ABSTRACT
PURPOSE: Urokinase receptor (uPAR) promotes extracellular matrix proteolysis, regulates adhesion and cell migration, transduces intracellular signals through interactions with the lateral partners. The expression of uPAR and urokinase (uPA) is significantly upregulated in peripheral nerves after injury, however, little is known about uPAR function in nerve regeneration or the molecular mechanisms involved. The purpose of this study is to investigate the role of uPAR in nerve regeneration after traumatic injury of n. Peroneus communis in uPA-/-, uPAR-/- or control mice (WT) and in neuritogenesis in an in vitro Neuro 2A cell model. RESULTS: Electrophysiological analysis indicates that nerve recovery is significantly impaired in uPAR-/- mice, but not in uPA-/- mice. These data correlate with the reduced amount of NF200-positive axons in regenerating nerves from uPAR-/- mice compared to uPA-/- or control mice. There is an increase in uPAR expression and remarkable colocalization of uPAR with α5 and ß1 integrin in uPA-/- mice in recovering nerves, pointing to a potential link between uPAR and its lateral partner α5ß1-integrin. Using an in vitro model of neuritogenesis and α325 blocking peptide, which abrogates uPAR-α5ß1 interaction in Neuro 2A cells but has no effect on their function, we have further confirmed the significance of uPAR-α5ß1 interaction. CONCLUSION: Taken together, we report evidence pointing to an important role of uPAR, rather than uPA, in peripheral nerve recovery and neuritogenesis.
Subject(s)
Integrin alpha5beta1/metabolism , Nerve Regeneration/genetics , Receptors, Urokinase Plasminogen Activator/genetics , Urokinase-Type Plasminogen Activator/genetics , Animals , Cell Line, Tumor , Cell Movement/genetics , Extracellular Matrix/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Nerve Regeneration/physiologyABSTRACT
Automated Fmoc solid-phase technique was used to synthesize Cys-containing linear peptide fragments of monocyte chemoattractant protein-1 and chemokine domain of fractalkine along with their analogues with Cys residue being either modified or replaced with Ser. Chimeric symmetric and asymmetric disulfides were also prepared from the former linear precursors. A SAR study on a set of the newly synthesized peptides revealed that capacity to stimulate migration of monocytes and to influence cell motility in vitro, in general, critically depends on the presence of Cys free thiol group in the molecule. Notably, all analogs lacking this feature, including chimeric disulfides, demonstrated lack of effect on monocyte migration.
Subject(s)
Cell Movement/drug effects , Chemokine CCL2 , Monocytes/metabolism , Peptides , Cell Line, Tumor , Humans , Monocytes/cytology , Peptides/chemical synthesis , Peptides/chemistry , Peptides/pharmacologyABSTRACT
Effects of apelin-12 H-Arg-Pro-Arg-Leu-Ser-His-Lys-Gly-Pro-Met-Pro-Phe-OH (A12) and its modified analogue H-(NMe)Arg-Pro-Arg-Leu-Ser-His-Lys-Gly-Pro-Nle-Pro-Phe-OH (I) on activity of antioxidant enzymes, formation of malonic dialdehyde (MDA) and generation of reactive oxygen species (ROS) were studied in ex vivo and in vivo models of myocardial ischemia and reperfusion (I/R) injury in Wistar rats. Preischemic infusion of peptide A12 or AI enhanced cardiac function recovery of isolated perfused heart and was accompanied by a marked attenuation of ROS generation detected by electron paramagnetic resonance (EPR) technique in myocardial effluent at early reperfusion compared with control. Intravenous administration (i.v.) of peptides in narcotized rats with regional myocardial ischemia limited infarct size and reduced activity of lactate dehydrogenase and MB-fraction of creatine kinase in plasma at the end of reperfusion. Treatment with peptide A12 prevented reduction or augmented activity of myocardial u/Zn superoxide dismutase, catalase and glutathione peroxidase by the end of reperfusion in both I/R models compared with control. Increased MDA content in the area at risk of rat heart in situ at the end of reperfusion was reduced to the initial value under the effect of i.v. A12 administration. Therefore, cardioprotective action of natural apelin-12 and its structural analog AI involve reduction of short-lived ROS generation and improvement of the antioxidant state of ischemic heart during reperfusion.
