Department of Blood Circulation

Версія за 15:28, 12 січня 2017

DEPARTMENT OF CIRCULATION PHYSIOLOGY

Background

Department of Circulation Physiology was organized in 1960 (head - prof. M.I.Gurevich) on the basis of the Department of Circulation and Respiration Physiology (head-academician M.M.Gorev), created in 1953 as a structural unit of the Bogomolets Institute of Physiology. Since 1986, the head of the department has been corresponding member of NAS of Ukraine V.F.Sagach.

The Structure Of The Department

In the department (2016), two doctors of sciences (VF Sagach and L.M.Shapoval), 3 Senior Research officers (O.V.Akopova, O.I.Bondarenko, N.A.Strutynska) 4 research officers (Yu.V.Hoshovska, NO Dorofeyeva, Yu.P.Korkach, S.V.Chorna), 5 junior research officers (K.O.Drachuk, R.A.Dobrovolska, O. V.Dmytrenko, O.M.Semenykhina, B.S.Kop'yak), 3 engineers (T.A.Ilyina, I.Yu.Okhay, F.V.Dobrovolskyy) and four post graduate students are working.

Defended Theses

2010 - S.O.Talanov "Changes in the functional state of the cardiovascular system of rats with chronic nigrostriatal dopamine deficit " for the degree of candidate of biological sciences, specialty 03.00.13- Human and Animal Physiology

2012 - Yu.V.Hoshovska "The role of mitochondrial uncoupling proteins UCP2 and UCP3 in the functioning of the heart" for the degree of candidate of biological sciences, specialty 03.00.13- Human and Animal Physiology

2013 - N.O. Dorofeyeva "The role of hydrogen sulphide in changing the functioning of the heart in hypertensive animals" for the degree of candidate of biological sciences Pathological physiology spetsialnistyu14.03.04-

2014 - O.M. Semenykhina "The role of hydrogen sulphide in the regulation of vascular smooth muscle tone and cardioprotective mechanisms of action" for the degree of candidate of biological sciences, specialty 03.00.13- Human and Animal Physiology

2016 - O.V.Akopova "The role of mitochondrial transmembrane pores in the regulation of Ca 2+ exchange, energetic and free-radical processes in the mitochondria," for the degree of doctor of biological sciences, specialty 03.00.04- Biochemistry

State Awards

1994 - OO Moybenko., VF Sagach – O.O. Bogomolets Award of National Academy of Sciences of Ukraine for the monograph "Immunogenicity of disorders of the cardiovascular system" -Kyiv, Naukova Dumka, 1992, 202 p.

1996 - V.F.Sagach, L.M.Shapoval, A.I.Solovyov, O.V.Bazylyuk, A.V.Dmytriyeva, M.M.Tkachenko,.S.M.Marchenko received the State Prize in Science and Technology for a series of scientific works "THE ROLE OF ENDOTHELIUM AND BIOLOGICALLY ACTIVE SUBSTANCES OF ENDOTHELIAL ORIGIN IN THE REGULATION OF BLOOD CIRCULATION AND HEART ACTIVITY”

2003 - V.F.Sagach and M.M.Tkachenko among other authors received the State Prize in Science and Technology for a series of scientific papers "STUDY OF FUNDAMENTAL MECHANISMS OF ACTION OF NITRIC OXIDE IN THE CARDIOVASCULAR SYSTEM AS THE BASIS OF PATHOGENETIC TREATMENT OF ITS DISEASES"

Prizes and scholarships to young scientists

2012 - Hoshovs'ka Y., JP Korkach, Chorna SV received the Prize of the President of Ukraine for young scientists for a series of scientific works "THE PERMEABILITY OF MITOCHONDRIAL MEMBRANES AS A NEW TARGET CARDIOPROTECTION”

2014- Dorofeeva NO, Chorna SV received Presidential Scholarship for Young Scientists

2015 - Hoshovs'ka Y.V. received scholarship for young scientists from National Academy of Sciences of Ukraine "

