Expression Of Formyl Peptide Receptor Research Articles

Cytomegalovirus durably primes neutrophil oxidative burst.

Cytomegalovirus (CMV) is a ubiquitous herpes virus that infects most humans, thereafter persisting lifelong in tissues of the host. It is a known pathogen in immunosuppressed patients, but its impact on immunocompetent hosts remains less understood. Recent data have shown that CMV leaves a significant and long-lasting imprint in host immunity that may confer some protection against subsequent bacterial infection. Such innate immune activation may come at a cost, however, with potential to cause immunopathology. Neutrophils are central to many models of immunopathology, and while acute CMV infection is known to influence neutrophil biology, the impact of chronic CMV infection on neutrophil function remains unreported. Using our murine model of CMV infection and latency, we show that chronic CMV causes persistent enhancement of neutrophil oxidative burst well after resolution of acute infection. Moreover, this in vivo priming of marrow neutrophils is associated with enhanced formyl peptide receptor expression, and ultimately constitutive c-Jun N-terminal kinase phosphorylation and enhanced CD14 expression in/on circulating neutrophils. Finally, we show that neutrophil priming is dependent on viral load, suggesting that naturally infected human hosts will show variability in CMV-related neutrophil priming. Altogether, these findings represent a previously unrecognized and potentially important impact of chronic CMV infection on neutrophil responsiveness in immunocompetent hosts.

Peptides Derived from Soybean β-Conglycinin Induce the Migration of Human Peripheral Polymorphonuclear Leukocytes.

Food-derived peptides have various biological activities. When food proteins are ingested orally, they are digested into peptides by endogenous digestive enzymes and absorbed by the immune cell-rich intestinal tract. However, little is known about the effects of food-derived peptides on the motility of human immune cells. In this study, we aimed to understand the effects of peptides derived from a soybean protein β-conglycinin on the motility of human peripheral polymorphonuclear leukocytes. We illustrated that MITL and MITLAIPVNKPGR, produced by digestion using in-vivo enzymes (trypsin and pancreatic elastase) of β-conglycinin, induces the migration of dibutyryl cAMP (Bt2 cAMP)-differentiated human promyelocytic leukemia 60 (HL-60) cells and human polymorphonuclear leukocytes in a dose- and time-dependent manner. This migration was more pronounced in Bt2 cAMP-differentiated HL-60 cells; mRNA expression of formyl peptide receptor (FPR) 1 increased significantly than in all-trans-retinoic acid (ATRA)-differentiated HL-60 cells. This migration was inhibited by tert-butoxycarbonyl (Boc)-MLP, an inhibitor of FPR, and by pretreatment with pertussis toxin (PTX). However, the effect was weak when treated with WRW4, a selective inhibitor of the FPR2. We then demonstrated that MITLAIPVNKPGR induced intracellular calcium responses in human polymorphonuclear leukocytes and Bt2 cAMP-HL60 cells. Furthermore, pre-treatment by fMLP desensitized the calcium response of MITLAIPVNKPGR in these cells. From the above, MITLAIPVNKPGR and MITL derived from soybean β-conglycinin induced polymorphonuclear leukocyte migration via the FPR1-dependent mechanism. We found chemotactic peptides to human polymorphonuclear leukocytes, which are the endogenous enzyme digests of soybean protein.

Resolvin D1 attenuates acid-induced DNA damage in esophageal epithelial cells and rat models of acid reflux

The role of resolvin D1 (RvD1) in gastroesophageal reflux disease (GERD) remains largely unknown. Here, we investigated the potential role of RvD1 in acid-induced DNA damage in esophageal epithelial cells, patients with refractory GERD and a rat model of acid reflux. Weak acid exposure induced longer comet tails, reactive oxygen species (ROS) generation, oxidative DNA damage and DNA double-strand breaks (DSBs) in cells and RvD1 (0.1 μM) blocked all these effects. Mechanistic analyses showed that apart from ROS-reducing effects, RvD1 possessed a strong capacity to promote DNA damage repair, augmenting cell cycle checkpoint activity and DSB repair by modulating phosphatase and tensin homolog (PTEN) in cells. We also detected the surface expression of formyl peptide receptor 2 (FPR2), a receptor for RvD1, in the esophageal epithelial cells, and inhibition of FPR2 abrogated the protective effects of RvD1 on cells. Furthermore, a positive correlation between RvD1 and PTEN was observed predominantly in the esophageal epithelium from patients with refractory GERD (r = 0.67, P < 0.05). Additionally, RvD1 administration upregulated PTEN, suppressed DNA DSBs and alleviated microscopic damage in the rat model of gastric reflux. FPR2 gene silencing abolished the therapeutic effects of RvD1 on the rat model. Taken together, RvD1 binding to FPR2 protects the esophageal epithelium from acid reflux-induced DNA damage via a mechanism involving the inhibition of ROS production and facilitation of DSB repair. These findings support RvD1 as a promising approach that may be valuable for the treatment of GERD.