Subject(s)
Intercellular Signaling Peptides and Proteins/pharmacokinetics , Myocardial Ischemia/metabolism , Myocardial Reperfusion Injury/metabolism , Myocardium/metabolism , Oxidative Stress , Animals , Disease Models, Animal , Intercellular Signaling Peptides and Proteins/chemistry , Intercellular Signaling Peptides and Proteins/therapeutic use , Male , Malondialdehyde/metabolism , Myocardial Ischemia/drug therapy , Myocardial Reperfusion Injury/drug therapy , Rats , Rats, WistarABSTRACT
Linear peptides corresponding to fragment 83-98 of the first loop and fragments 168-192 and 171-182 of the second extracellular loops of M2-muscarinic receptor (marker of early cardiac disorders and arrhythmias) were synthesized by Fmoc-SPPS method. A new conformational antigen was synthesized by method of selective ligation of linear peptides by disulfide bond with native localization. Peptides were studied in reaction with sera from patients with idiopathic arrhythmias. A new conformational antigen was recognized by sera from patients with idiopathic arrhythmias with high reactivity.
Subject(s)
Arrhythmias, Cardiac/immunology , Peptide Fragments/immunology , Receptor, Muscarinic M2/immunology , Vaccines, Synthetic/pharmacology , Amino Acid Sequence , Arrhythmias, Cardiac/blood , Arrhythmias, Cardiac/drug therapy , Autoantibodies/blood , Autoantibodies/immunology , Autoantibodies/isolation & purification , Humans , Peptide Fragments/chemical synthesis , Peptide Fragments/pharmacology , Protein Conformation , Receptor, Muscarinic M2/metabolism , Receptors, Adrenergic, beta-1/immunology , Vaccines, Synthetic/chemistry , Vaccines, Synthetic/immunologyABSTRACT
Novel peptides originating from the peptide inhibitor of myosin light chain kinase, L-PIK (Arg-Lys-Lys-Tyr-Lys-Tyr-Arg-Arg-Lys), have been studied for ability to attenuate the thrombin-induced hyperpermeability of endothelial cell monolayer in culture. Peptides [NalphaMeArg1]-Lys-Lys-Tyr-Lys-Tyr-Arg-(D)Arg8-Lys and H-Arg(NO2)Lys-Lys-Tyr-Lys-Tyr-Arg-Arg-Lys-NH2 (designated PIK2 and PIK4, respectively) appeared to be the most effective inhibitors of endothelial cell monolayer hyperpermebility, and surpassed other known peptide inhibitors of myosin light chain kinase derived from original L-PIK. Our results validate PIK2 and PIK4 as the leading molecules for the development of novel drugs intended to counteract pathological hyperpermeability of vascular endothelium.
Subject(s)
Capillary Permeability/drug effects , Endothelial Cells/drug effects , Endothelium, Vascular/drug effects , Myosin-Light-Chain Kinase/antagonists & inhibitors , Peptides/pharmacology , Amino Acid Sequence , Cell Line , Diffusion Chambers, Culture , Electric Impedance , Endothelial Cells/cytology , Endothelial Cells/enzymology , Endothelium, Vascular/cytology , Endothelium, Vascular/enzymology , Fluorescein-5-isothiocyanate/analogs & derivatives , Humans , Kinetics , Molecular Sequence Data , Myosin-Light-Chain Kinase/metabolism , Peptides/chemical synthesis , Serum Albumin , Spectrometry, Fluorescence , Structure-Activity Relationship , Thrombin/pharmacologyABSTRACT
We have studied anti-tumor properties of bone marrow derived peptide Phe-Leu-Gly-Phe-Pro-Thr (MP-1) synthesized by classical methods. It was shown that MP-1 enhanced the effect ofcytostatic therapy of lymphatic leukemia P388 and increased latent growth period of P388 tumors implanted in irradiated mice. MP-1 also decreased metastasis of mouse breast adenocarcinoma Ca-755 after surgery.
