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The contribution of the Nobel Prize laureates to the development of dynamic biochemistry and bioenergetics. E. Buchner, A. Kossel, R. Willstätter, O. Meyerhof, A. Hill, O. Warburg, A. Szent-Györgyi

V. M. Danilova, R. P. Vynogradova, S. V. Komisarenko

Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kiyv;
e-mail: valdan@biochem.kiev.ua

Received: 29 November 2018; Accepted: 13 December 2018

Thanks to the great discoveries of the Nobel laureates of the first half of the 20th century – E. Buchner, A. Kossel, R. Willstätter, O. Meyerhof, A. Hill, O. Warburg, A. Szent-Györgyi, we have gained a deep understanding of the mechanisms of organic­ substances conversion and oxidation in living­ organisms­. This article gives an analysis of the research activity of these distinguished scientists, who, through decoding the main ways of conver­ting carbohydrates and energy in living organisms, laid the foundations of dynamic biochemistry and bioener­getics (one of the branches of biochemical science).

Levels of angiogenic regulators and MMP-2, -9 activities in Martorell ulcer: a case report

O. M. Petrenko1, A. A. Tykhomyrov2

1Bogomolets National Medical University, Kyiv, Ukraine;
2Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv, Ukraine;
e-mail: artem_tykhomyrov@ukr.net

Received: 27 July 2018; Accepted: 13 December 2018

Martorell hypertensive ischemic leg ulcers (HYTILU) represent a unique form of lower extremity non-healing ulcers that develop in association with poorly controlled high blood pressure. The present study was performed in order to assess levels of protein regulators of angiogenesis (vascular endothelial growth factor, or VEGF, and angiostatins) and to evaluate activities of matrix metalloproteinases (MMPs) (gelatinases MMP-2 and -9) in wound cutaneous tissue in the case of patient with 2-years HYTILU history. VEGF and angiostatin levels were analyzed by Western blot, MMP activities were evaluated by gelatin zymography. We report here for the first time that wound tissue in HYTILU is characterized with increased levels of VEGF (by 75 folds vs. histologically normal tissue, P < 0.01) and dramatic overproduction of angiostatin levels, which are undetectable in healthy cutaneous tissue. Approximately 10-fold elevation in MMP-2 and -9 activities is observed in wound tissue as compared with uninjured cutaneous tissue. Obtained results indicate that increased production of angiogenic inhibitors, angiostatins, may counteract VEGF-induced pro-angiogenic signaling, and together with MMP overactivation, contributes to failed healing of ischemic ulcer. Further extended studies are needed to clarify how changes of angiogenic profile and imbalance of proteolytic activities in non-healing Martorell ulcers can be considered during their management procedures to improve efficacy of surgery debridement and/or skin grafting.

Evaluation of biochemical indicators in blood plasma of rats with tetracycline-induced hepatosis and their correction by milk phospholipids

V. A. Gryshchenko1, V. V. Musiychuk1, V. O. Chernyshenko2,
O. V. Gornytska2, T. M. Platonova2

1National University of Life and Environmental Sciences of Ukraine, Kyiv;
2Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
е-mail: viktoriya_004@ukr.net

Received: 13 July 2018; Accepted: 13 December  2018

Tetracycline is a drug with direct cytotoxic action on the liver, and therefore it is widely used in pharmaceutical studies of therapeutic effectiveness of hepatoprotective preparations. The aim of the present work was to determine the biochemical indicators in blood plasma of rats with tetracycline-induced hepatosis and correction properties of milk phospholipids under tetracycline-induced hepatosis in rats. To achieve this, Wistar rats were administered 250 mg/kg of 4% tetracycline hydrochloride suspension once a day intragastrically. As the corrective therapy, 1% solution of BAS “FLP-MD” was administered in liposomal form based on milk phospholipids. Under modeled steatohepatitis, significant destructive changes were observed in the cell membranes of hepatocytes in experimental rats. It was confirmed by higher activity of transaminase (in particular, activity of АSТ increased 4 times, that of ALT 1.7 times and the AST/ALT ratio was increased 2.4 times in blood plasma). The synthesis of clotting factors in livers of animals with hepatosis was inhibited. The content of fibrinogen in blood plasma decreased by 21%, factor II (prothrombin) by 27.8%, Xa-factor by 27.9%, and protein C by 40.6%. The animals also had hypochromic anemia, azotemia and bilirubinemia. The calcium-phosphor metabolism and hyperkalemia were observed. The liposomal BAS “FLP-MD” based on milk phospholipids diminished harmful effects of tetracycline, in particular supporting blood coagulation factors’ level restoration, and also by the activity of transaminases. According to the results, it may be used in prophylactics and pharmaceutical correction of steatohepatitis.

