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Lectinocytochemical study of rat stomach mucosa under the conditions of cyclooxygenase-1/-2 blockage and pretreatment witH H-Glu-Asp-Gly-OH

C. M. Nasadyuk1*, E. A. Sogomonyan2, A. M. Yashchenko2, A. Y. Sklyarov1

1Department of Biochemistry, Danylo Halytsky Lviv National Medical University, Ukraine;
2Department of Histology, Danylo Halytsky Lviv National Medical University, Ukraine;

Received: 22 December 2019; Accepted: 27 March 2020

Assessment of glycoconjugate expression on cell membranes using the lectin histochemistry technique may be a feasible approach for evaluating the functional state of the cell. The aim of this study was to evaluate carbohydrate determinants of rat stomach mucosa cell membranes under the conditions of COX-1/-2 blockage­ with indomethacin and pretreatment with the tripeptide H-Glu-Asp-Gly-OH. Male Wistar rats were divided into 3 groups (n = 6 per group): 1st group (control) received vehicle; 2nd – indomethacin (35 mg/kg); 3rd – H-Glu-Asp-Gly-OH (10 µg) 30 min before indomethacin. Rats were sacrificed 24 hours later. Gastric mucosa (GM) carbohydrate determinants were studied by lectin-peroxidase technique. The lectins panel included α-fucose- (LABA), syalo- (WGA, SNA), mannose- (Con A, LCA) and galactose-specific (HPA, PNA, SBA) lectins. Intensity of lectin-receptor reaction was scored: 0 – no reaction; 1 – weak; 2 – mild; and 3 – strong reaction. COX-1/2 blockage caused GM lesions, attenuated by H-Glu-Asp-Gly-OH. WGA and SNA showed the highest affinity to GM. Indomethacin decreased SNA-labeling of epitheliocytes and mucocytes and LABA-labeling of chief cells. H-Glu-Asp-Gly-OH reversed the glycosylation changes, caused by COX-1/COX-2 blockage only in regards to labeling of chief cells with LABA, epitheliocytes and mucocytes with LCA, mucocytes with SNA. Predominantly H-Glu-Asp-Gly-OH under COX-1/COX-2 blockage had an effect opposite to indomethacin alone but glycosylation changes under these conditions differed significantly also from the control. COX-1/COX-2 blockage causes alteration of glycosylation processes in rat GM, mainly reduction of NeuNAc(α2-6)DGal and α-Fuc content. H-Glu-Asp-Gly-OH under the conditions of COX-1/COX-2 blockage leads to more profound changes in GM lectin-binding pattern compared to the independent effect of indomethacin and to control.

Freezing influences, the exposure of IgG glycans in sera from multiple sclerosis patients

M. Bozhenko1, M. Boichuk1, G. Bila2, T. Nehrych1*, R. Bilyy2*

1Department of Neurology, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine;
2Department of Histology, Cytology and Embryology, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine;

Received: 08 January 2020; Accepted: 27 March 2020

N-glycan residues attached to Asn297 of the immunoglobulin IgG molecule are responsible for changing­ its structural conformation and are used as markers of many inflammatory diseases. Freezing stabilizes protein structure, while recent solution NMR data showed greatly altered IgG glycan mobility at different­ temperatures. The aim of the current work was to investigate whether freezing sera samples from multiple sclerosis (MS) patients and normal healthy donors (NHD) influences exposure of IgG glycans. The developed lectin immunosorbent assay was used to evaluate exposure of native IgG glycans with fucose-binding AAL lectin and sialic acid-binding SNA lectin. Sera samples were divided and either immediately frozen at -20 °C or stored at 4 °C. Lectin exposure was compared between 5 MS patient groups (n = 75) vs NHD (n = 23) and in paired samples with and without freezing. A significant increase in the exposure of fucose residues on IgG glycans in MS patients, compared to NHD, was observed. This increase was only observed if sera were frozen before analysis. The exposure of sialic acid was decreased in MS vs NHD samples after freezing sera samples. The exposure of core fucose residues and terminal sialic residues differed significantly in paired sera samples after freezing. Combined parameters of fucose and sialic acid exposure on native IgG glycans using frozen sera samples serve as a discriminative marker between MS and NHD. For AAL exposure, the discrimination of MS was characterized by AUROC of 0.906, sensitivity of 76.7% and specificity of 59.0% (P < 0.0001).

