Platelet-activating factor and protease-activated receptor 2 cooperate to promote neutrophil recruitment and lung inflammation through nuclear factor-kappa B transactivation.

Although it is well established that platelet-activated receptor (PAF) and protease-activated receptor 2 (PAR2) play a pivotal role in the pathophysiology of lung and airway inflammatory diseases, a role for a PAR2-PAFR cooperation in lung inflammation has not been investigated. Here, we investigated the role of PAR2 in PAF-induced lung inflammation and neutrophil recruitment in lungs of BALB/c mice. Mice were pretreated with the PAR2 antagonist ENMD1068, PAF receptor (PAFR) antagonist WEB2086, or aprotinin prior to intranasal instillation of carbamyl-PAF (C-PAF) or the PAR2 agonist peptide SLIGRL-NH2 (PAR2-AP). Leukocyte infiltration in bronchoalveolar lavage fluid (BALF), C-X-C motif ligand 1 (CXCL)1 and CXCL2 chemokines, myeloperoxidase (MPO), and N-acetyl-glycosaminidase (NAG) levels in BALF, or lung inflammation were evaluated. Intracellular calcium signaling, PAFR/PAR2 physical interaction, and the expression of PAR2 and nuclear factor-kappa B (NF-КB, p65) transcription factor were investigated in RAW 264.7 cells stimulated with C-PAF in the presence or absence of ENMD1068. C-PAF- or PAR2-AP-induced neutrophil recruitment into lungs was inhibited in mice pretreated with ENMD1068 and aprotinin or WEB2086, respectively. PAR2 blockade impaired C-PAF-induced neutrophil rolling and adhesion, lung inflammation, and production of MPO, NAG, CXCL1, and CXCL2 production in lungs of mice. PAFR activation reduced PAR2 expression and physical interaction of PAR2 and PAFR; co-activation is required for PAFR/PAR2 physical interaction. PAR2 blockade impaired C-PAF-induced calcium signal and NF-κB p65 translocation in RAW 264.7 murine macrophages. This study provides the first evidence for a cooperation between PAFR and PAR2 mediating neutrophil recruitment, lung inflammation, and macrophage activation.

LyeTx I-b Peptide Attenuates Tumor Burden and Metastasis in a Mouse 4T1 Breast Cancer Model.

Cationic anticancer peptides have exhibited potent anti-proliferative and anti-inflammatory effects in neoplastic illness conditions. LyeTx I-b is a synthetic peptide derived from Lycosa erythrognatha spider venom that previously showed antibiotic activity in vitro and in vivo. This study focused on the effects of LyeTxI-b on a 4T1 mouse mammary carcinoma model. Mice with a palpable tumor in the left flank were subcutaneously or intratumorally injected with LyeTx I-b (5 mg/kg), which significantly decreased the tumor volume and metastatic nodules. Histological analyses showed a large necrotic area in treated primary tumors compared to the control. LyeTxI-b reduced tumor growth and lung metastasis in the 4T1 mouse mammary carcinoma model with no signs of toxicity in healthy or cancerous mice. The mechanism of action of LyeTx I-b on the 4T1 mouse mammary carcinoma model was evaluated in vitro and is associated with induction of apoptosis and cell proliferation inhibition. Furthermore, LyeTx I-b seems to be an efficient regulator of the 4T1 tumor microenvironment by modulating several cytokines, such as TGF-ß, TNF-α, IL-1ß, IL-6, and IL-10, in primary tumor and lung, spleen, and brain. LyeTx I-b also plays a role in leukocytes rolling and adhesion into spinal cord microcirculation and in the number of circulating leukocytes. These data suggest a potent antineoplastic efficacy ofLyeTx I-b.

Correction to: A role for mast cells and mast cell tryptase in driving neutrophil recruitment in LPS-induced lung inflammation via protease-activated receptor 2 in mice.

The original article can be found online.

A role for mast cells and mast cell tryptase in driving neutrophil recruitment in LPS-induced lung inflammation via protease-activated receptor 2 in mice.