Cellular fatty acid composition of Aeromonas genus – destructor of aromatic xenobiotics

T. V. Gudzenko, O. G. Gorshkova, N. V. Korotaieva,
O. V. Voliuvach, А. М. Ostapchuk, V. O. Іvanytsia

Оdesa I. I. Mechnikov National University, Ukraine;
e-mail: tgudzenko@ukr.net

Received: 17 September 2018; Accepted: 13 December 2018

The aim of this study was a determination of the fatty acid composition of cellular lipids and identification of the strains, isolated from the wastewater of pharmaceutical production, – the destructor of aromatic xenobiotics. The phenotypic characteristics and cellular fatty acid (FA) composition confirmed the strain belonging to the Aeromonas ichthiosmia with the similarity index of library data MIDI Sherlock – 0.564. Analysis of the cellular FA composition of the strain Aeromonas ichthiosmia ONU552 was carried out using the MIDI Sherlock microorganism identification system based on the gas chromatograph Agilent 7890. Chromatographic analysis showed that the fatty acid profile of the strain Aeromonas ichthiosmia ONU552 contains 26 fatty acids with the total number of carbon atoms from 10 to 18. 85.27% of saturated and unsaturated fatty acids had unbranched structure. The total content of unsaturated fatty acids – 16:1 w7c/16:1 w6c, 18:1 w7c, 16:1 w7c alcohol, 17:1 w8c, 17:1 w6c, 16:1 w5c, was 50% of the total fatty acid pool. Less than 1.5% branched fatty acids were predominantly in the iso form: 13:0 iso (0.20%); 15:0 iso (0.97%); 17:1 iso w9c (1.35%), 17:0 iso (1.49%); in the anteiso form, only one acid 17:0 (0.27%) was identified. It was shown that the characteris­tic of the fatty acid composition of the strain Aeromonas ichthiosmia ONU552 – the destructor of aromatic xenobio­tics, was the presence of hydroxyacids 12:0 3OH, 15:0 3OH, 15:0 iso 3OH and dominance of hexadecanoic (16:0) and hexadecenoic (16:1 w7c/16:1 w6c) of fatty acids.

Role of AP-1 transcriptional factor in development of oxidative and nitrosative stress in periodontal tissues during systemic inflammatory response

A. M. Yelins’ka, O. Ye. Akimov, V. O. Kostenko

Ukrainian Medical Stomatological Academy, Poltava, Ukraine;
e-mail: riseofrevan5@gmail.com

Received: 04 August 2018; Accepted: 13 December 2018

Chronic systemic inflammatory response syndrome (SIRS) underlies many diseases (sepsis, atherosclerosis, diabetes mellitus). According to research data of recent years the key role in the development of SIRS is played by the activation of various nuclear transcription factors. The work was aimed at studying the role of such transcription factor as activator protein 1 (AP-1) in the development of oxidative and nitrosative stress in soft periodontal tissues during chronic systemic inflammatory response (SIRS). The experiment was carried out on 24 the Wistar rats. We induced SIRS by bacterial lipopolysaccharide of Salmonella typhi (0.4 μg/kg) intraperitoneal injection. We studied changes in the functioning of the nitric oxide (NO) cycle, the production of superoxide anion radical (O2•-) and the activity of antioxidant enzymes in soft periodontal tissues homogenate. We used SR11302 as an Ap-1 inhibitor (15 mg/kg) for 2 months. We established that during the SIRS modeling, the activity of antioxidant enzymes in soft periodontal tissues decreased with a simultaneous increase in the production of O2•-. SIRS elevated the production of NO by inducible NO-synthase (iNOS) and nitrite reductases. The nonoxidative cleavage of L-arginine under this condition was also increased. The concentration of peroxynitrite (ONOO) was shown to be elevated more than 2-fold. The inhibition of AP-1 by SR11302 normalized the functional state of the NO cycle, reduced O2•- production and restored the activity of antioxidant enzymes. In this way, under SIRS conditions, “vicious circle” of ONOO formation is formed. SIRS in soft periodontal tissues poses a threat of oxidative and nitrosative stress development. Usage of AP-1 activation inhibitor SR11302 breaks “vicious circle” of ONOO formation.

