gw9508 and Inflammation

The free fatty acid receptor 4 (FFA4), also known as GPR120, is a G protein-coupled receptor that is activated by long-chain fatty acids and that has been associated with regulation of appetite, release of insulin controlling hormones, insulin sensitization, anti-inflammatory and potentially anti-obesity activity, and is progressively appearing as an attractive potential target for the treatment of metabolic dysfunctions such as obesity, type 2 diabetes and inflammatory disorders. Ongoing investigations of the pharmacological functions of FFA4 and validation of its potential as a therapeutic target depend critically on the appropriateness and quality of the available pharmacological probes or tool compounds. After a brief summary of the pharmacological functions of FFA4 and some general considerations on desirable properties for these pharmacological tool compounds, the individual compounds that have been or are currently being used as tools for probing the function of FFA4 in various in vitro and in vivo settings will be discussed and evaluated.

Topics: Animals; Biphenyl Compounds; Humans; Inflammation; Insulin Resistance; Methylamines; Obesity; Phenylpropionates; Propionates; Receptors, G-Protein-Coupled

Over the last decade, a subfamily of G protein-coupled receptors that are agonized by endogenous and dietary free-fatty acids (FFA) has been discovered. These free-fatty acid receptors include FFA2 and FFA3, which are agonized by short-chained FFA, as well as FFA1 and FFA4, which are agonized by medium-to-long chained FFA. Ligands for FFA1 and FFA4 comprise the family of long chain polyunsaturated omega-3 fatty acids including α-linolenic acid (ALA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), suggesting that many of the long-known beneficial effects of these fats may be receptor mediated. In this regard, FFA4 has gathered considerable interest due to its role in ameliorating inflammation, promoting insulin sensitization, and regulating energy metabolism in response to FFA ligands. The goal of this review is to summarize the body of evidence in regard to FFA4 signal transduction, its mechanisms of regulation, and its functional role in a variety of tissues. In addition, recent endeavors toward discovery of small molecules that modulate FFA4 activity are also presented.

Topics: Animals; Energy Metabolism; Fatty Acids, Nonesterified; Fatty Acids, Unsaturated; Gene Expression Regulation; Humans; Inflammation; Ligands; Liver; Macrophages; Metabolic Diseases; Methylamines; Organ Specificity; Pancreas; Propionates; Protein Isoforms; Receptors, G-Protein-Coupled; Signal Transduction

GPR120 has been reported to ameliorate inflammation in diabetes and diabetic complications. In this study, GW9508, the GPR120 agonist, was utilized in human retinal microvascular endothelial cells (HRMECs) exposed to high glucose (HG) to investigate the involvement of GPR120 in cellular viability and apoptosis as well as the association with the NLRP3 inflammasome.. The expression of GPR120 in HRMECs cultured under HG was firstly detected by Western blotting. HRMECs were then assigned to the normal control, GW9508, HG, and HG + GW9508 groups. The expression of the NLRP3 inflammasome consists of NLRP3, ASC, and caspase-1 and was detected by Western blotting and the downstream IL-1. GPR120 is expressed in HRMECs, and HG can reduce its expression in a time-dependent manner. GW9508 can attenuate inflammation by reducing the expression of NLRP3, ASC, caspase-1, IL-1. Activation of GPR120 protects retinal vascular endothelial cells from HG through inhibiting NLRP3 inflammasome. Thus, GPR120 might be a potential therapeutic target to reduce retinal endothelial damage in diabetic retinopathy.. ZIEL: Es gibt Studien, wonach GPR120 mit Diabetes und diabetischen Komplikationen einhergehende Entzündungen mindern kann. In dieser Studie wurde der GPR120-Agonist GW9508 bei menschlichen mikrovaskulären retinalen Endothelzellen (HMRECs) eingesetzt, die hohen Glukosekonzentrationen ausgesetzt wurden. Ziel der Studie war es, die Auswirkungen von GPR120 auf Zellviabilität und Apoptose sowie auf mögliche Interaktionen mit dem NLRP3-Inflammasom zu untersuchen.. Die Exprimierung von GPR120 in HMRECs, die in hohen Glukosekonzentrationen kultiviert wurden, wurde erstmals mit Western Blot entdeckt. Die HMRECs wurden in 4 Gruppen eingeteilt: eine Kontrollgruppe, eine GW9508-Gruppe, eine Hohe-Glukose-Gruppe (HG-Gruppe) und eine HG + GW9508-Gruppe. Das NLRP3-Inflammsom exprimiert NLRP3, ASC und Caspase-1. Deren Exprimierung wurde mit Western Blot und die nachgelagerte Exprimierung von IL-1. GPR120 wird in HMRECs exprimiert, und HG kann diese Exprimierung zeitabhängig reduzieren. GW9508 mindert Entzündungen, indem es die Exprimierung von NLRP3, ASC, Caspase-1, IL-1. Die GPR120-Aktivierung schützt retinale vaskuläre Endothelzellen vor HG durch Hemmung des NLRP3-Inflammasoms. GPR120 könnte potenziell ein neues therapeutisches Zielmolekül sein, das zur Reduzierung von Schäden an retinalen Endothelzellen eingesetzt werden könnte.