Formyl peptide receptor-2 is upregulated in the blood and placenta of patients with gestational diabetes mellitus.

AimTo investigate the expression of formyl peptide receptor 2 (FPR2) in maternal blood, umbilical blood, and placenta of patients with gestational diabetes mellitus (GDM), and to analyze the changes of other pro‐inflammatory cytokines in blood, including interleukin 33 (IL‐33), IL‐1β, tumor necrosis factor alpha (TNF‐α), and C‐reactive protein (CRP), so as to reveal the pathogenesis of GDM.MethodsFPR2, IL‐33, IL‐1β, T TNF‐α, and CRP in maternal blood and umbilical cord blood of 50 pregnant women with GDM and 30 normal pregnant women were analyzed by ELISA method to explore the correlation between inflammatory factors and blood glucose. The expression of FPR2 in placental tissues was analyzed by PCR and immunohistochemistry.ResultsThe expression of FPR2 in maternal blood of gestational diabetes patients was significantly higher than that of normal pregnant women, and other inflammatory factors IL‐33 and IL‐1β in maternal blood were also significantly increased. The expression of FPR2 in umbilical cord blood of gestational diabetes was higher than that of normal pregnant women, but the difference was not significant. Other inflammatory factors IL‐33, IL‐1β, and CRP in umbilical cord blood were also significantly increased. The expression of FPR2mRNA and protein in placental tissues of gestational diabetes was significantly higher than that of normal pregnant women.ConclusionsThe level of FPR2, IL‐33, and IL‐1β in maternal blood was related to the pathogenesis of GDM and these inflammatory factors could be used as special candidate direction of marks for the prevention, clinical treatment and drug design of GDM, laying a new theoretical foundation for the treatment of GDM.

Analysis of the vomeronasal organ transcriptome reveals variable gene expression depending on age and function in rabbits

The vomeronasal organ (VNO) is a chemosensory organ specialized in pheromone detection that shows a broad morphofunctional and genomic diversity among mammals. However, its expression patterns have only been well-characterized in mice. Here, we provide the first comprehensive RNA sequencing study of the rabbit VNO across gender and sexual maturation stages. We characterized the VNO transcriptome, updating the number and expression of the two main vomeronasal receptor families, including 128 V1Rs and 67 V2Rs. Further, we defined the expression of formyl-peptide receptor and transient receptor potential channel families, both known to have specific roles in the VNO. Several sex hormone-related pathways were consistently enriched in the VNO, highlighting the relevance of this organ in reproduction. Moreover, whereas juvenile and adult VNOs showed significant transcriptome differences, male and female did not. Overall, these results contribute to understand the genomic basis of behavioural responses mediated by the VNO in a non-rodent model.

Acute cigarette smoke-induced eQTL affects formyl peptide receptor expression and lung function.

Cigarette smoking is one of the most prevalent causes of preventable deaths worldwide, leading to chronic diseases, including chronic obstructive pulmonary disease (COPD). Cigarette smoke is known to induce significant transcriptional modifications throughout the respiratory tract. However, it is largely unknown how genetic profiles influence the smoking-related transcriptional changes and how changes in gene expression translate into altered alveolar epithelial repair responses. We performed a candidate-based acute cigarette smoke-induced eQTL study, investigating the association between SNP and differential gene expression of FPR family members in bronchial epithelial cells isolated 24 h after smoking and after 48 h without smoking. The effects FPR1 on lung epithelial integrity and repair upon damage in the presence and absence of cigarette smoke were studied in CRISPR-Cas9-generated lung epithelial knockout cells. One significant (FDR < 0.05) inducible eQTL (rs3212855) was identified that induced a >2-fold change in gene expression. The minor allele of rs3212855 was associated with significantly higher gene expression of FPR1, FPR2 and FPR3 upon smoking. Importantly, the minor allele of rs3212855 was also associated with lower lung function. Alveolar epithelial FPR1 knockout cells were protected against CSE-induced reduction in repair capacity upon wounding. We identified a novel smoking-related inducible eQTL that is associated with a smoke-induced increase in the expression of FPR1, FPR2 and FPR3, and with lowered lung function. in vitro FPR1 down-regulation protects against smoke-induced reduction in lung epithelial repair.

