The structural analogue of apelin-12 prevents energy disorders in the heart in experimental type 1 diabetes mellitus.

Type 1 diabetes mellitus (T1DM) is the most severe form of diabetes, which is characterized by absolute insulin deficiency induced by the destruction of pancreatic beta cells. The aim of this study was to evaluate the effect of a structural analogue of apelin-12 ((NαMe)Arg-Pro-Arg-Leu-Ser-His-Lys-Gly-Pro-Nle-Pro-Phe-OH, metilin) on hyperglycemia, mitochondrial (MCh) respiration in permeabilized cardiac left ventricular (LV) fibers, the myocardial energy state, and cardiomyocyte membranes damage in a model of streptozotocin (STZ) diabetes in rats. Metilin was prepared by solid-phase synthesis using the Fmoc strategy and purified using HPLC. Four groups of animals were used: initial state (IS); control (C), diabetic control (D) and diabetic animals additionally treated with metilin (DM). The following parameters have been studied: blood glucose, MCh respiration in LV fibers, the content of cardiac ATP, ADP, AMP, phosphocreatine (PCr) and creatine (Cr), the activity of creatine kinase-MB (CK-MB) and lactate dehydrogenase (LDH) in blood plasma. Administration of metilin to STZ-treated rats decreased blood glucose, increased state 3 oxygen consumption, the respiratory control ratio in MCh of permeabilized LV fibers, and increased the functional coupling of mitochondrial CK (mt-CK) to oxidative phosphorylation compared with these parameters in group D. In STZ-treated animals metilin administration caused an increase in the PCr content and prevention of the loss of total creatine (ΣCr=PCr+Cr) in the diabetic hearts, as well as restoration of the PCr/ATP ratio in the myocardium and a decrease in the activity of CK-MB and LDH in plasma to initial values. Thus, metilin prevented energy disorders disturbances in cardiomyocytes of animals with experimental T1DM.

[The anti-ischemic and antioxidant activity of the pharmacological agonist of galanin receptor GalR2 and carnosine in in vitro and in vivo model systems]. / Protivoishemicheskaia i antioksidantnaia aktivnost' farmakologicheskogo agonista retseptora galanina GalR2 i karnozina v model'nykh sistemakh in vitro i in vivo.

Antioxidant and anti-ischemic properties of the pharmacological agonist of galanin receptor GalR2 WTLNSAGYLLGPßAH (Gal) and its C-terminal fragment, dipeptide carnosine (ßAH), were studied in the model of regional ischemia and reperfusion of the rat heart in vivo in the dose range of 0.5-5.0 mg/kg and Cu²âº-induced free radical oxidation of low density lipoproteins (LDL) of human plasma in vitro for peptide concentrations of 0.01 mM and 0.1 mM. Gal was obtained by automatic solid phase synthesis using the Fmoc methodology; its structure was characterized by 1H-NMR spectroscopy and MALDI-TOF mass spectrometry. Intravenous administration of the optimal dose of Gal (1 mg/kg) to rats after ischemia was more effective than carnosine in reducing of the myocardial infarct size and the activity of creatine kinase-MB and lactate dehydrogenase in blood plasma at the end of reperfusion. It also improved the metabolic state of the reperfused myocardium and reduced the formation of peroxidation products during reperfusion. Gal reduced more effectively the formation of adducts of hydroxyl radicals in the interstitium of the area at risk (AAR) of the rat heart than carnosine. Carnosine at a dose of 1 mg/kg more effectively increased the activity of catalase and glutathione peroxidase in the AAR by the end of reperfusion compared to Gal. In a model of Cu²âº-initiated oxidation of human plasma LDL 0.1 mM carnosine demonstrated a significantly more pronounced reduction in the formation of lipid radicals compared to Gal. The results show that Gal can be considered as a promising agent that reduces myocardial injury during reperfusion and oxidative stress.

The Mechanisms of Cardiac Protection Using a Synthetic Agonist of Galanin Receptors during Chronic Administration of Doxorubicin.