Hypertriglyceridemia is associated with long-term risk of cardiovascular events and specific comorbidity in very high-risk hypertensive patients

O. Ya. Korolyuk, O. M. Radchenko

Danylo Halytskyi Lviv National Medical University, Department of Internal Medicine No 2, Lviv, Ukraine;

Received: 19 December 2019; Accepted: 27 March 2020

Although hypertriglyceridemia (HTG) frequently occurs in hypertensive patients and may increase cardiovascular risk, the need for and manner of its reduction remain controversial. The objectives of this study were to compare lipid profiles, parameters of glucose homeostasis, comorbidity, and 5-year survival without cardiovascular events in very high-risk hypertensive (VHRH) patients with and without HTG, who received moderate intensity atorvastatin therapy. After initial assessment, 107 VHRH subjects were divided into two groups, i.e., without (n = 49) and with HTG (n = 58). During observation once annually patients were interviewed about prior hospitalizations with further screening for diabetes. Combined endpoint included­ hospitalization due acute myocardial infarction, decompensated heart failure, stroke or death. Survival was analyzed by Kaplan-Meier’s method. Nonparametric methods were used for statistical analysis. Higher median­ values of logarithmic value of triglycerides-to-HDL-cholesterol ratio, lipid accumulation product, fasting insulin, and HOMA index were observed in group 2 (< 0.002) that reflect predominance of small dense LDL particles, ectopic lipid deposition and insulin resistance. Patients with HTG more commonly had type 2 diabetes (58.6% vs 34.5%, including first-detected cases during initial assessments and observation, P = 0.02), liver steatosis (81.0% vs 55.1%, P = 0.006), and lithogenic gallbladder disorders (55.2% vs 34.7%, P=0.05). Women with HTG frequently had a history of hysterovariectomy (55.2% vs 19.0%, Р = 0.018). Despite long-term statin therapy, they often failed to reach recommended LDL-C targets and had worse survival due to significantly higher incidence of combined endpoint (39.6% vs 22.4%, P = 0.027). Further studies are necessary to find safe and effective strategy for secondary prevention in this population.

Homo Scientist can lead Homo Sapiens to the next evolutionary stage, to Homo Ingenious

Sandor G. Vari

International Research and Innovation in Medicine Program,
Cedars – Sinai Medical Center, Los Angeles, CA, USA
and Regional Cooperation in the Fields of Health, Science and Technology
Association (RECOO P HST Association), Budapest, Hungary

The Nobel laureates’ contributions to the study of carbohydrate metabolism and its regulation. A. Harden, H. Euler-Chelpin, C. F. Cori, G. T. Cori, E. Sutherland, L. F. Leloir, H. Krebs, F. Lipmann, P. Mitchell

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

Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kiyv;

Received: 13 November 2019; Accepted: 29 November 2019

Carbohydrate metabolism is a complex and multi-stage process. Many scientists (biochemists, physiologists, chemists) worked on deciphering this process, but only some of them were awarded the Nobel Prize. Thus, in the early XXth century, the work of A. Garden and H. Euler-Chelpin with yeast cells revealed that the conversion of carbohydrates (sugars) into end products occurs in living cells in several steps with the involvement of enzymes and that this conversion requires the presence of phosphoric acid residue. These studies were the beginning of exploring the chemical reactions (the reactions of intermediate metabolism) that are fundamentals for the vital functions of cells. In 1932, Hans Krebs discovered the ornithine cycle, a sequence of chemical reactions, in which the end product of nitrogen metabolism, urea, is formed in the liver of animals. The apogee of his research was the discovery of tri- and dicarboxylic acid cycle, which combines the oxidation of almost all organic compounds in living organisms. Fritz Lipmann, who in 1945 discovered coenzyme A and identified its role in the activation of organic compounds, furthered the works of H. Krebs. At the time, it became clear how inactive acetic acid and other organic acids are activated in living organisms to be oxidized in the tricarboxylic acid cycle. The great work was done by the spouses Gerty and Carl Cori,  and Bernardo Houssay, as well as their students and followers, in particular, Luis Leloir to clarify the mechanism of conversion (synthesis and breakdown) of glycogen in the liver and muscles. The peak of studying the carbohydrate metabolism was the research of Earl Sutherland who in 1958 revealed the regulation of the activity of enzymes involved in the conversion of chemical compounds (the example – phosphorylase), with the participation of the adenylate cyclase enzyme and c-AMP. The discovery of c-AMR established one of the fundamental principles of almost all vital processes. And the culmination of research on carbohydrate metabolism was the unrivaled works of Peter Mitchell, who studied the course of biochemical reactions in cells relative to certain intracellular “landmarks” and who formulated the chemiosmotic theory of oxidative phosphorylation that underlies bioenergetics.

