Tag Archives: glutathione

Oxidative/antioxidant balance and matrix metalloproteinases level in the knee cartilage of rats under experimental osteoarthritis and probiotic administration

O. Korotkyi*, K. Dvorshchenko, L. Kot,
T. Vovk, M. Tymoshenko, L. Ostapchenko

ESC “Institute of Biology and Medicine”, Taras Shevchenko National University of Kyiv, Ukraine;
*e-mail: korotkyi@gmail.com

Received: 28 June 2020; Accepted: 13 November 2020

The aim of this work was to investigate the effect of the poly-strain probiotic on oxidative-antioxidant balance and the level of matrix metalloproteinases (MMPs) in rat knee cartilage under experimental osteoarthritis. Osteoarthritis was induced by a single injection of monoiodoacetate into the knee joint of rats. Probiotic was administered daily for 14 days. Knee cartilages homogenate was used to evaluate  the content of reactive oxygen species (superoxide anion and hydrogen peroxide), products of lipids peroxidation (diene conjugates, TBA-active compound, Shiff bases), to determine superoxide dismutase and catalase activity and activity of glutathione-dependent  antioxidant enzymes, the level of reduced and oxidized glutathione. The level of MMPs -1, -2, -3, -8 expression was estimated by ELISA. Osteoarthritis was found to cause a significant increase in the reactive oxygen species level, lipid peroxidation products content, superoxide dismutase and catalase activity, level of all studied MMPs, and also depletion of glutathione-dependent antioxidant system and the decrease in the ratio between reduced and oxidized glutathione.The administration of the probiotic was followed by the tendency for the restoration of the parameters to the values of the control group. Thus, the administration of the probiotic to rats with osteoarthritis may be considered as an anti-inflammatory and antioxidant agent for further clinical trials.

Effect of probiotic composition on oxidative/antioxidant balance in blood of rats under experimental osteoarthritis

O. Korotkyi, K. Dvorshchenko, A. Vovk, A. Dranitsina, M. Tymoshenko, L. Kot, L. Ostapchenko

ESC “Institute of Biology and Medicine”, Taras Shevchenko National University of Kyiv, Ukraine;
e-mail: korotky@ukr.net

Received: 20 June 2019; Accepted: 18 October 2019

Osteoarthritis (OA) is a widespread pathology of the musculoskeletal system. OA may associate with a wide range of disorders, that lead to the development of various strategies on how to prevent and treat the disease. Recent studies discussed interactions between the microbiome and a wide range of pathologies, including OA. In this study, we investigated the effect of probiotic cultures on oxidative/antioxidant balance in blood of rats during OA. Experimental OA was induced by a single injection of sodium monoiodoacetate into the knee joint. A probiotic composition (Multiprobiotic Simbiter®) was administered by peroral catheter once per day for 14 days. We investigated the next parameters: expression of Nos2 gene in the blood, superoxide dismutase activity, catalase activity, glutathione peroxidase activity, glutathione transferase activity, glutathione reductase activity, contents of superoxide, hydrogen peroxide, TBA-reactive compounds, oxidized and reduced glutathione in the serum of the blood. Monoiodoacetate-induced OA caused significant changes on oxidative/antioxidant balance in the blood of rats: increasing of the contents of superoxide anion radical, hydrogen peroxide, thiobarbituric acid-reactive compounds, oxidized glutathione, upregulating of Nos2 expression, increasing of catalase activity; conversely, superoxide dismutase activity, glutathione peroxidize activity, glutathione transferase activity, glutathione reductase activity, the content of reduced glutathione were significantly decreased, compared to control group. Administration of probiotics to animals with OA led to positive changes in the studied parameters approaching the values of control group (some of them were statistically significant).

