW264.7 were used to study the function of PAR2 activation in vitro. BMDM was stimulated by the small molecular PAR2 agonist, 2-furoyl-LIGRLO-amide trifluoroacetate salt, followed by transcription factor microarray to screen the significantly activated signaling pathways under PAR2 activation. Western blot analysis, quantitative real-time polymerase chain reaction (qRT-PCR) was used to evaluate the expression of targeted genes and transcription factors. Immunofluorescence was used to observe the subcellular distribution of transcription factors.Our results demonstrated that M1-like polarization was presented by PAR2 agonist treatment with significant upregulation of interleukin-1β, interleukin-6, monocyte chemotactic protein-1, and tumor necrosis factor-α in BMDM and RAW264.7.Microarray identified forkhead box protein O1 (FOXO1) was significantly increased under PAR2 agonist stimulation, which was confirmed by qPCR and Western blot analysis. Immunofluorescence demonstrated that increased FOXO1 accumulated in the nucleus, which is necessary to promote transcription for targeted genes. We further knocked down FOXO1 expression using small interfering RNA, which alleviated PAR2-induced proinflammatory gene expression. The PAR2/FOXO1 pathway mediated stimulation of proinflammatory genes was further confirmed by tryptase, an endogenous ligand of PAR2. In conclusion, this study demonstrated that PAR2 activation-induced M1 polarization and inflammation through the FOXO1-dependent pathway. K E Y W O R D Sforkhead box protein O1, macrophage polarization, protease-activated receptor 2 J Cell Biochem. 2019;120:9799-9809.wileyonlinelibrary.com/journal/jcb

Macrophage TLR4 and PAR2 Signaling: Role in Regulating Vascular Inflammatory Injury and Repair

“…Human monocytes primarily express PAR1, but upon differentiation into macrophages increase expression of PAR2 ( 172 ). PAR2 activation alone is able to skew macrophages into either the M1 or M2 phenotype ( 173 – 175 ). Stimulation of BMDM or RAW cells with the small molecule PAR2 agonist, 2-furoyl-LIGRLO-amide trifluoroacetate salt, skewed MΦ into M1-like cells due to activation of the forkhead box protein O1 (FOXO1) ( 173 ).…”

Protease profile of normal and neoplastic mast cells in the human bone marrow

“…In particular, in the airways, this leads to the formation of functional prerequisites for exacerbation of bronchoconstriction, mucus secretion by mucocytes. PAR2 promotes M1 macrophage polarization and in ammation via FOXO1 pathway 65 67 . In addition, an important effect of tryptase is the activation of secretion by cells of a speci c tissue microenvironment of pro-in ammatory mediators into the intercellular matrix, creating an increased background content of cytokines chemokines 68,69. In light of the previously described effects of tryptase, the progression of allergic reactions is also important 70 .…”

Molecular Biological Peculiarities of the Must Cells Secretome of the Lower Limb Skin in Trophic Ulcers of Various Etiologies

“…Рецепторы PAR-2 локализованы на разных клеточных типах специфичного тканевого микроокружения кожи. Активация этих ре- цепторов на макрофагах приводит к их поляризации к первому типу, усиливая воспаление [12]. Триптаза опосредованно через активацию PAR-2 вовлечена в сигнальные пути развития отека и зуда.…”