Topics: Apoptosis; bcl-2-Associated X Protein; Caspase 1; Endothelial Cells; Glucose; Humans; Inflammasomes; Inflammation; Interleukin-18; NLR Family, Pyrin Domain-Containing 3 Protein; RNA, Small Interfering

Oxidized low-density lipoprotein (ox-LDL)-induced endothelial inflammation plays an important role in the development of cardiovascular diseases. G protein-coupled receptors (GPCR) are gaining traction as potential treatment targets due to their roles in mediating a wide range of physiological processes. GPR120 is a recently identified omega-3 fatty acid receptor. We hypothesized that agonism of GPR120 might attenuate ox-LDL-induced endothelial dysfunction. In the present study, we tested the effects of two GPR120 agonists-GW9508 and TUG-891-in mitigating endothelial insult induced by ox-LDL in human aortic endothelial cells (HAECs). Real-time PCR, western blot, and ELISA analyses were used in our experiments. Our findings demonstrate that GPR120 is downregulated by exposure to ox-LDL, suggesting a role for GPR120 in mediating ox-LDL insult. Furthermore, we found that agonism of GPR120 could suppress oxidative stress and inflammation by inhibiting the production of reactive oxygen species and the expression of proinflammatory cytokines. Importantly, we show that agonism of GPR120 prevents the attachment of monocytes to endothelial cells by suppressing the expression of VCAM-1 and E-selectin. Finally, we show that agonism of GPR120 exerts a remarkable atheroprotective effect by elevating the expression level of Krüppel-like factor 2 (KLF2). Together, our results demonstrate a potential role for specific agonism of GPR120 in the prevention of endothelial damages induced by ox-LDL.

Topics: Cell Adhesion; Cell Survival; Cytokines; Down-Regulation; E-Selectin; Endothelial Cells; Humans; Inflammation; Kruppel-Like Transcription Factors; Lipoproteins, LDL; Methylamines; Monocytes; Oxidative Stress; Propionates; Reactive Oxygen Species; Receptors, G-Protein-Coupled; Vascular Cell Adhesion Molecule-1

Glucagon-like peptide (GLP)-2, secreted by L cells in the small intestine, has anti-inflammatory effects in the gastrointestinal tract. A GLP-2 analogue has been an effective treatment for Crohn disease (CD). G-protein-coupled receptor (GPR) 40 and GPR120 are probably involved in GLP-2 production, the mechanisms of which remain unclear. In our experiments, normal ileal mucosa expressed GPR40, but rarely expressed GPR120. However, both GPRs were overexpressed in the L cells of the inflamed ileal mucosa of CD patients. Mucosal inflammation induced the overexpression of GPR40, GPR120, and several inflammatory cytokines, with correlations between ileal concentrations of tumor necrosis factor (TNF)-α and GPR expression levels; however, inflammation did not induce the expression of proglucagon, a precursor of GLP-2 in CD patients. In rat L cells and GLUTag cells, TNF-α treatment increased GPR120 mRNA expression without affecting GPR40 mRNA expression. Dual agonists of GPR40 and GPR120, GW9508 and linoleic acid, respectively, increased GLP-2 production from L cells, but these agonists decreased it in the presence of TNF-α. The GPR40 antagonist, GW1100, inhibited the GW9508-induced increase in GLP-2 production, and silencing GPR120 resulted in further elevation of GLP-2 production. Thus, GPR120-dependent signaling inhibited the stimulatory effects of GPR40 on GLP-2 expression, and TNF-α treatment decreased GLP-2 expression by up-regulating GPR120 expression in L cells.