The use of the anticancer drug doxorubicin (Dox) is limited by its cardiotoxic effect. The aim of this work was to study the effect of a new synthetic agonist of the galanin receptor GalR1-3 [ßAla14, His15]-galanine (2-15) (G) on the metabolism, antioxidant enzyme activity, and cardiac function in rats with cardiomyopathy (CM) caused by chronic administration of Dox. Coadministration of peptide G and Dox significantly increased the fractional shortening (FS) and ejection fraction (EF) by an average of 30 ± 4% compared with the indices in the Dox group. The reduced severity of cardiac dysfunction under the action of G was accompanied by a 2.5-fold decrease in the activity of creatine kinase-MB (CK-MB) in blood plasma. The protective mechanism of the action of peptide G is caused by a reduced lipid peroxidation (LP) that is due to the increased activity of Cu,Zn superoxide dismutase (Cu,Zn-SOD) and glutathione peroxidase (GSH-Px) in the damaged heart. Administration of peptide G significantly increased the adenine nucleotide pool (ΣAH), ATP content, and the levels of phosphocreatine (PCr) and total creatine (ΣCr) in the damaged myocardium. It also reduced lactate accumulation relative to its content in the Dox group. The better energy supply of cardiomyocytes after treatment with peptide G prevented the accumulation of cytotoxic ammonia and disruption in the metabolism of the key myocardial amino acids (glutamic acid (Glu), aspartic acid (Asp), and alanine (Ala)). Peptide G significantly improved the morphological parameters of the heart in rats treated with Dox. The results show promise in using peptide G to efficiently correct functional, morphological, and metabolic damage to the heart caused by anthracycline chemotherapy.

[Cardiometabolic efficacy and toxicological evaluation of a pharmacological galanin receptor agonist]. / Kardiometabolicheskaia éffektivnost' i toksikologicheskaia kharakteristika farmakologicheskogo agonista retseptorov galanina.

The goal of this study was to examine effects of a novel galanin receptor agonist GalR1-3 [bAla14, His15]-galanine 2-15 (G), obtained by automatic solid-phase synthesis, on the metabolic state of the area at risk and the size of acute myocardial infarction (MI) in rats in vivo and evaluate its toxicity in BALB /c mice. In anesthetized rats, regional ischemia was simulated by coronary artery occlusion and then coronary blood flow was restored. The peptide G was administered intravenously (i.v.) with a bolus after a period of regional ischemia in the dose range of 0.25-3.0 mg/kg. The sizes of MI and the activities of creatine kinase-MB (СK-MB) and lactate dehydrogenase (LDH) in blood plasma were estimated. The effect of administration of the optimal dose of G (1.0 mg/kg) on myocardial content of adenine nucleotides (AN), phosphocreatine (PCr), creatine (Cr) and lactate was studied. I.v. administration of G to rats at a dose of 1.0 mg/kg slightly affected hemodynamic parameters, but reduced MI size by 40% and decreased plasma LDH and CK-MB activity by the end of reperfusion compared to control. These effects were accompanied by a significant improvement in energy state of area at risk (AAR) - an increase in myocardial content of ATP, åAN, PCr and åCr, and combined with a decrease in myocardial lactate level compared with the control. Toxicity of peptide G was studied with a single intraperitoneal injection of 0.5-3.0% solution of the peptide substance to mice. The absence of signs of intoxication and death of animals after G injection in the maximum possible dose did not allow determining the value of the average lethal dose. The results indicate therapeutic potential of the peptide G for preventing myocardial ischemia and reperfusion injury and feasibility for further study of its pharmacological properties and mechanisms of action.

[Protective action of a modified fragment of galanine in rats with doxorubicin-induced heart failure]. / Zashchitnoe deistvie modifitsirovannogo fragmenta galanina pri serdechnoi nedostatochnosti u krys, vyzvannoi doksorubitsinom.

The use of the anticancer drug doxorubicin (Dox) is limited due to its cardiotoxic effect. Using the method of automatic solid-phase peptide synthesis, we obtained a synthetic agonist of galanin receptors GalR1-3 [RAla14, His15]-galanine (2-15) (G), exhibiting cardioprotective properties. It was purified by high performance liquid chromatography (HPLC). The homogeneity and structure of the peptide was confirmed by HPLC, 1H-NMR spectroscopy and mass spectroscopy. The purpose of this study was to study the effect of G on the metabolism and cardiac function of rats with chronic heart failure (CHF) caused by Dox. Experiments were performed using male Wistar rats weighing 280-300 g. The control group of animals (C) was intraperitoneally treated with saline for 8 weeks; the doxorubicin group (D) of rats was intraperitoneally treated with Doх; the group of Doх + peptide G (D+G) received intraperitoneally injections of Doх and subcutaneously injections of peptide G; the peptide G group (G) was subcutaneously treated with G. At the beginning and at the end of the study, the concentration of thiobarbituric acid reactive substances (TBARS) and the activity of creatine kinase-MB (CK-MB) were determined in blood plasma; the animals were weighed, and cardiac function was assessed using echocardiography. At the end of the experiments, the hearts were used for determination of metabolites and assessment of oxidative phosphorylation in mitochondria. After 8-week treatment, animals of group D were characterized by severe heart failure, the lack of weight gain and an increase in plasma TBARS concentration and CK-MB activity. These disorders were accompanied by a decrease in the content of myocardial high-energy phosphates, a reduction inmitochondrial respiratory parameters, accumulation of lactate and glucose in the heart, and disturbances in the metabolism of alanine and glutamic and aspartic acids. Coadministration of G and Dox prevented the increase in plasma CK-MB activity and significantly reduced the plasma TBARS concentration. At the end of the experiments animals of group D+G had higher myocardial energy state and the respiratory control index of mitochondria than animals of group D, there was a decrease in anaerobic glycolysis and no changes in the amino acid content compared to the control. The peptide G significantly improved the parameters of cardiac function and caused weight gain in animals of group D+G in comparison with these parameters in group D. The obtained results demonstrate the ability of a novel agonist of galanin receptors GalR1-3 to attenuate Dox-indiced cardiotoxicity.

