Tripeptide FEG Research Articles

Prevention and Amelioration of Rodent Ventilation-Induced Lung Injury with Either Prophylactic or Therapeutic feG Administration.

Mechanical ventilation is a well-established therapy for patients with acute respiratory failure. However, up to 35% of mortality in acute respiratory distress syndrome may be attributed to ventilation-induced lung injury (VILI). We previously demonstrated the efficacy of the synthetic tripeptide feG for preventing and ameliorating acute pancreatitis-associated lung injury. However, as the mechanisms of induction of injury during mechanical ventilation may differ, we aimed to investigate the effect of feG in a rodent model of VILI, with or without secondary challenge, as a preventative treatment when administered before injury (prophylactic), or as a therapeutic treatment administered following initiation of injury (therapeutic). Lung injury was assessed following prophylactic or therapeutic intratracheal feG administration in a rodent model of ventilation-induced lung injury, with or without secondary intratracheal lipopolysaccharide challenge. Prophylactic feG administration resulted in significant improvements in arterial blood oxygenation and respiratory mechanics, and decreased lung oedema, bronchoalveolar lavage protein concentration, histological tissue injury scores, blood vessel activation, bronchoalveolar lavage cell infiltration and lung myeloperoxidase activity in VILI, both with and without lipopolysaccharide. Therapeutic feG administration similarly ameliorated the severity of tissue damage and encouraged the resolution of injury. feG associated decreases in endothelial adhesion molecules may indicate a mechanism for these effects. This study supports the potential for feG as a pharmacological agent in the prevention or treatment of lung injury associated with mechanical ventilation.

Tripeptide feG Prevents and Ameliorates Acute Pancreatitis-Associated Acute Lung Injury in a Rodent Model

The synthetic tripeptide feG (D-Phe-D-Glu-Gly) is a novel pharmacologic agent that decreases neutrophil recruitment, infiltration, and activation in various animal models of inflammatory disease. We aimed to investigate the effect of feG as both a preventive treatment when administered before acute lung injury and as a therapeutic treatment administered following initiation of acute lung injury. Lung injury was assessed following prophylactic or therapeutic intratracheal feG administration in a “two-hit” rodent model of acute pancreatitis plus intratracheal lipopolysaccharide. Following both prophylactic and therapeutic feG administration, there were significant improvements in arterial blood oxygenation and respiratory mechanics and decreased lung edema, BAL protein concentration, histologic tissue injury scores, BAL cell infiltration, and lung myeloperoxidase activity. Most indices of lung damage were reduced to baseline control values. feG reduced leukocyte infiltration, ameliorated the severity of inflammatory damage, and restored lung function when administered either prophylactically or therapeutically in a two-hit rat model of acute pancreatitis plus intratracheal lipopolysaccharide.

Prevention and amelioration of rodent endotoxin-induced lung injury with administration of a novel therapeutic tripeptide feG

BackgroundThe synthetic tripeptide feG is a novel pharmacological agent that decreases neutrophil recruitment, infiltration, and activation in various animal models of inflammatory disease. In human and rat cell culture models, feG requires pre-stimulation in order to decrease in vitro neutrophil chemotaxis. We aimed to investigate the effect of feG on neutrophil chemotaxis in a lipopolysaccharide-induced acute lung injury model without pre-stimulation. MethodsThe efficacy of feG as both a preventative treatment, when administered before lung injury (prophylactic), or as a therapeutic treatment, administered following lung injury (therapeutic), was investigated. ResultsProphylactic or therapeutic feG administration significantly reduced leukocyte infiltration, ameliorated the severity of inflammatory damage, and restored lung function. feG was demonstrated to significantly decrease bronchoalveolar lavage cell infiltration, lung myeloperoxidase activity, lung oedema, histological tissue injury scores, and improve arterial blood oxygenation and respiratory mechanics. ConclusionsfeG reduced leukocyte infiltration, ameliorated the severity of inflammatory damage, and restored lung function when administered prophylactically or therapeutically in a rodent model of lipopolysaccharide-induced acute lung injury, without the need for pre-stimulation, suggesting a direct rather than indirect mechanism of action in the lung.