Subject(s)
Antineoplastic Agents , Leukemia P388 , Oligopeptides , Animals , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/chemical synthesis , Cell Line, Tumor , Cisplatin/administration & dosage , Combined Modality Therapy , Leukemia P388/drug therapy , Leukemia P388/radiotherapy , Mice , Oligopeptides/administration & dosage , Oligopeptides/chemical synthesisABSTRACT
The apelin-12 and a number of its analogs, resistant to degradation of proteases, were synthesized by Fmoc- method of SPPS. By-products of synthesis were examined. It was found that serine hydroxyl group was sulfating during the final deprotection of apelin-12 (I) and its analogs. Sulfate moiety of Arg-protecting group transfer into hydroxyl group of Ser. Amount of by-product depends on presence of water in cleavage mixture. Furthermore, the final deprotection of amide analogs of apelin-12 (III, IV) is closed with formation of by-product--4-hydroxybenzylamide, its amount range on 20-8% on reaction mixture accordance HPLC data and also depend on composition of cleavage mixture. Effects of the synthesized peptides on recovery of cardiac function after ischemia were examined in a model of isolated perfused rat heart. Infusions of any of the peptides (I-V) before ischemia resulted in a significant improvement of contractile and pump function recovery compared to the control. Cardioptotective efficacy of the peptides increased in the following rank (I) < (II) = (III) < (IV) = (V).
Subject(s)
Cardiotonic Agents , Intercellular Signaling Peptides and Proteins , Myocardial Ischemia/drug therapy , Myocardial Ischemia/physiopathology , Recovery of Function/drug effects , Animals , Cardiotonic Agents/chemical synthesis , Cardiotonic Agents/chemistry , Cardiotonic Agents/pharmacokinetics , Intercellular Signaling Peptides and Proteins/chemical synthesis , Intercellular Signaling Peptides and Proteins/chemistry , Intercellular Signaling Peptides and Proteins/pharmacology , Male , Rats , Rats, WistarABSTRACT
Apelin 12 (A-12) was synthesized by the automatic solid phase method with the use of Fmoc technology. The synthesized peptide was purified by preparative HPLC and identified by 1H-NMR spectroscopy and mass spectrometry. Acute myocardial infarction was induced by 40-min LAD occlusion followed by 60-min reperfusion in narcotized Wistar rats. A-12 was administrated at the onset of the reperfusion at doses of 0.07, 0.35 and 0.70 micromole/kg; N(G)-nitro-L-arginine methyl ester (L-NAME), a NOS inhibitor, was applied at a dose of 10 mg/kg 10 min prior to reperfusion alone or before A-12 administration (0.35 micromole/kg); saline was used in control. The indicated A-12 doses induced a transient reduction of the arterial systolic blood pressure (ASBP) to 85, 58, and 56% of the initial level, respectively, which was accompanied by its recovery by the end of reperfusion. All A-12 doses significantly limited myocardial infarct size by 26, 40 and 33%, respectively, compared to the value in control. After administration of A-12 at dose of 0.35 micromol/kg, this effect was combined with reduction of MB-creatine kinase (MB-CK) and lactate dehydrogenase (LDH) activities in plasma at the end of reperfusion by 56 and 47%, respectively, compared to the values in control. Inhibition of NO formation by L-NAME increased SABP but did not affect myocardial infarct size compared with that in control. Coadministration of L-NAME and A-12 resulted in lesser reduction of ASBP during reperfusion than injection of A-12 alone. This intervention led to an increase in infarct size by 26% with concomitant 1.8- and 1.5-times elevation of MB-CK and LDH activities, respectively, compared to the values in the A-12 group. The results indicate that NO is involved as a mediator of the effects of A-12 on the overall protection consisting in a limitation of infarct size and reduction of postischemic cardiomyocyte membrane damage. Cardioprotective mechanisms of apelin action are discussed.