Anti-inflammatory effect of traditional Chinese medicine preparation Penyanling on pelvic inflammatory disease

Ethnopharmacological relevancePenyanling is made up of Smilacis Glabrae Rhizoma (SG, from Smilar glabra Roxb.), Angelicae Sinensis Radix (AS, from Angelica sinensis (Oliv.) Diels), Salviae Miltiorrhizae Radix et Rhizoma (SM, from Salvia miltiorrhiza Bunge), Sargentodoxae Caulis (SC, from Sargentodoxa cuneata (Oliv.) Rehd.et Wils.), Linderae Radix (LR, from Lindera aggregata (Sims) Kosterm.), Paeoniae Radix Rubra (PR, from Paeonia lactiflora Pall.), Sparganii Rhizoma (SR, from Sparganium stoloniferum (Graebn.) Buch.-Ham.), Corydalis Rhizoma (CoR, from Corydalis yanhusuo W. T. Wang), Cyperi Rhizoma (CyR, from Cyperus rotundus Linn.), Glycyrrhizae Radix et Rhizoma (GR, from Glycyrrhiza uralensis Fisch.), and Patrinia Scabiosaefolia (PS, from Patrinia scabiosaefolia Fisch. ex Trev.) recorded in Chinese Pharmacopoeia. It has been used on pelvic inflammatory disease (PID) for more than twenty years.Aim of the study: This study was carried out to illustrate its pharmacological action and clarify its substantial composition. Materials and methodsThe anti-inflammatory effects of Penyanling were studied on a PID rat model and a lipopolysaccharides (LPS)-stimulated THP-1 cell line. Histological changes and levels of inflammatory factors in the uterine tube of the PID rat were examined. Levels of nuclear factor-kappa B (NF-κB) in the nuclear of THP-1 cells and NF-κB, IκB-α, and FPR2 in the cytoplasm were tested by Western blot analysis. Substances within Penyanling were scanned with liquid chromatography-quadrupole-time of flight-mass spectrometry (LC-Q-TOF-MS). The contents of total flavonoids, phenolics, and saponins were quantified. ResultsThe anti-inflammatory effects of Penyanling were observed on PID rats, such as suppressing the infiltrations of lymphocytes and neutrophils in the uterine tube, decreasing the release of interleukin (IL)-1β, IL-6, IL-8, and monocyte chemotactic protein (MCP)-1, and promoting the production of lipoxin A4 (LXA4). On the other hand, Penyanling regulated the activity of NF-κB signal pathway on the LPS-stimulated THP-1 cell line, which suggested the potential mechanism of its anti-inflammatory effect. Besides, it could promote the expression of formyl peptide receptor 2 (FPR2), which suggested its effect on enhancing the resolution of inflammation. Seventy-six substances were identified by their accurate molecular weights, mass fragment patterns, retention times, and standards if available. Most of these substances were flavonoids, phenolics, saponins, and alkaloids. The contents of total flavonoids, phenolics, and saponins within Penyanling were 0.186, 1.371, and 4.321 mg/mL, respectively. ConclusionPenyanling showed an anti-inflammatory effect on PID, and its potential mechanism involved suppressing NF-κB signal pathway and promoting the resolution of inflammation. The main substances within it were flavonoids, phenolics, saponins, and alkaloids.

Data describing expression of formyl peptide receptor 2 in human articular chondrocytes.