P4.8 Structural features of the tripeptide feG accounting for different biological activities

The tripeptide FEG (Phe-Glu-Gly), derived from the c-terminus of the salivary gland peptide-T (SGP-T), TDFEGG, and its D-isomer feG are potent anti-inflammatory peptides that reduce type1 immediate hypersensitivity reactions such as vascular leak, intestinal motility disturbances, systemic hypotension, airways constriction and hyper-reactivity, and pulmonary inflammation. They also inhibit LPS-induced shock, cerulein-induced pancreatitis, zymosan-induced arthritis and other acute inflammatory responses.

Chronic use of the immunomodulating tripeptide feG–COOH in experimental feline asthma

We have previously documented that a single dose of feG–COOH prior to allergen challenge significantly decreased eosinophilic airway inflammation in cats with experimental asthma, but did not result in complete resolution of airway inflammation. This study was undertaken to determine if a chronic (2 weeks) course of feG–COOH in experimentally asthmatic cats would induce complete remission of airway inflammation and clinical signs of asthma. Experimental asthma was induced using Bermuda grass allergen (BGA) and cats were randomly selected to receive either feG–COOH (1 mg/kg, PO) or saline for 2 weeks, followed by a 2-week washout period. Cats then received the alternate treatment. Aerosol challenge with BGA was performed weekly throughout the study and bronchoalveolar lavage fluid (BALF) and blood were collected prior to and after each of the 2-week treatment periods. Regular use of feG–COOH had no significant effect on airway inflammation, BALF and plasma TNF bioactivity or a clinical sign compared to placebo. Regular use of feG–COOH can thus not be recommended as the sole therapy for feline allergic asthma.

The tripeptide feG inhibits leukocyte adhesion

BackgroundThe tripeptide feG (D-Phe-D-Glu-Gly) is a potent anti-inflammatory peptide that reduces the severity of type I immediate hypersensitivity reactions, and inhibits neutrophil chemotaxis and adhesion to tissues. feG also reduces the expression of β1-integrin on circulating neutrophils, but the counter ligands involved in the anti-adhesive actions of the peptide are not known. In this study the effects of feG on the adhesion of rat peritoneal leukocytes and extravasated neutrophils to several different integrin selective substrates were evaluated.ResultsThe adhesion of peritoneal leukocytes and extravasated neutrophils from rats to adhesive proteins coated to 96-well plates was dependent upon magnesium (Mg2+) ion, suggestive of integrin-mediated adhesion. feG inhibited leukocyte adhesion, but only if the cells were stimulated with PAF (10-9M), indicating that feG's actions in vitro require cell activation. In the dose range of 10-10M to 10-12M feG inhibited the adhesion of peritoneal leukocytes to fibrinogen and fibronectin, but not IgG, vitronectin or ICAM-1. feG inhibited the binding of extravasated neutrophils to heparin, IgG, fibronectin and CD16 antibody. Antigen-challenge of sensitized rats reduced the adhesion of peritoneal leukocytes to most substrates and abolished the inhibitory effects of feG. However, pretreating the animals with intraperitoneal feG (100 μg/kg) 18 h before collecting the cells from the antigen-challenged animal restored the inhibition of adhesion by in vitro feG of peritoneal leukocytes and extravasated neutrophils to fibronectin.ConclusionThe modulation of leukocyte adhesion by feG appears to involve actions on αMβ2 integrin, with a possible interaction with the low affinity FcγRIII receptor (CD16). The modulation of cell adhesion by feG is dual in nature. When administered in vivo, feG prevents inflammation-induced reductions in cell adhesion, as well as restoring its inhibitory effect in vitro. The mechanism by which in vivo treatment with feG exerts these effects remains to be elucidated.

A New Conformationally Restricted Mimetic of Dipeptide EG – Synthesis of an Analogue of FEG

AbstractStarting from the chiral pyrrolidin‐2‐one 2, the carboxy group at C‐4 underwent homologation, and subsequent removal of the 1‐(4‐methoxyphenyl)ethyl group gave lactam 6. Alkylation of N‐1 with benzyl bromoacetate led to 7, a new conformationally restricted analogue of the dipeptide EG (Glu‐Gly). The usefulness of 7 was demonstrated by its eventual conversion into 8, an orthogonally protected analogue ofbioactive tripeptide FEG. In order to provide the biological activity of the new mimetic 9, available from 8 after the removal of the protecting groups, the conformational preference of 9 was ascertained by a detailed conformational analysis and a comparison with that of FEG (1). (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)