Diagnostic significance of biochemical indicators of liver fibrogenesis in adolescents with obesity

O. V. Buznytska

Kharkіv Medical Academy of Postgraduate Education,
V. N. Karazin Kharkіv National University, Ukraine
e-mail: ebuznickaa@ukr.net; missbuzelena@gmail.com

Received: 27 September 2018; Accepted: 13 December 2018

Non-alcoholic fatty liver disease occurs in most obese people, the main pathway of which is the process of fibrogenesis. The aim of this work was to determine the potential biomarkers for early diagnosis of liver fibrogenesis in adolescents with obesity. The levels of liver fibrosis markers, such as fibronectin, collagen type IV, N-terminal propeptides and C-terminal telopeptides of type I collagen, were assessed with the use of IFA method in serum of 226 patients with obesity aged 8-18 years. A significant increase in levels of type IV collagen and fibronectin was observed in children with obesity (P < 0.05). As diagnostic criteria for fibrogenesis and fibrolysis, the levels of N-terminal propeptides and C-terminal telopeptides of type I collagen, respectively, were determined. The serum level of N-terminal propeptides of type I collagen significantly exceeds the normal values in all children with obesity compared to the control group (P < 0.05). The biochemical markers (type IV collagen, fibronectin, N-terminal propeptides and C-terminal telopeptides of type I collagen) were proven to have high diagnostic informative value in the early diagnosis of liver fibrogenesis in obese adolescents. It was shown that the signs of fibrosis in non-alcoholic fatty liver disease already occur at the stage of steatosis.

HBD-2 interactions with erythrocyte membranes in vitro

M. V. Makarenko1, D. O. Semeniuk2, I. O. Starenka2, A. P. Pogribna3,
I. V. Sokol4, L. I. Martinova1, D. O. Govsieiev4

1Institute of Postgraduate Education,
Bogomolets National Medical University, Kyiv Ukraine;
2ESC Institute of Biology and Medicine,
Taras Shevchenko National University of Kyiv, Ukraine;
e-mail: dmyt.semenyuk@gmail.com;
3Institute of Molecular Biology and Genetics, NAS of Ukraine, Kyiv;
4Kyiv City Maternity Hospital No 5

Received: 24 September  2018; Accepted: 13 December 2018

HBD-2 – a member of β-defensin family of antimicrobial peptides – is known to permeabilize cell membranes of susceptible cells, but the mechanism of such interactions is poorly understood. In our study, we have used a hemolytic model to explore the kinetic properties of HBD-2 interactions with membranes of human erythrocytes. We ran hemolytic assays with a wide range of both HBD-2 and erythrocyte concentrations, as well as varying pH values, incubation times, and osmotic strengths; each in the presence or the absence of inhibitory substances such as proteins and salts. The results show that HBD-2 cell membrane permeabilization is both dose- and time-dependent (with plateau effect observed in each case), and inversely dependent on erythrocyte concentration. HBD-2 interactions with cell membranes highly depend on pH value and the presen­ce of inhibitors but are not affected by tested osmotic strength range. Our findings suggest that interactions of HBD-2 with cell membranes are mainly electrostatic in nature and are limited by released cell content. We have developed a speculative model of such interaction based on our results.

Intermittent fasting causes metabolic stress and leucopenia in young mice

O. M. Sorochynska1, M. M. Bayliak1, Y. V. Vasylyk1,
O. V. Kuzniak1, I. Z. Drohomyretska1, A. Ya. Klonovskyi1,
J. M. Storey2, K. B. Storey2, V. I. Lushchak1

1Vasyl Stefanyk Precarpathian National University, Ivano-Frankivsk, Ukraine;
2Institute of Biochemistry, Carleton University, Ottawa, Canada;
e-mail: lushchak@pu.if.ua