Subject(s)
Intercellular Signaling Peptides and Proteins , Myocardial Contraction/drug effects , Myocardial Ischemia/drug therapy , Myocardial Reperfusion Injury/drug therapy , Myocytes, Cardiac/drug effects , Nitric Oxide/metabolism , Animals , Cardiotonic Agents/administration & dosage , Cardiotonic Agents/chemical synthesis , Cardiotonic Agents/pharmacokinetics , Disease Models, Animal , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/pharmacokinetics , Intercellular Signaling Peptides and Proteins/administration & dosage , Intercellular Signaling Peptides and Proteins/chemical synthesis , Intercellular Signaling Peptides and Proteins/pharmacokinetics , Male , Models, Cardiovascular , Monitoring, Physiologic/methods , Myocardial Contraction/physiology , Myocardial Ischemia/metabolism , Myocardial Ischemia/physiopathology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/physiopathology , Myocytes, Cardiac/metabolism , NG-Nitroarginine Methyl Ester/administration & dosage , NG-Nitroarginine Methyl Ester/pharmacokinetics , Nitric Oxide Synthase/antagonists & inhibitors , Nitric Oxide Synthase/metabolism , Rats , Rats, Wistar , Signal Transduction/drug effectsSubject(s)
Intercellular Signaling Peptides and Proteins/chemistry , Myocardial Infarction/drug therapy , Peptides/pharmacology , Amino Acid Sequence , Animals , Blood Pressure/drug effects , Creatine Kinase, MB Form/metabolism , Heart Rate/drug effects , Intercellular Signaling Peptides and Proteins/pharmacology , L-Lactate Dehydrogenase/metabolism , Male , Myocardial Infarction/metabolism , Myocardial Infarction/pathology , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/prevention & control , Peptides/chemistry , Rats , Rats, Wistar , Structure-Activity RelationshipABSTRACT
Leukocyte chemotaxis to the area of tissue damage is mediated by chemokines. According to the primary structure, chemokines are divided into four families, fractalkine (CX3CL1) is the only one member of CX3C family and the only membrane-bound chemokine. Fractalkine molecule includes the extracellular N-terminal chemokine domain, mucin-like rod, the transmembrane and the intracellular domains. In membrane-bound state fractalkine has the properties of an adhesion molecule. Chemokine domain of fractalkine (CDF) is released from cell membrane by proteolysis, and this soluble form acts as a chemoattractant for leukocytes expressing fractalkine receptor CX3CR1. Fractalkine is involved in development of a number of pathological processes caused by inflammation, and therefore a search for fractalkine inhibitors is very important. For this purpose we identified several antigenic determinants--the fragments of CDF, and the following peptides were synthesized--P41-52 H-Leu-Glu-Thr-Arg-Gln-His-Arg-Leu-Phe-Cys-Ala-Asp-NH2, P53-60 H-Pro-Lys-Glu-Gln-Trp-Val-Lys-Asp-NH2 and P60-71 H-Asp-Ala-Met-Gln-His-Leu-Asp-Arg-Gln-Ala-Ala-Ala-NH2. The peptide effects on adhesion and migration of human peripheral blood monocytes expressing fractalkine receptors were investigated. In the presence of CDF and P41-52 we observed the increased adhesion and migration of monocytes compared with spontaneous values. Peptides P53-60 and P60-71 significantly inhibited monocyte adhesion and migration stimulated by CDF. Since the chemotactic activity of chemokines was shown to be dependent on their binding to glycosaminoglycans of the cell surface and extracellular matrix, the effect ofpeptides on the interaction of CDF with heparin was analyzed by ELISA. Peptide P41-52 competed with CDF for heparin binding, while peptides P53-60 and P60-71 had no significant activity.