The formyl peptide receptor 2 (FPR2) belongs to the family of seven-transmembrane G protein-coupled receptors (GPCR) and are expressed by many different cells but mainly studied in immune cells. FPR2 is involved in host defense against bacterial infections and clearance of damaged cells through the oxidative burst and chemotaxis of neutrophils. In addition, FPR2 has also been implicated as an immunomodulator in sterile inflammations, e.g. inflammatory joint diseases. Here we present data regarding FPR2 expression in human articular chondrocytes, isolated from healthy individuals and osteoarthritic patients, on both mRNA and protein level using qPCR and Imagestream flow cytometry. We also present data after receptor stimulation and monitoring of production of nitric oxide, reactive oxygen species, IL-6, IL-8 and MMP-3. The presented data show that human articular chondrocytes from patients with osteoarthritis as well as from healthy individuals express FPR2 both at mRNA and protein level. The biological relevance of FPR2 expression in chondrocytes needs to be further investigated.

Contribution of FPR and TLR9 to hypoxia-induced chemoresistance of ovarian cancer cells.

Background/purposeThe aim of this study was to investigate the role and mechanisms of the formyl peptide receptor (FPR) and the toll-like receptor 9 (TLR9) in hypoxia-induced chemoresistance of human ovarian cancer cells.Materials and methodsSKOV3 cells were exposed to hypoxia for 24 hours, the supernatant was collected to stimulate normoxia-cultured SKOV3, and the inhibition rate of cell growth was detected with CCK8 test. The agonist of TLR9 CpG ODN and the agonist of FPR fMLF were applied to investigate the chemosensitivity of SKOV3 cells to cisplatin. The cells were also treated with FPR antagonist t-Boc or TLR9 antagonist CQ. Western blot was applied to detect protein levels of FPR, TLR9, MRP, P-gp, p53 and Beclin-1. Immunofluorescence staining was applied to observe the distribution of TLR9 in SKOV3 cells.ResultsHypoxia exposure reduced the inhibition rate of cisplatin on SKOV3 cells. WB showed that FPR and TLR9 were expressed in human ovarian cancer tissues and SKOV3 cells, and the levels were increased with longer hypoxia time. After SKOV3 was stimulated with fMLF or ODN2006, cisplatin-induced inhibition rate was significantly decreased. tBoc and CQ significantly attenuated hypoxia supernatant-induced chemoresistance of SKOV3 cells. Hypoxia supernatants significantly increased MRP, P-gp, p53 and Beclin-1 proteins in SKOV3 cells, which were significantly reduced by tBoc.ConclusionHypoxia upregulates the expression of FPR and TLR9, and promotes the release of ligands for both receptors in human ovarian cancer cell line. FPR and TLR9 may be noval targets for chemosensitizing to ovarian cancer cells.

Mitochondrial Damage-Associated Molecular Patterns of Injured Axons Induce Outgrowth of Schwann Cell Processes.

Activated Schwann cells put out cytoplasmic processes that play a significant role in cell migration and axon regeneration. Following nerve injury, axonal mitochondria release mitochondrial damage-associated molecular patterns (mtDAMPs), including formylated peptides and mitochondrial DNA (mtDNA). We hypothesize that mtDAMPs released from disintegrated axonal mitochondria may stimulate Schwann cells to put out cytoplasmic processes. We investigated RT4-D6P2T schwannoma cells (RT4) in vitro treated with N-formyl-L-methionyl-L-leucyl-phenylalanine (fMLP) or cytosine-phospho-guanine oligodeoxynucleotide (CpG ODN) for 1, 6 and 24 h. We also used immunohistochemical detection to monitor the expression of formylpeptide receptor 2 (FPR2) and toll-like receptor 9 (TLR9), the canonical receptors for formylated peptides and mtDNA, in RT4 cells and Schwann cells distal to nerve injury. RT4 cells treated with fMLP put out a significantly higher number of cytoplasmic processes compared to control cells. Preincubation with PBP10, a selective inhibitor of FPR2 resulted in a significant reduction of cytoplasmic process outgrowth. A significantly higher number of cytoplasmic processes was also found after treatment with CpG ODN compared to control cells. Pretreatment with inhibitory ODN (INH ODN) resulted in a reduced number of cytoplasmic processes after subsequent treatment with CpG ODN only at 6 h, but 1 and 24 h treatment with CpG ODN demonstrated an additive effect of INH ODN on the development of cytoplasmic processes. Immunohistochemistry and western blot detected increased levels of tyrosine-phosphorylated paxillin in RT4 cells associated with cytoplasmic process outgrowth after fMLP or CpG ODN treatment. We found increased immunofluorescence of FPR2 and TLR9 in RT4 cells treated with fMLP or CpG ODN as well as in activated Schwann cells distal to the nerve injury. In addition, activated Schwann cells displayed FPR2 and TLR9 immunostaining close to GAP43-immunopositive regenerated axons and their growth cones after nerve crush. Increased FPR2 and TLR9 immunoreaction was associated with activation of p38 and NFkB, respectively. Surprisingly, the growth cones displayed also FPR2 and TLR9 immunostaining. These results present the first evidence that potential mtDAMPs may play a key role in the induction of Schwann cell processes. This reaction of Schwann cells can be mediated via FPR2 and TLR9 that are canonical receptors for formylated peptides and mtDNA. The possible role for FPR2 and TLR9 in growth cones is also discussed.