State Patents From Government

1. Zhaboyedov GD, Lavrik NS, Kotsyuruba AV Kurilina AI, JP Korkach. Utility model patent №49934 from 11.05.2010.
2. Shymanska TV, Hoshovs'ka J., V. Sagach. Method to prevent reperfusion cardiac dysfunction / Utility model patent , number 77864 from 02.25.13.
3. Prylutskyy YI, Shapoval LM, Sagach VF, Dimitrenko AV, Radchenko NV "The use of single-walled carbon nanotubes in antihypertentional therapy." Patent for an invention, No. 20120821 from 05/07/2013
4. Talanov SA, Patalakh II, VF Sagach. Method of activation of endothelium in isolated vascular fragments for modeling endothelium-dependent processes // Utility model patent № 79466. - 25.04.2013.
5. Talanov SA, Patalakh II, VF Sagach The device for application of isolated fragments of vessels in the research of functions of activated endothelium // Utility model patent № 79918. - 13.05.2013.
6. Talanov SA, Kop'yak BS, Hoshovs'ka UV, Sagach VF Method for improving modeling hemiparkinsonizmu in rats. Utility model patent №87583 from 2014
7. Talanov CA Patalakh II, Sagach VF. Method for activation of endothelium in isolated vascular fragment in simulation of endothelium- dependent processes and device for their modeling. Patent for an invention, №104363 from 01.27.2014
8. Strutynskyy RB, Strutynska NA, Sagach VF Moybenko OO Yahupolskyy YL "Method for preventing calcium-induced mitochondrial pore opening in the heart» Utility model paten№ 12691; 10.04.2015.

Participation In The Programs

1. Joint project of the State Fund for Fundamental Research of Ukraine and Byelorussian Republican Foundation for Basic Research. “The role of nitric oxide in the reactions of the cardiovascular system under stress and hyperglycemia" (2007-2008).
2. Joint project of the State Fund for Fundamental Research of Ukraine and Byelorussian Republican Foundation for Basic Research 2009-2010. The project "Role of nitric oxide in the reactions of the cardiovascular system in animals adapted to exercise"
3. Combined program of NAS of Ukraine "New medical-biological problems and human environment" 2006 -2009. Project “ Inhibitors of mitochondrial permeability transition pore opening as a means of correction of the cardiovascular system functioning "
4. Combined program of interdisciplinary research of NAS of Ukraine "Fundamental principles of molecular and cellular biotechnology, 2010-2014" The role of ion-exchange membrane processes in the regulation of cyclosporine-sensitive pore and free radical reactions in the mitochondria. "
5. Project: “Endothelial Electrical Signaling in a murine model of Atherosclerosis. Identification of potential cannabinoid targets for treatment of the disease”. Funded by Swiss Science Foundation (SNSF).
6. Departmental scientific program of NAS of Ukraine 2006-2009. "The study of mechanisms of regulation of functional systems under normal and pathological states" theme "Studying the role of changes in the functional state of mitochondria in the reactivity of the cardiovascular system"
7. Departmental scientific program of NAS of Ukraine 2010-2013 "Study of the role of mitochondria in reactions of the cardiovascular system in different functional states of the body"
8. Departmental scientific program of NAS of Ukraine 2014-2018 — "Study of the mechanisms of regulation of functional systems under normal and pathological states " theme of the department: "Research on the role of signaling sulfur compounds in reactions of the cardiovascular system of rats in different states of the body" 
9. Departmental scientific program of NAS of Ukraine "Fundamental principles of Genomics and Proteomics", the theme of the Institute "Studies of molecular genetic mechanisms of physiological and pathophysiological processes and development of methods for its correction", theme of the depertment "Study of genetic mechanisms of changes in the functioning of mitochondria during prolonged exercise in old animals" 2006-2010
10. Departmental research NAS Ukraine program "Functional genomics, proteomics and metabolomics in Systemic Biology; Topic: "Genetic and epigenetic mechanisms of regulation of visceral functions in normal and pathological conditions"; theme "Study of genetic and epigenetic mechanisms of energy supply of adaptation of the cardiovascular system to stress of different genesis" 2012-2016

Major Methods Used In The Department

The experiments are performed on living rats, isolated segments of the vessels and the heart of rats, as well as on the endothelium and the mitochondria from isolated rat hearts.