OBJECTIVE: This study aims to investigate the role of protease-activated receptor (PAR) 2 and mast cell (MC) tryptase in LPS-induced lung inflammation and neutrophil recruitment in the lungs of C57BL/6 mice. METHODS: C57BL/6 mice were pretreated with the PAR2 antagonist ENMD-1068, compound 48/80 or aprotinin prior to intranasal instillation of MC tryptase or LPS. Blood leukocytes, C-X-C motif chemokine ligand (CXCL) 1 production leukocytes recovered from bronchoalveolar lavage fluid (BALF), and histopathological analysis of the lung were evaluated 4 h later. Furthermore, we performed experiments to determine intracellular calcium signaling in RAW 264.7 cells stimulated with LPS in the presence or absence of a protease inhibitor cocktail or ENMD-1068 and evaluated PAR2 expression in the lungs of LPS-treated mice. RESULTS: Pharmacological blockade of PAR2 or inhibition of proteases reduced neutrophils recovered in BALF and LPS-induced calcium signaling. PAR2 blockade impaired LPS-induced lung inflammation, PAR2 expression in the lung and CXCL1 release in BALF, and increased circulating blood neutrophils. Intranasal instillation of MC tryptase increased the number of neutrophils recovered in BALF, and MC depletion with compound 48/80 impaired LPS-induced neutrophil migration. CONCLUSION: Our study provides, for the first time, evidence of a pivotal role for MCs and MC tryptase in neutrophil migration, lung inflammation and macrophage activation triggered by LPS, by a mechanism dependent on PAR2 activation.

Inhibition of neutrophil migration and reduction of oxidative stress by ethyl p-coumarate in acute and chronic inflammatory models.

BACKGROUND: It is well known that medicinal plants and their products are relevant candidates for the treatment of inflammatory conditions. Ethyl p-coumarate is a phenylpropanoid that has similar structure to others anti-inflammatory and antioxidant substances. However, these activities have never been tested. PURPOSE: The aim of this study was to investigate the effect of ethyl p-coumarate on inflammatory and oxidative stress parameters. STUDY DESIGN: This is an experimental study to evaluate the anti-inflammatory and antioxidant activities of ethyl p-coumarate in acute and chronic models of inflammation. METHODS: The anti-inflammatory effect of ethyl p-coumarate was evaluated in Swiss mice by carrageenan-induced paw edema model (1%, 50 µl), followed by histological analysis, and edema induced by compound 48/80 (12 µg/paw), histamine (100  µg/paw), serotonin (100 µg/paw) and prostaglandin E2 (3 nmol/paw) in comparison to indomethacin treatment (10 mg/kg, p.o.). In addition, peritonitis was induced by carrageenan (500 µg/cavity) to neutrophil and total leukocytes counting, myeloperoxidase (MPO), interleukin 6 (IL-6) and 8 (IL-8), nitrite (NO2-), glutathione (GSH) and malondialdehyde (MDA) measurements. The arthritis model was induced with Freund's complete adjuvant (id. 0.1 ml) in female Wistar rats, with measurement of joint diameter and X-ray. Changes in gastric tissue of Swiss mice were analyzed in comparison to indomethacin (20  mg/kg, p.o.). RESULTS: After treatment with ethyl p-coumarate, the animals had no apparent toxic effects, and significantly inhibited paw edema induced by edematogenic agents, neutrophil (p < 0.001) and total leukocyte (p < 0.001) migration, MPO (p < 0.01), IL-6 (p < 0.05) and IL-8 (p < 0.5), MDA (p < 0.5), GSH (p < 0.5), NO2- (p < 0.001), joint thickness and bones changes. Furthermore, were not observed significant formation of gastric lesions. CONCLUSION: Taken together, these results suggest that ethyl p-coumarate exhibits anti-inflammatory activity through the inhibition of inflammatory mediators and leukocyte migration without causing gastric lesions.

Photobiomodulation reduces neutrophil migration and oxidative stress in mice with carrageenan-induced peritonitis.