Modulation of Neutrophil Function by Hormones

Endocrine regulation of the neutrophilic component of innate immunity with a particular focus on the production of reactive oxygen species (ROS) is the subject of this review. Endocrine hormones contribute to neutrophil mediated pathology in several ways either by directly activating the cells, as is the case with an angiotensin activation of NADPH oxidase on neutrophils and monocytes to contribute to atherosclerosis, or by having biphasic effects depending upon the stage of systemic inflammation, i.e. in a hyper-inflammatory phase or a hypo-inflammatory or anergic state, as seen with growth hormone. Some factors, such as a novel anti-inflammatory peptide from salivary glands, the tripeptide FEG, and hypothalamic proline-rich peptide have more subtle effects and appear to negatively regulate the priming events required for neutrophil activation. Investigations on the impact and role of endocrine hormones on neutrophil function and ROS production have contributed to the understanding of the efficacy of some medications in treating inflammation, as is the case with the angiotensin receptor blockers, and should lead to the development of novel drugs that modulate innate immune system function and the severity of an inflammatory response. Keywords: Hormones, peptides, reactive oxygen species, superoxide, leukocytes, anti-inflammatory

The tripeptide feG regulates the production of intracellular reactive oxygen species by neutrophils

BackgroundThe D-isomeric form of the tripeptide FEG (feG) is a potent anti-inflammatory agent that suppresses type I hypersensitivity (IgE-mediated allergic) reactions in several animal species. One of feG's primary actions is to inhibit leukocyte activation resulting in loss of their adhesive and migratory properties. Since activation of neutrophils is often associated with an increase in respiratory burst with the generation of reactive oxygen species (ROS), we examined the effect of feG on the respiratory burst in neutrophils of antigen-sensitized rats. A role for protein kinase C (PKC) in the actions of feG was evaluated by using selective isoform inhibitors for PKC.ResultsAt 18h after antigen (ovalbumin) challenge of sensitized Sprague-Dawley rats a pronounced neutrophilia occurred; a response that was reduced in animals treated with feG (100 μg/kg). With antigen-challenged animals the protein kinase C (PKC) activator, PMA, significantly increased intracellular ROS of circulating neutrophils, as determined by flow cytometry using the fluorescent probe dihydrorhodamine-123. This increase was prevented by treatment with feG at the time of antigen challenge. The inhibitor of PKCδ, rottlerin, which effectively prevented intracellular ROS production by circulating neutrophils of animals receiving a naïve antigen, failed to inhibit PMA-stimulated ROS production if the animals were challenged with antigen. feG treatment, however, re-established the inhibitory effects of the PKCδ inhibitor on intracellular ROS production. The extracellular release of superoxide anion, evaluated by measuring the oxidative reduction of cytochrome C, was neither modified by antigen challenge nor feG treatment. However, hispidin, an inhibitor of PKCβ, inhibited the release of superoxide anion from circulating leukocytes in all groups of animals. feG prevented the increased expression of the β1-integrin CD49d on the circulating neutrophils elicited by antigen challenge.ConclusionfeG reduces the capacity of circulating neutrophils to generate intracellular ROS consequent to an allergic reaction by preventing the deregulation of PKCδ. This action of feG may be related to the reduction in antigen-induced up-regulation of CD49d expression on circulating neutrophils.

Frontline: Inhibition of allergen‐induced pulmonary inflammation by the tripeptide feG: a mimetic of a neuro‐endocrine pathway