Subject(s)
Cell Adhesion , Cell Movement , Chemokine CX3CL1 , Monocytes/cytology , Peptide Fragments , Amino Acid Sequence , Cell Adhesion/drug effects , Cell Adhesion/immunology , Cell Adhesion/physiology , Cell Movement/drug effects , Cell Movement/immunology , Cell Movement/physiology , Chemokine CX3CL1/chemical synthesis , Chemokine CX3CL1/chemistry , Chemokine CX3CL1/immunology , Chemotaxis, Leukocyte , Humans , Monocytes/metabolism , Peptide Fragments/chemical synthesis , Peptide Fragments/chemistry , Peptide Fragments/immunologyABSTRACT
Apelin 12 (A-12) was synthesized by the automatic solid phase method with use of Fmoc 1H-NMR spectroscopy and mass spectrometry. Effects of apelin-12 (a peptide comprised of 12 aminoacids, A-12) on recovery of energy metabolism and cardiac function were studied in isolated working rat hearts perfused with Krebs buffer (KB) containing 11 mM glucose that were subjected to global ischemia and reperfusion. A short-term infusion of microM 140 A-12 in KB prior to ischemia enhanced myocardial ATP, the total adenine nucleotide pool (SigmaAN = ATP + ADP + AMP) and the energy charge of cardiomyocites ((ATP + 0.5ADP)/SigmaAN) at the end of reperfusion compared with control (KB infusion) and reduced lactate content and lactate/pyruvate ratio in reperfused myocardium to the initial values. This effect was accompanied by improved recovery of coronary flow and cardiac function. Coadministration of 140 microM A-12 and 100 microM L-NAME (the nonspecific NOS inhibitor) profoundly attenuated the peptide influence on metabolic and functional recovery of reperfused hearts. The results indicate involvement of NO, formed under the peptide action, in mechanisms of cardioprotection that are tightly associated with recovery of energy metabolism in postischemic heart.
Subject(s)
Energy Metabolism/drug effects , Intercellular Signaling Peptides and Proteins/pharmacology , Myocardial Ischemia/metabolism , Adenosine Triphosphate/metabolism , Animals , Enzyme Inhibitors/pharmacology , In Vitro Techniques , Intercellular Signaling Peptides and Proteins/chemical synthesis , Isotonic Solutions/pharmacology , Lactates/metabolism , Male , Myocardial Ischemia/drug therapy , Myocardial Ischemia/physiopathology , Myocardial Reperfusion , NG-Nitroarginine Methyl Ester/pharmacology , Nitric Oxide/metabolism , Rats , Rats, WistarABSTRACT
Apelin-12 (A-12) peptide was synthesized by automated solid phase method and purified by reverse phase HPLC. Its homogeneity and structure were confirmed by HPLC, (1)H-NMR spectroscopy, and mass spectroscopy. Acute myocardial infarction was induced by 40-min occlusion of the left coronary artery with subsequent 60-min reperfusion in narcotized Wistar rats. Peptide A-12 was injected (intravenous bolus, 0.07 or 0.35 µmol/kg) to experimental animals simultaneously with the beginning of reperfusion. Injections of A-12 in these doses led to reduction of systolic BP to 67 and 85% of the initial level, respectively, which was virtually restored completely by the end of reperfusion, and to a significant reduction of the infarction focus in the myocardium (by 21 and 34% in comparison with the control, respectively). Injection of A-12 in a dose of 0.35 µmol/kg led to reduction of plasma concentrations of necrosis markers in comparison with the control by the end of reperfusion: MB-creatine kinase by 56%, lactate dehydrogenase by 30%. The results attest to vasodilatory effects of A-12 under conditions of heart reperfusion in vivo; the peptide injected after local ischemia limits the myocardial infarction size and reduces damage to cardiomyocyte membrane.