Methods Used In Experiments

Physiological methods: registration of cardiohemodynamics; retrograde perfusion of coronary artery by the Langendorf method, recording of oxygen tension in solutions, registration of active myogenic responses of isolated vascular smooth muscle preparations, registration of electrical responses of the endothelium, injections of substances in populations of brain neurons using stereotaxic coordinates;

biophysical methods: recording of membrane potential of endothelial cells (cultivated and intact) by perforated patch -clamp under current clamp mode;

biochemical methods: determination of hydrogen sulfide, activities of hydrogen sulfide producing enzymes, activities of nitric oxide synthases (neuronal, endothelial and inducible), the levels of indicators of oxidative and nitrosative stress (nitrate, nitrite, nitrosothiols, peroxide hydrogen, superoxide radicals, hydroxyl radicals, diene conjugates, malondialdehyde) in homogenates of heart, aorta, and medulla oblongata, and in suspension of mitochondria, process of mitochondrial respiration and oxidative phosphorylation in the heart mitochondria of rats, isolation of mitochondria from rat heart, spectrophotometric registration of mitochondrial swelling and the mitochondrial factor release; molecular methods: PCR in real time; Western-blot analysis to determine the expression of uncoupling proteins (UCP2 and UCP3) in the mitochondria of the heart

models used: experimental Parkinson’s disease, experimental diabetes mellitus in rats.

The Main Achievements of the Department

In recent years, efforts of the scientific staff have been aimed at studying the role of endogenous hydrogen sulfide in various elements of the cardiovascular system (heart, blood vessels, cardiovascular neurons, mitochondria), and interaction between hydrogen sulfide and nitric oxide, another powerful transmitter in the body.

It has been revealed that hydrogen sulfide is actively involved in the heterometric regulation of cardiac function. Modulation of hydrogen sulfide synthesis affects the level of mitochondrial membrane potential and the respiration rate. Following blockade of enzymes for hydrogen sulfide synthesis, either cystathionine-γ-lyase or 3-mercaptopyruvate sulfur transferase, or simultaneous inhibition of both enzymes, the mitochondrial membrane potential in the heart is reduced significantly. The maximum reduction of the latter is observed after inhibiting 3-mercaptopyruvate sulfur transferase by aspartate. The mitochondrial respiration rate increases significantly after combined blockade of both cytosolic and mitochondrial enzymes. Changes in mitochondrial membrane potential coincide with the physiological response of the heart to the volume load. Blockade of hydrogen sulfide synthesis in the mitochondria by aspartate, accompanied by a significant decrease in membrane potential, results in the inhibition of cardiac response to volume load compared with that in control conditions. The latter reflects a reduction in the functional reserves of the heart under inhibited hydrogen sulfide synthesis.

It has been shown that hydrogen sulfide can carry neuroprotective effect by reducing 6-hydroxydopamine-induced degeneration of dopaminergic neurons in the midbrain (model of Parkinson's disease). Moreover, hydrogen sulfide exerts positive effect on the contractile function of the heart in rats, reducing the sensitivity of mitochondrial permeability transition pores to calcium, normalizing mitochondrial respiration and increasing the degree of coupling of oxidation and phosphorylation.