Interactions between the neuro-endocrine system and immune system help maintain health. One interaction involves the superior cervical ganglia (SCG), which regulate the prohormone submandibular rat 1 (SMR1) produced by the submandibular gland (SMG). A peptide derived from SMR1, feG, has anti-inflammatory activity, and modification to D-isomer feG enhances bioactivity. We tested feG as a therapeutic agent for airways inflammation, using rats sensitized by OVA or Nippostrongylus brasiliensis (Nb). Treatment with feG but not fdG down-regulated OVA-challenge-induced increases in bronchoalveolar lavage (BAL)-derived macrophages, eosinophils and PMN (neutrophils) by 44%, 69% and 67%, respectively, at 24 h. We found that feG also reduced ICAM-1 on BAL-derived macrophages and eosinophils by 27% and 65%, and L-selectin on PMN by 55% following OVA challenge. Furthermore, feG but not fdG reduced the OVA-induced TNF increase in BAL fluid. We showed that feG also down-regulated both hyper-responsiveness to methacholine (by 27%) and microgranulomata formation in the lung parenchyma. In Nb-challenged rats, feG treatment inhibited ex vivo allergen-induced contraction of tracheal smooth muscle by up to 73%. In conclusion, feG, which is a mimetic of a peptide derived from a rat salivary gland prohormone, has anti-inflammatory properties in allergic airways inflammation in Brown-Norway rats. The role of the SCG-SMG neuro-endocrine pathway in allergic asthma and other inflammatory diseases requires additional study.

A Tree-Based Algorithm for Determining the Effects of Solvation on the Structure of Salivary Gland Tripeptide NH3+-D-PHE-D-GLU-GLY-COO−

A D-enantiomeric analog of the submandibular gland rat-1 tripeptide FEG (Seq: NH3+-Phe-Glu-Gly-COO−) called feG (Seq: NH3+-D-Phe-D-Glu-Gly-COO−) was examined by molecular dynamics simulations in water. Previous in vacuo simulations suggested a conformation consisting predominantly of interactions between the Phe side chain and glutamyl-carboxyl group and a carboxyl/amino termini interaction. The solvated peptide was simulated using two approaches which were compared—a single 400-ns simulation and a “simulation tree.” The “tree” approach utilized 45 10-ns simulations with different conformations used as initial structures for given trajectories. We demonstrate that multiple short duration simulations are able to describe the same conformational space as that described by longer simulations. Furthermore, previously described in vacuo interactions were confirmed with amendments: the previously described head-to-tail arrangement of the amino and carboxyl termini, was not observed; the interaction between the glutamyl carboxyl and Phe side chain describes only one of a continuum of conformations present wherein the aromatic residue remains in close proximity to the glutamyl carbonyl group, and also interacts with either of the two available carboxyl groups. Finally, utilizing only two separate 10-ns trajectories, we were able to better describe the conformational space than a single 60-ns trajectory, realizing a threefold decrease in the computational complexity of the problem.

Identification of a binding site for the anti-inflammatory tripeptide feG

The mechanism of action of feG, an anti-inflammatory peptide, was explored using data mining, molecular modeling, and enzymatic techniques. The molecular coordinates of protein kinase A (PKA) were used to create six virtual isoforms of protein kinase C (PKCα, βI, βII, δ, ι, and ζ). With in silico techniques a binding site for feG was identified on PKCβI that correlated significantly with a biological activity, the inhibition of intestinal anaphylaxis. Since feG selectively increased the binding of a PKCβI antibody, it is proposed that this peptide inhibits the reassociation of the hydrophobic tail of PKCβI with its binding site and prevents the enzyme from assuming an inactive conformation.

Modulation of neutrophil function by the tripeptide feG

BackgroundNeutrophils are critical in the defense against potentially harmful microorganisms, but their excessive and inappropriate activation can contribute significantly to tissue damage and a worsening pathology. Through the release of endocrine factors submandibular glands contribute to achieving a balance in neutrophil function by modulating the state of activation and migratory potential of circulating neutrophils. A putative hormonal candidate for these effects on neutrophils was identified as a heptapeptide named submandibular gland peptide T (SGP-T; sequence = TDIFEGG). Since the tripeptide FEG, derived from SGP-T, and its D-amino acid analogue feG had similar inhibitory effects on inflammatory reactions, we investigated the effects of feG on human and rat neutrophil function.ResultsWith human neutrophils feG had no discernible effect on oxidative burst or phagocytosis, but in picomolar amounts it reduced PAF-induced neutrophil movement and adhesion, and the binding of CD11b by 34% and that of CD16b close to control values. In the rat feG (10-11M) reduced the binding of CD11b and CD16 antibodies to PAF-stimulated circulating neutrophils by 35% and 43%, respectively, and at 100 micrograms/kilograms intraperitoneally feG reduced neutrophil in vivo migration by 40%. With ovalbumin-sensitized rats that were challenged with antigen, feG inhibited binding of antibodies against CD16b but not CD11b, on peritoneal leukocytes.ConclusionsThe inhibitory effect of feG on neutrophil movement may be mediated by alterations in the co-stimulatory molecules CD11b and CD16.