Subject(s)
Cardiotonic Agents/therapeutic use , Intercellular Signaling Peptides and Proteins/therapeutic use , Myocardial Reperfusion Injury/prevention & control , Animals , Blood Pressure/drug effects , Cardiotonic Agents/chemical synthesis , Cardiotonic Agents/pharmacology , Creatine Kinase, MB Form/blood , Heart Ventricles/pathology , Intercellular Signaling Peptides and Proteins/chemical synthesis , Intercellular Signaling Peptides and Proteins/pharmacology , L-Lactate Dehydrogenase/blood , Male , Myocardial Reperfusion Injury/blood , Nitrates/blood , Nitrites/blood , Rats , Rats, WistarABSTRACT
Apelin 12 (A 12) was synthesized by the automatic solid phase method with the use of Fmoc technology. The synthesized peptide was purified by preparative HPLC and identified by 1H NMR spectroscopy and mass spectrometry. Effects of A 12 were studied on isolated working rat hearts perfused with Krebs buffer (KB) containing 11 mM glucose. The hearts were subjected to 35 min global ischemia followed by 30 min reperfusion. A short term infusion of A 12 in KB (35, 70, 140, 280, and 560 M) was applied prior to ischemia (A 12 I) or at onset of reperfusion (A 12 R). KB infusion without A 12 was used in control. A 12 infusion enhanced recovery of coronary flow, contractile and pump function during reperfusion with the largest augmentation of these indices in A 12 I group. Thus after infusion of 140 M A 12 recovery of coronary flow, the LVDP HR product and cardiac output were 92+/-5, 81+/-5, and 77+/-5% of the initial values, respectively, in A 12 I group, 83+/-6, 61+/-5, and 52+/-5% in A 12 R group, and 76+/-2, 42+/-2, 32+/-2% in control by the end of reperfusion. Both A 12 groups exhibited significant reduction of ischemia/reperfusion contracture compared with control. Enhanced functional recovery in A 12 I group was combined with a decrease in lactate dehydrogenase leakage in perfusate at early reperfusion (at the average by 36+/-5% compared with control, <0.05). Preischemic infusion of 140 M A 12 markedly increased myocardial ATP content and twice decreased AMP accumulation at the end of reperfusion. These alterations resulted in enhanced preservation of the total adenine nucleotide pool (to 81+/-5% of the initial value vs. 66+/-3% in control, <0.05) and better recovery of the energy charge potential (0.77+/-0.01 vs. 0.60+/-0.06 in control, <0.005) in reperfused hearts. At the end of experiment myocardial lactate and lactate/pyruvate ratio were on average 5 fold lower in A 12 I treated hearts compared with control one and did not differ significantly from initial values. This finding implies that better restoration of energy metabolism in hearts protected with A 12 before ischemia might be attributed to ameliorated glucose oxidation during reperfusion. Therefore enhanced functional recovery of ischemic heart and lesser cell membrane damage induced by A 12 were associated with maintaining high energy phosphates, particularly ATP, in reperfused myocardium. Cardioprotective mechanisms of apelin action are discussed.
Subject(s)
Energy Metabolism , Heart/drug effects , Intercellular Signaling Peptides and Proteins , Myocardial Contraction/drug effects , Myocardial Ischemia , Recovery of Function/drug effects , Animals , Coronary Circulation/drug effects , Drug Administration Routes , Drug Administration Schedule , Energy Metabolism/drug effects , Energy Metabolism/physiology , Heart/physiopathology , Intercellular Signaling Peptides and Proteins/chemical synthesis , Intercellular Signaling Peptides and Proteins/pharmacology , Male , Models, Animal , Myocardial Ischemia/drug therapy , Myocardial Ischemia/metabolism , Myocardium/metabolism , Protective Agents/chemical synthesis , Protective Agents/pharmacology , Rats , Rats, WistarABSTRACT
Myosin light chain kinase (MLCK) is the key regulator of various forms of cell motility including endothelial and epithelial permeability in particular. One of the potential MLCK inhibitors to be used in humans is a membrane permeable peptide H-RKKYKYRRK-NH2 (L-PIK). In present work we used solid phase peptide synthesis and Fmoc-technology to produce five modifications of L-PIK. Based on (1)H NMR analysis revealed that these peptides demonstrated improved resistance to degradation in blood plasma. One of de novo synthesized peptides, L-[MeArg(1)]PIK inhibited MLCK activity in vitro with the same efficiency as L-PIK whereas other modified peptides showed reduced inhibitory activity. D-amino acid analog of PIK was the least active inhibitor. Thus, we have demonstrated the possibility to produce an effective MLCK peptide inhibitor with increased resistance to biodegradation that is suitable for further pharmacological development.