Glutathione influence on energy metabolism in rat liver mitochondria under experimental nephropathy

Ye. O. Ferenchuk, I. V. Gerush

Higher State Educational Establishment of Ukraine “Bukovinian State Medical University”, Chernivtsi;
e-mail: yelena_f@ukr.net

Received: 17 October 2018; Accepted: 14 March 2019

Mitochondrial oxidative damage and disorders of energy metabolism contribute to a wide range of pathologies and disease progression. In our work, the effect of glutathione on the activity of respiratory chain enzymes and the content of free SH-groups in rat liver mitochondria was examined with the use of folic acid-induced nephropathy model. Mitochondria were isolated by differential centrifugation, NADH-dehydrogenase, succinate dehydrogenase, cytochrome oxidase and H+-ATPase activity were determined. The activity of these enzymes and the content of the free SH-groups in the liver were shown to be decreased under conditions of nephropathy, evidently due to the intensification of the free radical processes. The introduction of glutathione increased the content of SH-groups and the activity of the Complexes II and V enzymes of mitochondrial respiratory chain but did not change the activity of cytochrome oxidase in mitochondria isolated from the liver of rats under experimental nephropathy. The results obtained demonstrate a positive effect of glutathione on mitochondrial succinate dehydrogenase and H+-ATPase activity normalization in the liver of rats with nephropathy. These findings may help to extend the understanding of mitochondrial energy metabolism under development of kidney diseases.

Cloning and functional analysis of the GSH1/MET1 gene complementing cysteine and glutathione auxotrophy of the methylotrophic yeast Hansenula polymorpha

V. M. Ubiyvovk1, O.V. Blazhenko1, M. Zimmermann2, M. J. Sohn3,4, H. A. Kang3,4

1Institute of Cell Biology, National Academy of Sciences of Ukraine, Lviv;
2Institute of Biology IV- Microbiology and Genetics RWTH, Aachen, Germany;
3Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea;
4Department of Life Science, Chung-Ang University, Seoul, Korea;
e-mail: Oleksandra.Blazhenko@googlemail.com

The Hansenula polymorpha GSH1/MET1 gene was cloned by complementation of glutathione-dependent growth of H. polymorpha gsh1 mutant isolated previously as N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) resistant and cadmium ion sensitive clone. The H. polymorpha GSH1 gene was capable of restoring cadmium ion resistance, MNNG sensitivity, normal glutathione level and cell proliferation on minimal media without addition of cysteine or glutathione, when introduced into the gsh1 mutant cells. It was shown that the H. polymorpha GSH1 gene has homology to the Saccharomyces cerevisiae MET1 gene encoding S-adenosyl-L-methionine uroporphyrinogen III transmethylase, responsible for the biosynthesis of sulfite reductase cofactor, sirohaem. The H. polymorpha GSH1/MET1 gene deletion cassette (Hpgsh1/met1::ScLEU2) was constructed and corresponding null mutants were isolated. Crossing data of the point gsh1 and null gsh1/met1 mutants demonstrated that both alleles were located to the same gene. The null gsh1/met1 mutant showed total growth restoration on minimal media supplemented with cysteine or glutathione as a sole sulfur source, but not with inorganic (sulfate, sulfite) or organic (methionine, S-adenosylmethionine) sources of sulfur. Moreover, both the point gsh1 and null gsh1/met1 mutants displayed increased sensitivity to the toxic carbon substrate methanol, formaldehyde, organic peroxide and cadmium ions.

Antioxidant activity of melatonin and glutathione interacting with hydroxyl- and superoxide anion radicals

T. Y. Kuznetsova1, N. V. Solovyova2, V. V. Solovyov1, V. O. Kostenko2

1Yu.Kondratyuk Poltava National Technical University, Ukraine;
2Ukrainian Medical Stomatological Academy, Poltava;
e-mail: kzt7@ukr.net