The tripeptide FEG ameliorates systemic inflammatory responses to rat intestinal anaphylaxis.

Food allergies are generally associated with gastrointestinal upset, but in many patients systemic reactions occur. However, the systemic effects of food allergies are poorly understood in experimental animals, which also offer the opportunity to explore the actions of anti-allergic drugs. The tripeptide D-phenylalanine-D-glutamate-Glycine (feG), which potentially alleviates the symptoms of systemic anaphylactic reactions, was tested to determine if it also reduced systemic inflammatory responses provoked by a gastric allergic reaction. Optimal inhibition of intestinal anaphylaxis was obtained when 100 microg/kg of feG was given 20 min before the rats were challenged with antigen. The increase in total circulating neutrophils and accumulation of neutrophils in the heart, developing 3 h and 24 h, respectively, after antigen challenge were reduced by both feG and dexamethasone. Both anti-inflammatory agents reduced the increase in vascular permeability induced by antigen in the intestine and the peripheral skin (pinna), albeit with different time courses. Dexamethasone prevented increases in vascular permeability when given 12 h before antigen challenge, whereas feG was effective when given 20 min before ingestion of antigen. The tripeptide prevented the anaphylaxis induced up regulation of specific antibody binding of a cell adhesion molecule related to neutrophil activation, namely CD49d (alpha4 integrin). Aside from showing that intestinal anaphylaxis produces significant systemic inflammatory responses in non-intestinal tissues, our results indicate that the tripeptide feG is a potent inhibitor of extra-gastrointestinal allergic reactions preventing both acute (30 min) and chronic (3 h or greater) inflammatory responses.

The tripeptide feG reduces endotoxin-provoked perturbation of intestinal motility and inflammation.

Lipopolysaccharide (LPS)-induced intestinal endotoxaemia perturbs motility and causes activation and influx of inflammatory cells into the muscle tissue. Because rat submandibular gland peptide T (SGP-T; Thr-Asp-Ile-Phe-Glu-Gly-Gly), its carboxyl-terminal fragment tripeptide, FEG (Phe-Glu-Gly) and its D-isomeric analogue, feG, modulate intestinal anaphylactic reactions, we examined whether these peptides also modulate LPS-induced intestinal endotoxaemia in conscious rats. The disruption of the fasting pattern of intestinal MMCs (migrating motor complexes), induced by intravenous LPS (20 microg kg-1) injection, was prevented by all three peptides. The extravasation of leucocytes into the peritoneal cavity and increased expression of the activation marker CD18 on mesenteric tissue leucocytes (18 h after intraperitoneal injection of LPS) were reduced by orally administered feG, which also significantly decreased the number of intestinal tissue leucocytes expressing the integrin CD18. We conclude that feG attenuates both the immediate (intestinal motility) and late ( approximately 18 h) inflammatory reactions provoked by endotoxaemia.

Regulation of leukocyte adhesion to heart by the tripeptides feG and feG(NH2)

The role of the D-isomeric form of the salivary gland tripeptide FEG (feG) and its carboxyl-amidated derivative, feG(NH2), in regulating leukocyte adherence to nonfixed atrial slices from Sprague-Dawley rats was examined under static conditions. Optimal binding of the leukocytes was seen if the leukocytes were treated with platelet activating factor (PAF; 10(-9)M). The increased adherence of PAF-treated peripheral blood leukocytes was totally inhibited by both feG and feG(NH2) (10-9M), as well as by antibodies against CD18 and CD49d. In contrast, the binding of peritoneal leukocytes was blocked only by CD49d antibody. Circulating leukocytes obtained from lipopolysaccharide (LPS) treated (2 mg/kg ip) rats did not bind to atrial slices obtained from normal hearts, but readily bound to atrial slices obtained from LPS-treated rats. This leukocyte binding was inhibited by in vivo feG treatment (100 microg/kg ip, 24 h before harvest) or by treating the isolated cells with feG (10(-9)M). The amidated peptide feG(NH2) reduced neutrophil accumulation in the atrium elicited by ip injection of LPS, whereas feG was ineffective. The reduction in neutrophil infiltration into the myocardium by feG(NH2) and the prevention of leukocyte interaction with myocytes seen with both feG and feG(NH2) probably results in hindered leukocyte migration in the inflamed heart, resulting in less tissue damage. The inhibition by these tripeptides on neutrophil adhesion to myocytes suggests that salivary glands hormones regulate the severity of cardiac inflammation.