Expression and Functions of Formyl Peptide Receptor 1 in Drug-Resistant Bladder Cancer.

Objective:To explore the correlation of formyl peptide receptor 1 expression with drug resistance and the functions of formyl peptide receptor 1 in drug-resistant bladder cancer.Methods:Expression of formyl peptide receptor 1 in T24 and T24/DDP cisplatin-resistant bladder cancer cell lines was tested by quantitative real-time Polymerase Chain Reaction and Western blotting. After incubation of T24/DDP with N-formyl-Met-Leu-Phe, the phosphor proteins were tested by Western blot analysis. We characterized the functions of formyl peptide receptor 1 in T24/DDP cells by assessing proliferation, migration, and changes of cell cycles.Results:Formyl peptide receptor 1 was expressed in both T24 and T24/DDP, and it was overexpressed in T24/DDP compared with T24. Formyl peptide receptor 1 activation promoted the expression of the messenger RNA of resistance-related proteins, such as multidrug resistance-associated protein 1 (MRP1) and lung resistance-related protein (LRP). The expression of 4 signal pathway proteins were upregulated: signal transducer and activator of transcription 3, Janus kinase 2, extracellular regulated protein kinases, and protein kinase B, while the expression of phosphatidylinositol 3-kinase was observed to be downregulated in drug-resistant bladder cancer cells. Formyl peptide receptor 1 activation also improved the expression of phospho-signal transducer and activator of transcription 3 and phospho-extracellular regulated protein kinases 1/2 and promoted the proliferation and migration of T24/DDP cells. In addition, formyl peptide receptor 1 inhibition led to the change in the cell cycle in T24/DDP.Conclusions:The overexpression of formyl peptide receptor 1 may be related to drug-resistant bladder cancer and promotes the deterioration of drug-resistant bladder cancer.

Formyl peptide receptor 2 mediated chemotherapeutics drug resistance in colon cancer cells.

To determine the expression of formyl peptide receptor 2 (FPRL2) and its drug resistance role in cancer colon cells, and its underlying mechanisms. The expression of FPRL2 and its legend (F2L) in colon cancer tissues or cancer cells was determined by immunohistochemistry assay and Real-time polymerase chain reaction (PCR), respectively. Chemosensitivity of 5-Fu and MMC in colon cancer cells were tested by cell counting kit-8 (CCK-8) method. Expression of p-ERK was determined by Western blot assay. The expression of FPRL2 and its legend was significantly higher in resistant colon cancer tissues than those in non-resistant colon cancer tissues. The FPRL2 positive cells were two-thirds in tested cell lines. All of cells were F2L positive. The IC50 (inhibitory concentration 50) by 5-Fu and MMC was significantly higher in FPRL2 positive cells than those negative cells. The expression of p-AKT was markedly increased in FPRL2 positive cells. Pretreatment with AKT inhibitor enhanced the drug-sensitivity of these cells to 5-Fu and MMC. The FPRL2 played a significant role in colon cancer drug resistance and this effect was through AKT pathway.