Subject(s)
Myosin-Light-Chain Kinase/antagonists & inhibitors , Oligopeptides , Peptide Hydrolases/chemistry , Plasma/enzymology , Protein Kinase Inhibitors , Nuclear Magnetic Resonance, Biomolecular , Oligopeptides/chemical synthesis , Oligopeptides/chemistry , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistryABSTRACT
Two fragments corresponding to the 125-133 and 206-218 sequences of a molecule of the beta(1) adrenoreceptor (autoantibodies to this protein are often found in patients with dilated cardiomyopathy) were synthesized by the solid phase method with the use of Fmoc technology. Two new conformational antigens were prepared by directed (regioselective) and undirected (spontaneous) formation of intramolecular and intermolecular disulfide bridges between the corresponding cysteine residues of the synthesized peptides. One of these antigens consisted of a mixture of disulfide isomers, and another antigen was an isomer with a natural arrangement of S-S bridges. Immunosorbents were obtained by immobilization of the synthesizes antigens on the bromocyanogenactivated sepharose and applied to the removal of autoantibodies in a beta(1)-adrenoreceptor from the blood plasma of patients. We demonstrated that the sorbents on the basis of the conformational antigens were more effective in comparison with those containing linear peptide precursors.
Subject(s)
Antigens/chemistry , Disulfides/chemical synthesis , Peptides/chemical synthesis , Receptors, Adrenergic, beta-1/chemistry , Antigens/immunology , Autoantibodies/blood , Autoantibodies/isolation & purification , Cardiomyopathy, Dilated/blood , Cardiomyopathy, Dilated/immunology , Chromatography, High Pressure Liquid , Disulfides/chemistry , Humans , Immunosorbent Techniques , Peptides/chemistry , Peptides/immunology , Protein Structure, Tertiary , Receptors, Adrenergic, beta-1/immunologyABSTRACT
Neuropeptide FF (H-Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-NH2) injected intravenously temporarily enhanced the arterial pressure (AP) and the heart rate (HR). However, its role in the regulation of blood circulation is obscure. To study the properties of the molecule, its analogue was synthesized, in which proline in position 7 was substituted with glycine, and leucine in the position 2 with norleucine. Modified neuropeptide FF (FFm) also temporarily and in a dose-dependent manner increased the AP and HR; however, the equal degree of increase was reached at doses of FFm being 5-7 times lesser as compared with the natural peptide. The application of the FFm at hemorrhagic shock excluded mortality of animals during the experiment, considerably increased the degree of AP and HR restoration in the remaining experiments, and improved the survival of animals in 24 hours. It has been found that the level of antibodies to the fragment of hFF1 receptor in the serum is lower in spontaneously hypertensive rats SHR as compared with Wistar rats, but it is increased in patients of cardiological profile as compared with donors. The findings suggest involvement of neuropeptide FF in the regulation of blood circulation; however, the precise mechanisms remain to be determined.