In the medulla oblongata of rats, a significant content of endogenous hydrogen sulfide has been detected. In hypertension, the level of hydrogen sulfide in the rat medulla oblongata is much lower that that in the normotensive rats. After preliminary inhibition of cystathionine -β-synthase (which is expressed primarily in neurons of the brain), or an inhibition of 3-mercaptopyruvate sulfur transferase (which is thought to be the mitochondrial enzyme), a significant weakening of the hemodynamic effects is observed. Changes in hemodynamics after inhibiting cystathionine-γ-lyase are neglegible. These data suggest that the effects of L-cysteine in the medulla oblongata are implemented mainly through the activation of cystathionine -β-synthase and 3-mercaptopyruvate sulfur transferase. It has been also shown that an inhibition of neuronal NO-synthase reduces the effects of L-cysteine introduced in the medullary nuclei, testifying to the interaction between hydrogen sulfide and nitric oxide systems in the cardiovascular control. We were the first to show that an administration of single-walled carbon nanotubes in the medullary nuclei results in hypotensive responses in both normotensive and spontaneously hypertensive rats. Single-walled carbon nanotubes influence the neural medullary cardiovascular control in rats following their intramedullary or peripheral (intravenous, intraperitoneal, subcutaneous) administration. Endogenous hydrogen sulfide and nitric oxide contribute to the implementation of the hypotensive effect of single-walled carbon nanotubes.

In the study on adult and old rats, it has been revealed that blockade of cystathionine-γ-lyase (CSE), enzyme for de novo synthesis of hydrogen sulfide, restores acetylcholine-induced relaxation of the aorta smooth muscles, suppressed in old rats. This effect is removed by inhibiting of nitric oxide (NO) synthesis. The levels of intramitochondrial pools of H2S and NO2-, as well as cNOS activity in the heart decrease with age. An introduction of propargyl glycine largely restores the pools of H2S and NO2- and stimulates reduced activity of cNOS. Thus, stimulation of H2S and NO synthesis by propargyl glycine results in recovering of suppressed endothelium-dependent relaxation of the aorta smooth muscles in old rats and improving the cardiac function via the stimulation of H2S and NO synthesis. NaHS, introduced in vivo, inhibits oxidative and nitrosative stress, restores cNOS coupling and increases the constitutive synthesis of NO.

It has been found that an activation of free radical processes, particularly during aging, ischemia-reperfusion of the myocardium, and ischemic pre-conditioning results in enhanced expression of UCP2 and UCP3 genes in the heart tissues. Blockade these uncoupling proteins reduces negative effects of oxidative stress, indicating the participation of these proteins in the formation of cardioprotective myocardial response.

The features of calcium accumulation by isolated heart mitochondria from adult rats under physiological conditions and in case of mitochondrial dysfunction due to their load by high concentrations of calcium ions have been determined. The possibility of using flow cytometry to study mitochondrial calcium homeostasis in isolated cardiomyocytes was demonstrated. The dependence of calcium accumulation on incubation temperature, availability of Mg2+ -ATP complex substrate and succinate has been shown. It was found that mitochondria from isolated heart have limited ability to store calcium. The data on the impact of exogenous hydrogen sulfide and inhibitors of its synthesis on the accumulation of Ca2+ by the mitochondria in cardiomyocytes of adult rats have been obtained. It was found that an accumulation of calcium in isolated mitochondria depends on the concentration of NaHS. Inhibition of 3-mercaptopyruvate sulfur transferase, mitochondrial enzyme, does not affect the accumulation of calcium, which may testify to mitochondrial regulation of calcium-transporting system by cytosolic hydrogen sulfide, not mitochondrial one.

It has been found that endogenous hydrogen sulfide pools are reduced in the heart mitochondria in aging. This process is normalized under the action of pyridoxal-5-phosphate, in other words, hydrogen sulfide content increases probably due to an activation of H2S-synthesizing enzymes. It has been revealed that degree of swelling of the heart mitochondria from old rats after the introduction of Ca2+ inductor pyridoxal-5-phosphate lowers significantly compared to the heart mitochondria without inductor, which may indicate an increase in resistance due to reduced mitochondrial membrane permeability in the presence of Ca2+ inductor. The activation of hydrogen sulfide biosynthesis results in reduced sensitivity of mitochondrial permeability transition pore to its inducer calcium in the heart mitochondria from old rats due to increased threshold concentration, which causes swelling of organelles. The experimental results suggest that an increase in the content of hydrogen sulfide through the activation of its biosynthesis in the body is effective in inhibiting mitochondrial permeability transition pore, and it can be useful to correct mitochondrial dysfunction in various pathological conditions of the cardiovascular system in aging.