[Metilin Improves Сardiac Function in Rabbits With Chronic Heart Failure].

AIM: To study effects of intravenous infusion of a cardioprotective drug metilin, developed at the "National Medical Research Center of Cardiology" on indices of cardiac function in rabbits in vivo after prolonged administration of doxorubicin. MATERIALS AND METHODS: Animals of the experimental group were intravenously injected with doxorubicin (2 mg / kg once a week) for 8 weeks, animals of the control group received the same volume of saline. Myocardial damage was characterized by an increase in concentration of malondialdehyde (MDA), troponin (TnI) and MB-fraction of creatine kinase (CK-MB) in venous blood and by disturbances in the left ventricle (LV) structure at morphological examination. Metilin effects on cardiac function were assessed by echocardiography and LV catheterization by the Millar catheter tip pressure transducer. RESULTS: Doxorubicin administration led to a decrease of the body mass of animals, an increase of the plasma concentration of cardiac markers CK-MB and TnI, lipid peroxidation (LPO) product MDA in venous blood, and pronounced disturbances in the structure of LV fibers and microvessels. At the same time, a significant decrease of myocardial contractility indices was observed. Manifestations of this decrease were increase of the end-diastolic and end-systolic dimensions (EDD and ESD, respectively), and decreases in the shortening fraction and ejection fraction (SF and EF, respectively) compared to baseline values. These changes indicated development of chronic heart failure (CHF) in animals of the experimental group. Against this background, intravenous infusion of metilin significantly increased SF and EF, but did not affect the heart rate. Beneficial effects of metilin on the indices of cardiac contractility and relaxation were maintained after the infusion was stopped. Noteworthy, metilin exerted greater influence on cardiac function of rabbits with CHF compared to control animals that did not receive doxorubicin. CONCLUSION: The obtained results indicate the potential of metilin to reduce LV dysfunction during chemotherapy with doxorubicin.

[Analysis of Urine Proteome in Patients With Postinfarction Cardiosclerosis Combined With Hypertensive Disease for Assessing Endothelial Dysfunction].

In our study urine protein composition of 18 healthy volunteers was compared with that of 18 patients with ischemic heart disease and concomitant hypertension. Liquid chromatography-mass-spectrometry (LC-MS) analysis of the second fraction of morning urine was carried out using nano-line high performance liquid chromatograph and hybrid mass spectrometer. The analysis revealed 23 proteins expressed in the endothelium, according to the information contained in the database Bgee, and 49 proteins, with direct functional link with the processes in the endothelium in the reconstruction of associative networks using ANDSystem program. Comparison of urine proteome of healthy people and patients with postinfarction cardiosclerosis revealed proteins specific for patients with cardiovascular disease. Thus, proteins vitronectin, syndecan-4, a histidine rich glycoprotein, endothelial protein C receptor, colony stimulating factor, cathepsin D and sekretogranin-1 may be considered as potential markers for cardiovascular diseases. Further research in this area should be conducted for clinical and experimental verification of these hypotheses.

[Cytochrome C content and mitochondrial activity in rat's cardiac and skeletal muscle cells in the course of 3-day gravitational unloading].