The purpose of the study is to investigate the effects of two doses of photobiomodulation (PBM) on inflammatory parameters including cell migration and oxidative stress in carrageenan-induced peritonitis models. Twenty-eight mice were divided into four groups: saline; untreated carrageenan (Cg; inflammation induced); and PMB treatment groups L1 and L5 (inflammation induced with carrageenan followed by laser irradiation at 1 and 5 J/cm2, respectively). After 30 min of inducing inflammation, laser irradiation was administered every hour, for 4 h. Peritoneal fluid was collected for analyses. The total leukocyte number in the peritoneal fluid in L1 (4.33 ± 2.34) and L5 (4.95 ± 2.86) after PBM was lower than that in Cg (10.93 ± 5.15 cells/ml). The average differential count of neutrophils in the Cg was 9.46 ± 4.31 cells/ml, which was higher than that in L1 (3.7 ± 2.08) and L5 (4.94 ± 2.57). Myeloperoxidase activity was also lower in L1 (1.89 ± 0.43) and L5 (4.84 ± 2.62) than in Cg (22.92 ± 4.52 UMPO/ml). Malondialdehyde content was lower in L1 (137.5 ± 12.33) and L5 (169.6 ± 22.77) than in Cg (345.7 ± 65.67 nmol/ml). Glutathione peroxidase concentration was significantly higher in L1 (155.2 ± 12.43) and L5 (145.9 ± 9.585) than in Cg (79.75 ± 9.567 µ/ml). Nitrite concentration was lower in L1 (0.3317 µM ± 0.0669) and L5 (0.2429 µM ± 0.0232) than in Cg (0.8380 µM ± 0.01615). Laser irradiation at 1 and 5 J/cm2 reversed the inflammation (as indicated by neutrophil infiltration and oxidative stress).

H2S is a key antisecretory molecule against cholera toxin-induced diarrhoea in mice: Evidence for non-involvement of the AC/cAMP/PKA pathway and AMPK.

Hydrogen sulphide (H2S) is a gasotransmitter that participates in various physiological and pathophysiological processes within the gastrointestinal tract. We studied the effects and possible mechanism of action of H2S in secretory diarrhoea caused by cholera toxin (CT). The possible mechanisms of action of H2S were investigated using an intestinal fluid secretion model in isolated intestinal loops on anaesthetized mice treated with CT. NaHS and Lawesson's reagent and l-cysteine showed antisecretory activity through reduction of intestinal fluid secretion and loss of Cl- induced by CT. Pretreatment with an inhibitor of cystathionine-γ-lyase (CSE), dl-propargylglycine (PAG), reversed the effect of l-cysteine and caused severe intestinal secretion. Co-treatment with PAG and a submaximal dose of CT increased intestinal fluid secretion, thus supporting the role of H2S in the pathophysiology of cholera. CT increased the expression of CSE and the production of H2S. Pretreatment with PAG did not reverse the effect of SQ 22536 (an AC inhibitor), bupivacaine (inhibitor of cAMP production), KT-5720 (a PKA inhibitor), and AICAR (an AMPK activator). The treatment with Forskolin does not reverse the effects of the H2S donors. Co-treatment with either NaHS or Lawesson's reagent and dorsomorphin (an AMPK inhibitor) did not reverse the effect of the H2S donors. H2S has antisecretory activity and is an essential molecule for protection against the intestinal secretion induced by CT. Thus, H2S donor drugs are promising candidates for cholera therapy. However, more studies are needed to elucidate the possible mechanism of action.

Evaluation of anti-inflammatory potential of aqueous extract and polysaccharide fraction of Thuja occidentalis Linn. in mice.