Free radical oxidation in liver mitochondria of tumor-bearing rats and its correction by essential lipophilic nutrients

O. V. Ketsa, M. M. Marchenko

Fedkovich Chernivtsy National University, Ukraine;

Received: 21 May 2019; Accepted: 29 November 2019

The role of free radical oxidation in the increase of mitochondrial membranes permeability in organs which are not involved in oncogenesis and the development of the methods for preventing mitochondria dysfunction remain topical problems. In this work, the interconnection of lipid peroxidation (LPO) in liver mitochondrial fraction with the processes of mitochondrial swelling and cytochrome с release to the cytosol under separate and combined administration of ω-3 polyunsaturated fatty acids (PUFAs) and retinol acetate (vitamin A acetate) to rats with transplanted Guerin’s carcinoma was studied.  During the intensive tumor growth (14 days) the increase of superoxide radical generation and the content of primary (triene conjugates, TC), secondary (ketodienes and coupled trienes, CD+CT) and terminal (Schiff bases) lipid peroxidation products­ in the mitochondrial fraction of tumor-bearing rats was detected, which contributed to the mitochondrial swelling and cytochrome с release to the cytosol. Separate administration of ω-3 PUFAs to tumor-bearing rats decreased both free radical processes in mitochondrial fraction and mitochondrial swelling. Separate administration of retinol acetate in a high dose (3000 IU/kg of body weight) intensified free radical processes in the mitochondrial fraction of tumor-bearing rats, while administration of retinol acetate in a physiological dose (30 IU/kg of body weight) did not lead to changes compared to tumor-bearing rats that did not receive the drug. The prooxidant effects of retinoid were partially eliminated in the case of combined administration with ω-3 PUFA.

Cytotoxic activity of the cluster rhenium compound with β-alanine ligands

K. V. Polokhina1, D. E. Kytova1, A. V. Shtemenko1, N. I. Shtemenko1,2

1Ukrainian State University of Chemical Technology, Dnipro, Ukraine;
2Dnipro University of Technology, Ukraine;

Received: 25 February 2019; Accepted: 29 November 2019

Earlier we have shown that cluster rhenium compounds not only inhibited tumor growth in vivo but also supported the antioxidant state of experimental animals. Further investigation of new dirhenium(III) and cluster rhenium compounds in human leukemic cells is of great importance. The aim of the recent work was to investigate the cytotoxic activity of the new cluster rhenium compound with β-alanine ligands [Re2Cl6(C3H7NO2)2]·1.5H2O (I) in the solutions and nanoliposomes alone and together with cisplatin in Jurkat cells. It was shown that I in solution had cytotoxicity close to cisplatin (LC50 = 2.06·10-6 M). The administration of the rhenium-platinum system with І showed  increased cytotoxic activity, especially high when both components of the system were in the mixed liposomes together (LC50 = 4.93·10-10 M). The new dirhenium dicarboxylate complex with zwitterionic amino acid ligands possesses an appreciable cytotoxic and proapoptotic activity against leukemic cells, especially in combination with cisplatin, guiding the search for novel active rhenium compounds and development of improved regimens for combined chemotherapy based on combination of rhenium-platinum compounds.

Indexes of citrulline metabolism in rat liver under the toxic injury against the background of alimentary protein deficiency

H. P. Kopylchuk, I. M. Nykolaichuk, I. S. Lylyk

Yuriy Fedkovych Chernivtsi National University, Ukraine;
Institute of Biology, Chemistry and Bioresources, Chernivtsi, Ukraine;

Received: 29 May 2019; Accepted: 29 November 2019

It is known that citrulline is converted into arginine in the series of metabolic transformations. Results of our previous studies showed that acetaminophen-induced toxic injury on the background of the alimentary deprivation of protein is accompanied by a decrease in arginine level in rat hepatocytes, but citrulline liver metabolism at these conditions remains incompletely clear. In this work, the content of citrulline in the rat liver mitochondrial and cytosolic fractions and the activity of citrulline-degrading enzymes – argininosuccinate synthase and argininosuccinate lyase were investigated. It was found that in the mitochondrial fraction a maximal reduction of the citrulline levels occurred after administration of acetaminophen toxic doses regardless of the protein amount in the ration, while in the cytosolic fraction the alimentary protein deficiency was a key factor in decreasing the activity of argininosuccinate synthase and arginino-succinate lyase. The data obtained indicated the disturbances of the urea cycle functioning and explained the   decrease of L-arginine level in hepatocytes in conditions of acetaminophen-induced toxic injury against the background alimentary protein deficiency.