The Expression of Formyl Peptide Receptor 1 is Correlated with Tumor Invasion of Human Colorectal Cancer

Formyl peptide receptors (FPRs) are G protein-coupled chemoattractant receptors expressed mainly in phagocytic leukocytes. High expression of FPRs has also been detected in several cancers but the functions of FPR1 in tumor invasion and metastasis is poorly understood. In this study, we investigated the expression of FPRs in primary human colorectal cancer (CRC) and analyzed the association of FPRs expression with clinicopathological parameters. The levels of FPRs mRNA, especially those of FPR1, were significantly higher in colorectal tumors than in distant normal tissues and adjacent non-tumor tissues. FPR1 mRNA expression was also associated with tumor serosal infiltration. FPR1 protein expression was both in the colorectal epitheliums and tumor infiltrating neutrophils/macrophages. Furthermore, the functions of FPR1 in tumor invasion and tissue repair were investigated using the CRC cell lines SW480 and HT29. Higher cell surface expression of FPR1 is associated with significantly increased migration in SW480 cells compared with HT29 cells that have less FPR1 membrane expression. Finally, genetic deletion of fpr1 increased the survival rate of the resulting knockout mice compared with wild type littermates in a mouse model of colitis-associated colorectal cancer. Our data demonstrate that FPR1 may play an important role in tumor cell invasion in CRC patients.

Effect of hypothermia on splenic leukocyte modulation and survival duration in severely septic rats

BackgroundTherapeutic hypothermia (HT) in severe septic shock is associated with prolonged survival. We hypothesized that moderate HT would prolong survival and modulate the inflammatory response in rats with septic shock by exerting its therapeutic effect on splenic leukocytes. Materials and methodsSevere septic shock was created in rats by cecal ligation and incision (CLI). One hour after CLI or laparotomy, rats were randomized to sham, normothermia (NT), or 4 h of HT followed by 2 h of rewarming. HT (31 ± 1°C) was induced using a cooling blanket and monitored via a rectal temperature probe. ResultsSurvival duration was 2.78 ± 1.0 h in NT rats and 8.33 ± 0.32 h in HT rats (n = 8/group, P < 0.0001). In separate groups, 3 h after CLI, the spleen weight was significantly smaller in NT rats (769 ± 100 mg) than in HT rats (947 ± 157 mg, P = 0.04). Fluorescent immunostaining of formyl peptide receptors on leukocytes in spleen tissue showed considerably higher formyl peptide receptor expression in HT rats than in NT rats. Significantly elevated proinflammatory cytokines and myeloperoxidase enzyme in plasma were found in NT rats compared with HT rats. Anti-inflammatory cytokine, interleukin-10, was significantly higher in HT rats. Both proinflammatory cytokines and plasma myeloperoxidase were significantly reduced in splenectomized NT rats. ConclusionsModerate hypothermic therapy significantly prolongs the survival duration of rats with severe septic shock. HT dampens the inflammatory response during septic shock by modulating the spleen to an anti-inflammatory mode and preventing the spleen from releasing activated splenic leukocytes into the blood.

Follicular dendritic cells modulated by formyl peptide receptor ligands are capable of enhancing the germinal center reaction in mucosal immune system

Abstract In various vaccination protocols, induction of germinal center (GC) is one of the critical factors involved in successful immune induction because GC formation results in the production of high affinity antibodies via B cell clonal expansion, somatic hypermutation, and affinity-based B cell selection. Although GC formation in Peyer’s patch (PP) is slowly and spontaneously induced by intestinal microbiota, it is unknown whether follicular dendritic cells (FDCs) in PP directly contribute to the antigen-specific immune response induction. In this study, we found the expression of formyl peptide receptor (FPR-2), a receptor for cathelicidin LL-37 which could play a role as an adjuvant in mucosal vaccine, on FDCs. The interaction between LL-37 and FPR-2 on FDCs promotes the expression of CXCL13 which is closely associated with the GC formation. Based on these findings, we conclude that the adjuvant role of LL-37 in mucosal immune induction is closely associated with the enhancement of GC reaction in PPs by FDC stimulation.