Purpose of the investigation was to determine the cytochrome c content and associated gene expression in cardiomyocytes and m. soleus fibers obtained from rats following 6, 12, 18, 24 and 72 hours of tail suspension. Cellular respiration was measured using Sax's polarography as a parameter reflecting the mitochondrial activity. In cardiomyocytes, cytochrome c in the protein membrane-mitochondrial fraction grew 34.6% (p < 0.05) after 18 hours of suspension relative the control. Cycs and gapdh expression was not altered, while parameters of the cardiomyocyte cellular respiration were increased. In m. soleus fibers, cytochrome c made a fall following 24-hr suspension. mRNA cycs was found reduced significantly after 6 hours and cellular respiration intensity declined reliably after 72 hours of gravitational unloading.

[Identification proteins cardiovascular system in urine healthy subjects during "dry" immersion].

We analyzed the urine proteome in 14 healthy volunteers who underwent a 5-day dry immersion using proteomic analysis methods and bioinformatics approach. We identified 9 proteins related to cardiovascular system. It was shown that 5-day dry immersion modifies urine proteomic profile indicating renal, endocrine, circulatory and metabolic changes. Most of these changes are characterized by both a very rapid onset and very rapid restoration within return to normal conditions.

[Shifts in metabolism and its regulation under the effect of spaceflight factors].

The review deals with the results of studying the adaptive changes in metabolism and its neuroendocrine regulation in humans and animals under the effect of spaceflight factors and ground-based simulation of the gravitational unloading. The majority of the investigations were concerned with the water-electrolyte and mineral turnover, as well as protein, lipid and carbohydrates metabolism. Biochemical measurements of the body liquids (blood, urine and saliva) before, in and after space flight or in ground simulation experiments were used as indictors of the status of sympathoadrenal, hypothalamo-pituitary-adrenal and other systems involved in systemic regulation of metabolism, and also strength of stress-reaction to adversities. The authors generalized data on the interrelation and interaction of the neuroendocrine and psychophysiological status both in the real and simulated conditions of space flight.

Structure and functional characteristics of rat's left ventricle cardiomyocytes under antiorthostatic suspension of various duration and subsequent reloading.

The goal of the research was to identify the structural and functional characteristics of the rat's left ventricle under antiorthostatic suspension within 1, 3, 7 and 14 days, and subsequent 3 and 7-day reloading after a 14-day suspension. The transversal stiffness of the cardiomyocyte has been determined by the atomic force microscopy, cell respiration--by polarography and proteins content--by Western blotting. Stiffness of the cortical cytoskeleton increases as soon as one day after the suspension and increases up to the 14th day, and starts decreasing during reloading, reaching the control level after 7 days. The stiffness of the contractile apparatus and the intensity of cell respiration also increases. The content of non-muscle isoforms of actin in the cytoplasmic fraction of proteins does not change during the whole experiment, as does not the beta-actin content in the membrane fraction. The content of gamma-actin in the membrane fraction correlates with the change in the transversal stiffness of the cortical cytoskeleton. Increased content of alpha-actinin-1 and alpha-actinin-4 in the membrane fraction of proteins during the suspension is consistent with increased gamma-actin content there. The opposite direction of change of alpha-actinin-1 and alpha-actinin-4 content suggests their involvement into the signal pathways.

[Parameters of fibers cell respiration and desmin content in rat soleus muscle at early stages of gravitational unloading].

The aim of the work was to study the parameters of fibers cell respiration and desmin content in Wistar rat soleus muscle after 1, 3, 7 and 14 days of gravitational unloading. Gravitational unloading was simulated by antiorthostatic hindlimb suspension. The parameters of cell respiration were determined using the polarography, and desmin content was assessed by means of Western blotting. The results showed that the intensity of cell respiration is reduced after three days of gravitational unloading, reaches a minimum level after seven days and slightly increases by the fourteenth day of hindlimb unloading, as well as the content of desmin, which, however, to the fourteenth day returns to the control level. Taking into account that mitochondrial function depends on the state of cytoskeleton the data allow us to assume that early reduction of the intensity of cell respiration under unloading could be caused by degradation of the protein desmin that determines intracellular localization of mitochondria.

[Desmin content and cell breathing in rats' m. soleus fibers after 3- and 7-day recovery from 14-day suspension].

Cell breathing and relative desmin content were investigated in m. soleus fibers of rats after 3 and 7 days of recovery from 14-day suspension according to the Morey - Holton modification of the Ilyin - Novikov procedure. The cell breathing parameters were determined with the help of polarography; desmin content was measured using the western-blot technique. The results evidence that cell breathing intensity subsided during 14 days of gravitational unloading, reached minimum after 3 days of recovery and regained baseline values after 7 days of recovery. Post suspension desmin content did not differ from control values, made a significant drop in 3 days and returned to baseline values in 7 days of recovery. These data suggest an interdependence between cell breathing and desmin content in m. soleus fibers of rats as during gravitation unloading, so in the period of recovery.