Biochemical indicators of green photosynthetic bacteria Chlorobium limicola response to Cu(2+) action

T. B. Sehin1, S. O. Hnatush1, O. D. Maslovska1,
A. A. Halushka1, Y. H. Zaritska2

1Ivan Franko National University of Lviv, Ukraine;
2State Scientific-Research Control Institute of Veterinary
Medicinal Products and Feed Additives, Lviv, Ukraine;

Received: 11 March 2019; Accepted: 29 November 2019

Photolithotrophic sulfur bacteria are involved in  biota functioning and have  a biotechnological potential for bioremediation of contaminated environment, but the mechanisms of xenobiotics, in particular of heavy metal ions damaging action and the pathways of photolithotrophic bacteria adaptation  under these conditions have not been established. In this work, the biochemical indicators of green photosynthetic bacteria Chlorobium limicola response to Cu ions were studied. C. limicola cells were incubated during one hour in buffer containing copper (II) sulfate in 0.05–0.5 mM concentrations and grown for 8 days in GSB medium. The content of Cu2+ in cells was estimated by atomic absorption spectroscopy. The activity of enzymes of antioxidant defense, photosynthetic pigments and glutathione content, indexes of lipids unsaturation and membrane viscosity as markers of  membrane fluidity were estimated. It was shown that the response of green photosynthetic bacteria C. limicola to Cu2+ action varied  depending on cations concentration. Under the influence of metal salt at 0.05 mM concentration, the activity of antioxidant enzymes, GSH/GSSG ratio, the content of photosynthetic pigments and membrane fluidity indexes were higher as compared with control. Under the increase of copper (II) sulfate concentration to 0.25 mM, the activity of antioxidant  enzymes was lower compared to the response of the cells under the influence of 0.05 mM copper (II) and the GSSG content was increased.  Under the influence of 0.5 mM copper (II) the indexes of membrane fluidity did not differ from the control, but superoxide dismutase and peroxidase activity inhibition and  the further decrease of GSH/GSSG ratio were observed followed by the highest Cu2+ cations accumulation in cells and significant decrease of  bacteria biomass growth.

Production and physicochemical characterization of xanthan gum by native lactose consuming isolates of Xanthomonas citri subsp. citri

R. Moravej1, S. M. Alavi2, M. Azin3, A. H. Salmanian2

1Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran;
2Department of Plant Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran;
3Department of Biotechnology, Iranian Research Organization for Science and Technology (IROST), Tehran, Iran;

Received: 30 September 2019; Accepted: 29 November 2019

Xanthan is a biopolymer produced by Xanthomonas bacteria which is widely used in many industries such as food and oil. In this work, three Xanthomonas strains (X. citri/NIGEB-88, X. citri/NIGEB-386 and X. citri/NIGEB-K37) were used to evaluate their industrial potential to produce xanthan gum in whey medium. Bacteria growth rate, viscosity, biomass, dry weigh of produced xanthan and β-galactosidase activity were studied during the fermentation process and the presence of β-galactosidase genes was assessed by PCR technique. Strain NIGEB-386 had the best ability to utilize lactose in the whey medium. The highest amount of xanthan production and viscosity were 22.7 g/l and 2066.6 mPa·s, respectively. The presence of six β-galactosidase genes in strains NIGEB-386 and NIGEB-K37 was confirmed. The pyruvate and acetyl contents in xanthan gum were 2.1 and 0.29 %, respectively. Fourier-transform infrared spectroscopy analysis determined the position of the functional groups in the structure of the fermentation product. In whey medium, the performance of both NIGEB-386 and NIGEB-K37 strains were better than the X. campestris. The findings showed that Xanthomonas citri/NIGEB-386 is suitable for industrial production of xanthan using cheese whey as a low-cost medium.