Inhibition of Allergic Inflammation by C-Terminal Peptides of the Prohormone Submandibular Rat 1 (SMR-1)

Background: The C-terminal of the prohormone submandibular rat 1 protein (SMR-1) contains several small peptides that reduce the severity of allergic inflammation and septic shock, and are part of the cervical sympathetic trunk-submandibular gland (SMG) axis of neuroendocrine immunology. These peptides include the heptapeptide, submandibular gland peptide-T and the tripeptide FEG. The D-isomeric form of this tripeptide, feG, which is active when administered orally, reduces LPS-provoked leukocyte rolling on mesenteric venules and influx of inflammatory cells into the peritoneum and intestinal muscle. Methods: To investigate the mechanism of action of these peptides, the influx of inflammatory molecules into the airways, and several properties of human neutrophils were examined. Results: Oral feG (1 mg/kg) inhibited the influx of inflammatory cells into the airways lumen of allergen challenged, sensitized Brown Norway rats. This inhibition occurred whether feG was given 30 min prior to 6 h post allergen challenge. Moreover, feG in picomolar to nanomolar concentrations inhibited PAF elicited chemotaxis by 30–40%, but the peptides did not affect superoxide production or phagocytosis by neutrophils. feG reduced PAF-stimulated expression of CD11b. Conclusions: feG may exert its anti-inflammatory effects by modulating the expression and functions of β₂ integrins. The CST-SMG axis may be a major neuroendocrine pathway that modulates allergic asthma and other inflammatory responses.

Reduction of endotoxin-induced leukocyte activation in the rat intestine by a D-isomeric analog of salivary gland tripeptide FEG

Reduction of endotoxin-induced leukocyte activation in the rat intestine by a D-isomeric analog of salivary gland tripeptide FEG

Attenuation of intestinal and cardiovascular anaphylaxis by the salivary gland tripeptide FEG and its d-isomeric analog feG

The effects of the submandibular gland peptide-T (SGP-T; Thr-Asp-Ile-Phe-Gly-Gly; TDIFEGG), its carboxy-terminal fragment (the tripeptide FEG; Phe-Glu-Gly), and the d-isomeric analog (feG) on intestinal and cardiovascular anaphylactic reactions were studied. The tripeptides, FEG and feG, when administered intravenously or orally to egg albumin-sensitized Hooded Lister or Sprague–Dawley rats 30 min prior to challenge with the antigen, totally prevented the disruption of intestinal motility and the development of anaphylaxis provoked diarrhea and inhibited anaphylactic hypotension by 66%. Submandibular gland peptides participate in the regulation of systemic inflammatory reactions, and the d-amino acid tripeptide, feG, is a potent, orally active anti-anaphylactic agent.

Submandibular gland peptide-T (SGP-T): Modulation of endotoxic and anaphylactic shock

This report summarizes recent studies on the antishock activities of a novel peptide, submandibular gland peptide-T (SGP-T). This heptapeptide (TDIFEGG) was isolated from the submandibular glands of rats based on the ability of extracts of these glands to reduce the severity of the hypotension induced by lipopolysaccharide. SGP-T was also found to reduce the severity of anaphylactic shock in ovalbumin-sensitized rats. Structure–activity relationship studies revealed that the inhibition of intestinal anaphylaxis in vitro only required a portion of SGP-T, i.e., the tripeptide FEG, localized at the carboxy terminal of the parent heptapeptide. Both FEG and its d-isomeric form, feG, inhibited anaphylactic hypotension when administered intravenously prior to exposure to the antigen. The peptide feG also inhibited anaphylactic hypotension when given orally 1 h before exposure to the antigen. SGP-T may be a prototype to a family of small peptides that modulate the immunological and smooth muscle reactions to severe inflammatory (endotoxic and anaphylactic) reactions. Drug Dev. Res. 42:164–171, 1997. © 1997 Wiley-Liss, Inc.