[Changes in cell respiration of postural muscle fibers under long-term gravitational unloading after dietary succinate supplementation].

The intensity of cell respiration of the rat m. soleus, m. gastrocnemius c.m. and tibialis anterior fibers during 35-day gravitational unloading, with the addition of succinate in the diet at a dosage rate of 50 mg per 1 kg animal weight has been investigated. The gravitational unloading was modeled by antiorthostatic hindlimb suspension. The intensity of cell respiration was estimated by polarography. It was shown that the rate of oxygen consumption by soleus and gastrocnemius fibers on endogenous and exogenous substrates and with the addition of ADP decreases after the discharge. This may be associated with the transition to the glycolytic energy path due to a decrease in the EMG-activity. At the same time, the respiration rate after the addition of exogenous substrates in soleus fibers did not increase, indicating a disturbance in the function of the NCCR-section of the respiratory chain and more pronounced changes in the structure of muscle fibers. In tibialis anterior fibers, no changes in oxygen consumption velocity were observed. The introduction of succinate to the diet of rats makes it possible to prevent the negative effects of hypokinesia, although it reduces the basal level of intensity of cell respiration.

Effects of long-term microgravitation exposure on cell respiration of the rat musculus soleus fibers.

Cell respiration of the m. soleus fibers was studied in Wistar rats treated with succinic acid and exposed to microgravitation for 35 days. The results indicated that respiration rates during utilization of endogenous and exogenous substrates and the maximum respiration rate decreased in animals subjected to microgravitation without succinate treatment. The respiration rate during utilization of exogenous substrate did not increase in comparison with that on endogenous substrates. Succinic acid prevented the decrease in respiration rate on endogenous substrates and the maximum respiration rate. On the other hand, the respiration rate on exogenous substrates was reduced in vivarium control rats receiving succinate in comparison with intact control group. That could indicate changed efficiency of complex I of the respiratory chain due to reciprocal regulation of the tricarbonic acid cycle.

[Effects of creatine phosphokinase competitive inhibitor on system and tissue energy metabolism in rats in the norm and during unloading].

In an experiment with rats system and intracellular energy metabolism was assessed by cell reactions to chronic injection of beta-guanidine propionic acid (beta-GPA) stimulating AMP-dependent protein kinase (AMPK). Suspension was shown to inhibit the succinate dehydrogenase (SDH) activity, reduce glycogen in both types of muscle fibers, and stimulate the activity of alpha-glycerophosphate dehydrogenase (alpha-GPDH) in fast fibers. Supplementing the rat chow with beta-GPA did not modify these parameters during suspension; however, the blood urea level increased considerably in the suspended and control rats. In the controls, beta-GPA as well as suspension, stimulates growth of the aspartate aminotranspherase activity (AST) in blood. Yet, the suspension and beta-GPA injection had no additive effect. Moreover, their effects were opposite in rats subjected to suspension + beta-GPA. Glucose concentration was observed to become lowered in blood of resting rats treated with beta-GPA. This effect can be associated with a more intensive insulin-dependent glucose transport to muscles. The additional glucose, because of increased demand by fibers, underwent to oxidation and did not replenish the intracellular carbohydrate deposits These data suggest energy metabolism shifting toward activation of the processes of disintegration of substrates for energy production due to a sharp growth of energy demand.

Protective effect of energy metabolism regulators in alteration of gravitation load under experimental conditions.

Mitochondrial substrate-based preparations corrected disorders, caused by long-term exposure to abnormal gravitation vector in head-down tilt (hanging) test in rats. The preparations produced systemic and polyorgan protective effects consisting in correction of the blood prooxidant/antioxidant balance, energy metabolism in musculus soleus, and minimization of morphological changes in the liver and kidneys.

[DNA analysis of retroposon-like genetic LINE elements in blood plasma of rats exposed to radio-diapason electromagnetic waves]. / Analiz DNK retrotranspozonpodobnykh geneticheskikh élementov LINE v plazme krovi krys, podvergshikhsia vozdeistviiu élektromagnitnykh voli radiodiapazona.

The elevation of LINE-elements' DNA level was revealed in blood plasma of rats exposed to electromagnetic waves. The amount of full-size 5'-containing LINE-elements copies was increased especially. Connection of this effect with retrotransposon activation and genetic instability condition of organism development is supposed.