Topics: Adult; Aged; Aged, 80 and over; Animals; Benzoates; Case-Control Studies; Cohort Studies; Crohn Disease; Cytokines; Female; Gene Expression Regulation; Glucagon-Like Peptide 2; Humans; Immunoenzyme Techniques; Inflammation; Intestinal Mucosa; Male; Methylamines; Middle Aged; Propionates; Pyrimidines; Rats; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; Tumor Necrosis Factor-alpha; Up-Regulation; Young Adult

The G-protein-coupled receptor 40 (GPR40) is suggested to function as a transmembrane receptor for medium- to long-chain free fatty acids and is implicated to play a role in free fatty acids-mediated enhancement of glucose-stimulated insulin secretion from pancreas. However, the functional role of GPR40 in nervous system including somatosensory pain signaling has not been fully examined yet.. Intrathecal injection of GPR40 agonist (MEDICA16 or GW9508) dose-dependently reduced ipsilateral mechanical allodynia in CFA and SNL models and thermal hyperalgesia in carrageenan model. These anti-allodynic and anti-hyperalgesic effects were almost completely reversed by a GPR40 antagonist, GW1100. Immunohistochemical analysis revealed that GPR40 is expressed in spinal dorsal horn and dorsal root ganglion neurons, and immunoblot analysis showed that carrageenan or CFA inflammation or spinal nerve injury resulted in increased expression of GPR40 in these areas. Patch-clamp recordings from spinal cord slices exhibited that bath-application of either MEDICA16 or GW9508 significantly decreased the frequency of spontaneous excitatory postsynaptic currents in the substantia gelatinosa neurons of the three pain models.. Our results indicate that GPR40 signaling pathway plays an important suppressive role in spinal nociceptive processing after inflammation or nerve injury, and that GPR40 agonists might serve as a new class of analgesics for treating inflammatory and neuropathic pain.

Topics: Analgesics; Animals; Behavior, Animal; Disease Models, Animal; Excitatory Postsynaptic Potentials; Fatty Acids, Nonesterified; Ganglia, Spinal; Inflammation; Male; Methylamines; Mice, Inbred C57BL; Neuralgia; Neurons; Propionates; Receptors, G-Protein-Coupled; Spinal Nerves

Docosahexaenoic acid (DHA) is one of the major ingredients of fish oil and has been reported to have anti-inflammatory properties mediated through the GPR120 receptor. Whether cytosolic phospholipase A2 (cPLA2 ) and lipid mediators produced from cPLA2 activation are involved in the anti-inflammatory role of DHA in macrophages has not been reported. We report here that DHA and the GPR120 agonist, GW9508, activate cPLA2 and cyclooxygenase 2 (COX-2), and cause prostaglandin E2 (PGE2) release in a murine macrophage cell line RAW264.7 and in human primary monocyte-derived macrophages. DHA and GW9508 activate cPLA2 via GPR120 receptor, G protein Gαq and scaffold protein β-arrestin 2. Extracellular signal-regulated kinase 1/2 activation is involved in DHA- and GW9508-induced cPLA2 activation, but not p38 mitogen-activated protein kinase. The anti-inflammatory role of DHA and GW9508 is in part via activation of cPLA2 , COX-2 and production of PGE2 as a cPLA2 inhibitor or a COX-2 inhibitor partially reverses the DHA- and GW9508-induced inhibition of lipopolysaccharide-induced interleukin-6 secretion. The cPLA2 product arachidonic acid and PGE2 also play an anti-inflammatory role. This effect of PGE2 is partially through inhibition of the nuclear factor-κB signalling pathway and through the EP4 receptor of PGE2 because an EP4 inhibitor or knock-down of EP4 partially reverses DHA inhibition of lipopolysaccharide-induced interleukin-6 secretion. Hence, DHA has an anti-inflammatory effect partially through induction of PGE2.

Topics: Animals; Blotting, Western; Cell Line; Cytoplasm; Dinoprostone; Docosahexaenoic Acids; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Fish Oils; Humans; Inflammation; Macrophages; Methylamines; Mice; Phospholipases A2; Propionates; Real-Time Polymerase Chain Reaction; Receptors, G-Protein-Coupled; Receptors, Prostaglandin E, EP4 Subtype; RNA, Small Interfering; Transfection

This protocol describes microsphere-based protease assays for use in flow cytometry and high-throughput screening. This platform measures a loss of fluorescence from the surface of a microsphere due to the cleavage of an attached fluorescent protease substrate by a suitable protease enzyme. The assay format can be adapted to any site or protein-specific protease of interest and results can be measured in both real time and as endpoint fluorescence assays on a flow cytometer. Endpoint assays are easily adapted to microplate format for flow cytometry high-throughput analysis and inhibitor screening.

Topics: Animals; Biotinylation; Flow Cytometry; Fluorescence Resonance Energy Transfer; Green Fluorescent Proteins; High-Throughput Screening Assays; Humans; Inflammation; Kinetics; Microspheres; Peptide Hydrolases; Peptides; Reproducibility of Results; Temperature