Abstract 213: Deletion of 12/15 Lipoxygenase and Fatty Acids Interaction Alters Leukocyte Kinetics Leading to Improved Healing Following Myocardial Infarction

The 12/15 lipoxygenase (LOX) enzyme catalyzes oxygenation of fatty acids to form lipid mediators leading to non-resolving inflammation. However, how 12/15LOX interacts with PUFA (polyunsaturated fatty acids) in post-myocardial infarction (MI) cardiac healing is unclear. Here we assessed the role of 12/15LOX in post-MI cardiac remodeling in a PUFA (10% w/w, 22 Kcal) enriched environment. C57BL/6J wild-type (WT) and 12/15LOX null (12/15LOX –/– ) male mice of 8-12-weeks age were fed PUFA-enriched diet for 1 month and subjected to permanent coronary artery ligation. Post-MI mice were monitored for day (d)1 or d5 along with standard diet (SD)-fed MI controls. No-MI surgery mice served as d0 controls. 12/15LOX and PUFA+12/15LOX -/- mice reduced infarcted wall thinning with decreased lung edema index at d5 post-MI (all p&lt;0.05) despite comparable infarct area (49-52%). PUFA+WT and 12/15LOX -/- mice showed improved ejection fraction with reduced left ventricle (LV) hypertrophy index than SD+WT at d5 post-MI (p&lt;0.05). The neutrophil density was decreased in both PUFA+WT and 12/15LOX -/- mice at d1 post-MI (P&lt;0.05). The neutrophil clearance was rapid in 12/15LOX -/- alone and PUFA+12/15LOX -/- mice, with increases in expression of formyl peptide receptor 2 at d5 post-MI. The macrophage density was decreased in PUFA+WT and 12/15LOX -/- mice compared with their respective SD+WT control at d5 post-MI. PUFA+12/15LOX -/- mice displayed an increased expression of ccl2 (all p&lt;0.05) in the infarct area. 12/15LOX deletion stimulated 5-LOX and heme oxygenase-1 in PUFA+12/15LOX -/- mice with increased levels of 9-and 13-hydroxyoctadecadienoic acid indicating improved resolution of inflammation at d5 post-MI. The PUFA+12/15LOX -/- mice displayed higher expression of proresolving macrophages ym-1, mrc-1 and arg-1 compared with SD+12/15LOX -/- mice at d5 post-MI (all p&lt;0.05). Further, 12/15LOX -/- mice with or without PUFA showed reduced collagen deposition at d5 post-MI compared to SD+WT. In conclusion, deletion of 12/15LOX and short-term PUFA exposure attenuated cardiac remodeling via shortening the pro-inflammatory window, thus leading to an improved resolution of inflammation and LV function post-MI.

Flow cytometric analysis with a fluorescently labeled formyl peptide receptor ligand as a new method to study the pharmacological profile of the histamine H2 receptor.

The histamine H2 receptor (H2R) is a Gs protein-coupled receptor. Its activation leads to increases in the second messenger adenosine-3',5'-cyclic monophosphate (cAMP). Presently, several systems are established to characterize the pharmacological profile of the H2R, mostly requiring radioactive material, animal models, or human blood cells. This prompted us to establish a flow cytometric analysis with a fluorescently labeled formyl peptide receptor (FPR) ligand in order to investigate the H2R functionally and pharmacologically. First, we stimulated U937 promonocytes, which mature in a cAMP-dependent fashion upon H2R activation, with histamine (HA) or selective H2R agonists and measured increases in cAMP concentrations by mass spectrometry. Next, indicative for the maturation of U937 promonocytes, we assessed the FPR expression upon incubation with HA or H2R agonists. FPR expression was measured either indirectly by formyl peptide-induced changes in intracellular calcium concentrations ([Ca(2+)]i) or directly with the fluorescein-labeled FPR ligand fNleLFNleYK-Fl. HA and H2R agonists concentration-dependently induced FPR expression, and potencies and efficacies of fMLP-induced increases in [Ca(2+)]i and FPR density correlated linearly. Accordingly, flow cytometric analysis of FPR expression constitutes a simple, inexpensive, sensitive, and reliable method to characterize the H2R pharmacologically. Furthermore, we evaluated FPR expression at the mRNA level. Generally, quantitative real-time polymerase chain reaction confirmed functional data. Additionally, our study supports the concept of functional selectivity of the H2R, since we observed dissociations in the efficacies of HA and H2R agonists in cAMP accumulation and FPR expression.

Splenic leukocytes mediate the hyperglycemic exacerbation of myocardial infarct size in mice.