Subject(s)
Blood Pressure/drug effects , Hypotension/prevention & control , Oligopeptides/pharmacology , Peptide Fragments/pharmacology , Shock, Hemorrhagic/prevention & control , Animals , Autoantibodies/blood , Female , Heart Rate/drug effects , Hypertension/immunology , Hypertension/metabolism , Hypotension/physiopathology , Male , Oligopeptides/blood , Oligopeptides/chemical synthesis , Oligopeptides/chemistry , Oligopeptides/therapeutic use , Peptide Fragments/chemical synthesis , Peptide Fragments/chemistry , Peptide Fragments/therapeutic use , Rats , Rats, Inbred SHR , Rats, Wistar , Receptors, Neuropeptide/immunology , Receptors, Neuropeptide/physiology , Shock, Hemorrhagic/physiopathologyABSTRACT
A new anti-inflammatory peptide X (Ingramon)--a fragment of C-terminal sequence (65-76) of MCP-1, was elaborated. In Boyden chamber assay, Ingramon inhibited migration of human monocytic cells THP-1 and peripheral blood monocytes. In vitro in blood plasma, Ingramon was stable for at least 24 hours. Ingramon was accumulated in sites of inflammation in rats, and impaired monocyte and granulocyte accumulation occurred in different models of inflammation in rodents and primates. Ingramon exerted no effect on the MCP-1-dependent externalisation of p2 integrins CD lib CD18 (Mac-1). We propose that Ingramon might interfere with MCP-1-heparin/heparan sulphate binding on cell surface. Ingramon reception by volunteers showed its safety. The clinical trials of Ingramon in patients with acute coronary syndromes and percutaneous coronary interventions, have been launched.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/chemical synthesis , Chemokine CCL2/antagonists & inhibitors , Drug Design , Inflammation/drug therapy , Peptide Fragments/chemical synthesis , Animals , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Carotid Arteries/drug effects , Carotid Arteries/pathology , Chemotaxis, Leukocyte/drug effects , Disease Models, Animal , Drug Stability , Humans , In Vitro Techniques , Inflammation/metabolism , Macaca fascicularis , Male , Mice , Mice, Inbred C57BL , Monocytes/cytology , Monocytes/drug effects , Peptide Fragments/chemistry , Peptide Fragments/pharmacology , Peptide Fragments/therapeutic use , Rats , Rats, Wistar , Tunica Intima/drug effectsABSTRACT
The Val-Val-Tyr-Pro-Asp bone marrow peptide (MP-5) and an analogue (MP-5-Lys) were synthesized. Fluorescent derivatives, Ftc-MP-5 and MP-5-Lys(Ftc), were prepared. The biological activity of MP-5 and MP-5-Lys was studied in vitro and in vivo. The MP-5 peptide caused 60-84% inhibition of growth of the following mouse cancers: lymphatic leukemia P-388, melanoma B-16, and cervical carcinoma CUC-5. These peptides also restored functional activity of T lymphocytes that was inhibited by metabolic products of the HL-60 leukemic cell line. MP-5-Lys(Ftc) was shown to preserve the functional properties of MP-5 toward T lymphocytes, but Ftc-MP-5 was practically inactive.
Subject(s)
Antineoplastic Agents/chemical synthesis , Fluorescent Dyes/chemical synthesis , Immunologic Factors/chemical synthesis , Oligopeptides/chemical synthesis , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Fluorescent Dyes/chemistry , Fluorescent Dyes/pharmacology , Immunologic Factors/chemistry , Immunologic Factors/pharmacology , Mice , Oligopeptides/chemistry , Oligopeptides/pharmacology , T-Lymphocytes/cytology , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , Xenograft Model Antitumor AssaysSubject(s)
Glomerular Filtration Rate/drug effects , Kidney/drug effects , Potassium/urine , Sodium/urine , Vasotocin/pharmacology , Animals , Body Weight/drug effects , Creatinine/urine , Diuresis/drug effects , Female , Injections, Intramuscular , Kidney/physiology , Macaca mulatta , Male , Rats , Rats, Wistar , Time Factors , Urination/drug effects , Vasotocin/administration & dosage , Vasotocin/analogs & derivatives , Water/administration & dosage , Water/metabolismABSTRACT
The retro-enantio analogue of peptide 66-77 of the chemokine MCP-1 and two hexapeptide fragments 66-71 and 72-77 of the C-terminal sequence of this protein were synthesized using the Fmoc strategy of solid phase peptide synthesis. The effect of the synthetic peptides upon the MCP-1-stimulated migration of THP-1 mononuclear cells was studied in vitro. The activity of the retro-enantio analogue was found to be comparable with that of the initial peptide 66-77: both peptides inhibit the migration of monocytes and granulocytes into inflammation zones of experimental animals.