Inflammation is a protective reaction of the microcirculation. However, sustained inflammation can lead to undesired effects. Thuja occidentalis Linn has many pharmacological properties but has no anti-inflammatory activity described. Thus, this study aims evaluating the anti-inflammatory activity of the aqueous extract (AE) and the polysaccharide fraction (PLS) of T. occidentalis L. in mice. The results of our evaluations in various experimental models indicated that AE and PLS (3, 10, and 30mg/kg, i.p.) reduced (p˂0.05) paw edema induced by carrageenan, dextran sulfate (DEX), compound 48/80, serotonin (5-HT), bradykinin (BK), histamine (HIST), and prostaglandin E2 (PGE2). Furthermore, it inhibited neutrophils recruitment; decreased MPO activity, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels, vascular permeability, nitrite concentration, and MDA concentration; and maintained the GSH levels in the peritoneal exudate. The AE and PLS reduced neutrophil infiltration and cyclooxygenase (COX)-2 and inducible nitric oxide synthase (iNOS) immunostaining in paw tissue. Treatment with the AE and PLS (300mg/kg) did not induce gastric toxicity. In conclusion, these results show that the AE and PLS reduced the inflammatory response by inhibiting vascular and cellular events, inhibiting pro-inflammatory cytokine production, and reducing oxidative stress. Furthermore, they did not induce gastric toxicity at high doses.

Riparin A, a compound from Aniba riparia, attenuate the inflammatory response by modulation of neutrophil migration.

Inflammation is a local tissue response to attacks characterized by vascular and cellular events, including intense oxidative stress. Riparin A, a compound obtained from Aniba riparia, has been shown to have antioxidant activity and cytotoxicity in vitro. This study was aimed at evaluating the anti-inflammatory effect of riparin A against acute inflammation. The results of our evaluations in various experimental models indicated that riparin A reduced paw edema induced by carrageenan, compound 48/80, histamine, and serotonin. Furthermore, it decreased leukocyte and neutrophil counts, myeloperoxidase activity, thiobarbituric acid reactive substance (TBARS) levels, and cytokine (tumor necrosis factor-α and interleukin-1ß) levels increased by carrageenan-induced peritonitis, and reversed glutathione levels. Riparin A also reduced carrageenan-induced adhesion and rolling of leukocytes on epithelial cells and did not produce gastric-damage as compared with indomethacin. In conclusion, the data show that riparin A reduces inflammatory response by inhibiting vascular and cellular events, modulating neutrophil migration, inhibiting proinflammatory cytokine production, and reducing oxidative stress.

Carvacryl acetate, a derivative of carvacrol, reduces nociceptive and inflammatory response in mice.

AIMS: The present study aimed to investigate the potential anti-inflammatory and anti-nociceptive effects of carvacryl acetate, a derivative of carvacrol, in mice. MAIN METHODS: The anti-inflammatory activity was evaluated using various phlogistic agents that induce paw edema, peritonitis model, myeloperoxidase (MPO) activity, pro and anti-inflammatory cytokine levels. Evaluation of antinociceptive activity was conducted through acetic acid-induced writhing, hot plate test, formalin test, capsaicin and glutamate tests, as well as evaluation of motor performance on rotarod test. KEY FINDINGS: Pretreatment of mice with carvacryl acetate (75 mg/kg) significantly reduced carrageenan-induced paw edema (P<0.05) when compared to vehicle-treated group. Likewise, carvacryl acetate (75 mg/kg) strongly inhibited edema induced by histamine, serotonin, prostaglandin E2 and compound 48/80. In the peritonitis model, carvacryl acetate significantly decreased total and differential leukocyte counts, and reduced levels of myeloperoxidase and interleukin-1 beta (IL-1ß) in the peritoneal exudate. The levels of IL-10, an anti-inflammatory cytokine, were enhanced by carvacryl acetate. Pretreatment with carvacryl acetate also decreased the number of acetic acid-induced writhing, increased the latency time of the animals on the hot plate and decreased paw licking time in the formalin, capsaicin and glutamate tests. The pretreatment with naloxone did not reverse the carvacryl acetate-mediated nociceptive effect. SIGNIFICANCE: In conclusion, the current study demonstrated that carvacryl acetate exhibited anti-inflammatory activity in mice by reducing inflammatory mediators, neutrophil migration and cytokine concentration, and anti-nociceptive activity due to the involvement of capsaicin and glutamate pathways.