Acute hyperglycemia during acute myocardial infarction is associated with worse myocardial injury and increased mortality. Using a mouse model of myocardial ischemia/reperfusion injury, we tested the hypothesis that acute hyperglycemia activates splenic leukocytes and subsequently exacerbates myocardial infarct size. We then examined whether the adverse effects of hyperglycemia could be attenuated by a potent anti-inflammatory agent (an agonist of the adenosine A2A receptor) administered immediately prior to reperfusion. C57BL6 (WT) mice underwent 30-min LAD occlusion and 60-min reperfusion with or without prior splenectomy. Acute hyperglycemia before ischemia increased myocardial infarct size (IS) by 43% (p < 0.05). Splenectomy before ischemia did not change IS (vs. control, p = NS) but did serve to prevent the exacerbation of IS by hyperglycemia. Acute hyperglycemia activated splenic leukocytes by increasing formyl peptide receptor expression and reactive oxygen species production before ischemia, and enhanced splenic neutrophil release with resultant peripheral neutrophilia and increased myocardial neutrophil infiltration during reperfusion. Acute adoptive transfer of splenic leukocytes to splenectomized mice before ischemia restored the hyperglycemic exacerbation of infarct size. ATL146e, an adenosine 2A receptor (A2AR) agonist, abolished neutrophilia during reperfusion and reduced IS in hyperglycemic mice. ATL146e also reduced IS in splenectomized hyperglycemic mice with transfer of WT splenic leukocytes, but not with transfer of splenic leukocytes from A2AR knockout mice. Acute hyperglycemia prior to myocardial ischemia and reperfusion exacerbates IS by activating splenic leukocytes. ATL146e administered at reperfusion suffices to abrogate the hyperglycemic exacerbation of IS by acting on A2ARs on splenic leukocytes.

Abstract 3976: Expression of formyl peptide receptor 1 (FPR1) in neuroblastoma: Implications in tumorigenesis

Abstract The formyl peptide receptor 1 (FPR1) is a G protein-coupled receptor with pattern recognition properties. It is mainly expressed by myeloid cells and is involved in a broad range of host defense mechanisms. However, a variety of host-derived agonists of FPR1 have been identified, including formyl peptides released from disrupted mitochondria of necrotic cells. In the present study, we demonstrate that FPR1 is expressed in neuroblastoma cell lines and primary tumors and high expression is correlated with poor overall survival. Addition of the FPR1 agonist N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) to neuroblastoma cells in vitro causes enhanced proliferative activity, increase of intracellular calcium response and activation of STAT3 and MAPK/Erk signaling pathways. Currently, clonal neuroblastoma cell line with inducible activity of FPR1 gene is under testing in a mouse model to assess the significance of this receptor in vivo. Our findings so far suggest that FPR1 may play a significant role in neuroblastoma tumorigenesis and that therapeutic intervention of the FPR1 pathway may become an important clinical strategy in neuroblastoma therapy. Citation Format: Igor Snapkov, Per Kogner, John-Inge Johnsen, Baldur Sveinbjørnsson. Expression of formyl peptide receptor 1 (FPR1) in neuroblastoma: Implications in tumorigenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3976. doi:10.1158/1538-7445.AM2014-3976

Gene Expression of Formyl Peptide Receptor 1 in Eosinophils of Iraqi Patients with Extrinsic Allergic Asthma

The formyl peptide receptor 1 (FPR1) is a member of the seven transmembrane G-protein coupled receptors family, mainly expressed by polymorphonuclear and mononuclear phagocytes and play a role in chemotaxis, killing of microorganisms through phagocytosis, and plays a important role players in innate immunity and host defense. The expression of (FPR1) in eosinophils has been investigated intensively with many diseases. In this study we explain regulation of (FPR1) expression in eosinophils during allergic disease such as (extrinsic Asthma). The study were conducted by using Reverse Transcription Real-Time PCR technique, and the fold changes of (FPR1) were estimated in the house dust mite (HDM) hypersensitive asthmatic patients group and birch pollen hypersensitive asthmatic patients group as well as healthy control group. The results show that there are high significant increase fold changes of (FPR1) expression (up regulation) in eosinophils of HDM hypersensitive asthmatic patients group. In contrast, birch pollen hypersensitive asthmatic patients group have no significant increase in FPR1 expression when compared with healthy control group. In conclusion, we concluded that formyl peptide receptor 1 (FPR1) that expressed by eosinophils have potential role in immune regulation and allergic diseases mainly extrinsic asthma when patients exposed to exogenous aggressive factor such as house dust mite.