In biochemical studies, it has been shown that hydrogen sulfide donor NaHS at physiological concentrations inhibits calcium-induced mitochondrial permeability transition pore opening, indicating the protective effect on the pore formation in rat heart. An increase in the activity of arginase, nitrate reductase, and enzymes for constitutive nitric oxide synthesis has been detected in the mitochondria from rat heart after either the introduction of L-cysteine, substrate of hydrogen sulfide synthesis, or its antagonist propargyl glycine, and their joint actions that may be a manifestation of hormesis. Temporal correlation between increased pools of two powerful vasodilators, namely sulfur- containing and nitrogen- containing, was found in the blood plasma and the heart of rats. With aging, endogenous pools of hydrogen sulfide are reduced in the blood plasma and the heart mitochondria, at that oxidative and nitrosative stress is considerably weakened not only following a single injection of L-cysteine, but also propargyl glycine, an inhibitor of H2S synthesis. Reducing of endogenous pools of hydrogen sulfide in the heart mitochondria from old rats leads to an increase in peroxynitrite generation that is metabolized to generate *OH and * NO2-. In the heart mitochondria of old rats, propargyl glycine stimulates the activity of constitutive NO-synthase (cNOS), while the activity of inducible isoform (iNOS) is inhibited by propargyl glycine and L-cysteine, and their combined administration has an additive effect.

It has been shown that omega 3 polyunsaturated fatty acids (linoleic acid) cause hyperpolarization of endothelial cells due to the stimulation of calcium-dependent potassium channels of large conductance. This effect is receptor-independent and preserved in the membrane patch when you add omega 3 fatty acids to the inner surface of the membrane. Reduction of cholesterol level in the membrane of endothelial cells almost completely inhibits this effect, indicating the regulatory role of cholesterol in the effect of omega 3 fatty acids on calcium-dependent potassium channels of large conductivity. A similar effect is observed when adding anandamide, an agonist of cannabinoid receptors types I and II. Results can extend modern understanding of the mechanisms of action of omega 3 polyunsaturated fatty acids and agonists of cannabinoid receptors.

The effects of Ca2+ -induced mitochondrial permeability transition pore (MPTP) opening in the states of submaximal conductivity on the formation of ROS in the mitochondria of the rat brain and the liver were shown, as well as a linear dependence of ROS generation (JR) on the rate of Ca2+ -stimulated respiration (JO2) and exponential dependence of JR on membrane potential (ΔΨm). Based on the obtained results, it was concluded that in the conditions of submaximal conductivity, the role of MPTP in ROS generation is determined by its contribution to both the respiration rate and the modulation of ΔΨm. The data obtained indicate potential-dependent mechanism of regulating ROS generation in mitochondria of the liver and the brain of rats by MPTP opening in states of submaximal conductivity.

The levels of expression of mRNA cystathionine-γ-lyase, cystathionine-β-synthase and 3-mercaptopyruvate sulfur transferase were studied by PCR in real time method in the heart from adult and old rats in control and after the modulation of endogenous production of hydrogen sulfide. It has been revealed that the level of mRNA 3-MPST is significantly higher than mRNA CSE in the heart of adult and old animals. An increase in the expression of mRNA of two enzymes was found in the heart of old rats compared to adult animals. Moderate short-term inhibition of CSE together with a single injection of the amino acid L-cysteine is accompanied by increased expression of CSE mRNA in the heart of only adult rats, while in old animals substantial decline was observed. 3- MPST mRNA expression significantly increases (6.3-fold (p <0.05) in adults and 2.6-fold (p <0.05) in old animals), compared to control rats. Thus, under conditions of moderate short-term inhibition of CSE with following administration of amino acid L-cysteine, changes in expression of H2S-synthesizing enzymes occur, which are aimed at activating the endogenous synthesis of hydrogen sulfide as an important regulatory factor in the cardiovascular system under physiological and pathological conditions.