Received: 29 October 2018; Accepted: 13 December 2018

Overweight and obesity became the worldwide epidemic resulting from overeating especially when a so-called Western diet rich in carbohydrates and fats is used. It is widely accepted that limitation of food consumption could help to withstand such state of adult organism, but information about younger groups is contradictory. The present study was undertaken to characterize the effects of intermittent fasting, using an every other day (EOD) fasting/feeding protocol, on hematological parameters and biochemical blood plasma indices in young mice from one to two months old. It was shown that intermittently fasted mice were characterized by a reduced body weight, reduced total number of blood leucocytes, lower glucose and lactate levels and higher activity of alanine aminotransferase and aspartate aminotransferase in blood plasma as compared with the age-matched control mice. To gain the same mass EOD animals needed to eat more food than ad libitum fed animals. These differences may probably be explained by a need to expend certain resources to combat stress induced by intermittent fasting. Our data showed that EOD feeding at a young age may negatively influence young mammals.

Influence of sulfite on ATPase activity of coupling factor CF(1) isolated from spinach chloroplasts

O. B. Onoiko, A. P. Khomochkin, O. K. Zolotareva

M. G. Kholodhy Institute of Botany, National Academy of Sciences of Ukraine, Kyiv;
e-mail: membrana@ukr.net

Received: 25 October 2018; Accepted: 13 December 2018

The present study of sulfite participation in the regulation of the chloroplast ATP-synthase complex can help to understand the mechanisms of acid rains toxic effects on the plant photosynthetic apparatus. The aim of the work was to study the effect of sulfite on Ca2+– and Mg2+-dependent ATP hydrolysis catalysed by coupling factor CF1, exposed to preliminary short-term acid treatment. CF1 was isolated from spinach chloroplasts (Spinacia oleracea L.). The latent ATPase activity of CF1 was activated by sodium sulfite addition into incubation media. The rate of ATP hydrolysis was determined by the release of inorganic phosphate. In the presence of 25 mM exogenous sulfite  the Mg2+-dependent hydrolytic activity of CF1 was increased 7-fold and Ca2+-dependent activity 3-fold compared to the control. Carbonic anhydrase inhibitors acetazolamide or ethoxyzolamide eliminated sulfite-induced stimulation of ATP hydrolysis. The sulfite stimulating effect on Ca2+– and Mg2+-ATPase activity was increased dramatically after incubation (5 min) of isolated CF1 in a medium with pH 3.5 and its  subsequent transition to the alkali medium (pH 8.0). In this case, 1 mM sulfite-induced a 10-fold acceleration of ATP hydrolysis. Carbonic anhydrase specific inhibitors (50 μM) removed the sulfite effect. These data suggest that sulfite was able to replace bicarbonate in the CF1 structure after the release of bound HCO3 during acid treatment.

Spin-orbit coupling effects in O(2) activation by cofactor-independent 2,4-dioxygenase

B. F. Minaev1, R. R. Valiev2

1Bohdan Khmelnytsky National University of Cherkasy, Ukraine;
e-mail: bfmin43@ukr.net;
2Royal Institute of Technology, Stockholm, Sweden;
e-mail: valievrashid@mail.ru

Received: 22 August 2018; Accepted: 13 December 2018

The O2 (dioxygen) is paramagnetic molecule with two non-paired electron spins and triplet ground state (S = 1) while majority of organic molecules are diamagnetic species; they have all electron spins paired and the singlet ground state with the total spin S = 0. Oxygenases catalyze a concerted insertion of the triplet dioxygen into organic (diamagnetic) molecules in a strictly spin-forbidden process and this puzzle is not solved so far in modern enzymology. Many oxidases and oxygenases utilize the π-conjugated organic cofactor (like flavins, pterins) in a singlet ground state and reaction of cofactor with O2 is still spin-forbidden. It is clear that the protein environment in the enzyme active-site “helps” in some way to overcome spin prohibition, but this environment is definitely diamagnetic and the spin-puzzle still exists. Some oxidases and oxygenases use paramagnetic metal ions as a cofactor; in this case the spin prohibition is formally reduced. In recent years, a numbers of oxidative enzymes are discovered which do not contain any cofactor. In the present work, we considered a rather popular cofactor-free bacterial 2,4-dioxygenase and its oxygenolytic reactions with 2-n-alkyl-3-hydroxy-4(1H)-quinolones (AHQ’s). We presented results of quantum-chemical calculations of intermediate diradical proposed recently for direct reaction of dioxygen with AHQ substrate and made conclusion about the mechanism of spin-catalysis.