Based on the analysis of the results obtained by quantum chemical modeling of interaction between reduced glutathione (GSH) and melatonin (MLT) molecules with oxygen radicals (•OH and • OOˉ) it was found that this interaction occured following the acid-base mechanism, where MLT and GSH acted as a base in respect of •OH, and as acid in respect of •OOˉ. We have carried out the correlation of the results of quantum chemical calculations (density redistribution, energetic characteristics) under the interaction of MLT and GSH molecules with •OH and •OOˉ in changing macroscopic properties of the process of electroreduction of free oxygen radicals in the presence of antioxidants (potential and maximal current wave reduction waves). This was a direct experimental macroscale evidence of the results of theoretical modeling at the nanoscale level that pointed to a marked antioxidant activity of glutathione compared with melatonin.

Biochemical mechanism of the o,p’-DDD effect on the adrenal cortex

A. S. Mikosha, O. I. Kovzun

V. P. Komisarenko Institute of Endocrinology and Metabolism, National Academy of Medical Sciences of Ukraine, Kyiv;
e-mail: asmikosha@gmail.com

o,p’-Dichlorodiphenyldichloroethane (o,p’-DDD, mitotane) is used in the treatment of adrenocortical cancer and Cushing’s disease. This medicine induces numerous biochemical changes in the adrenal cortex, as well as disorder in the mitochondrial structure. Therewith, the level of produced corticosteroid hormones is significantly reduced. One of the possible causes can be a decrease in the NADPH level due to inhibition of the activity of its reduction system and increased NADPH consumption during the glutathione reduction catalyzed by glutathione reductase. o,p’-DDD is partially metabolized in the adrenal glands, and   the main metabolite (in terms of quantity) is o,p’-dichlorodiphenylacetic acid. However, attempts to find a physiologically active component among metabolites were unsuccessful. The most pronounced changes caused by o,p’-DDD were found in the mitochondria of the adrenal cortex. The respiration at the level of IV and I complexes is suppressed, the protein content of these complexes decreases. The phospholipid composition of the tissue altered and the concentration of diphosphatidylglycerol, the most important component of mitochondrial membranes, decreased. In our opinion, o,p’-DDD, owing to its high lipophilicity, accumulates in the mitochondria membranes and causes conformational disorder followed by disorder in mitochondrial functions. It was shown that o,p’-DDD acts as an inhibitor of acyl-CoA-cholesterol acyltransferase (ACAT, SOAT1). Therefore, adenocorticocytes accumulate free cholesterol, causing endoplasmic reticulum stress, mitochondrial swelling and caspases activation. Increased apoptosis leads to a decline in adrenal function and to a decrease in weight of adrenal glands.

Population-related peculiarities of molecular stress-responsive systems of bivalve mollusk under the effect of tetrazine pesticide

H. I. Falfushynska, L. L. Gnatyshyna, O. B. Stoliar

Volodymyr Hnatiuk Ternopil National Pedagogical University, Ukraine;
e-mail: halynka.f@gmail.com, oksana.stolyar@gmail.com

Tetrazine pesticides are widely used for the treatment of crops in most EU countries and USA. However, data about the effect of environmentally realistic concentrations of biocides on the molecular stress response system in non-target organisms are absent. The aim of our study was the comparison of adaptive capability of bivalve mollusk Anodonta cygnea from two populations under the effects of commercial pesticide Apollo in terms of biochemical parameters of the digestive gland. The differences between parameters of oxidative stress and glutathione transferase activity in specimens of control groups from clean (I group) and polluted (B group) areas have been shown. Under the effect of Apollo, the level of protein carbonyls and  microsomal oxidation processes increased, and the level of metallothioneins and oxyradical formation decreased in the specimens from both populations. However, the treatment provoked the activation of antioxidant processes in the I group and their inhibition in B group. Potentially the injury of cellular thiols, glutathione and metallothioneins, seems to be key point of tetrazine pesticides toxicity.

Manifestations of oxidative stress and molecular damages in ovarian cancer tissue

H. I. Falfushynska1,2, L. L. Gnatyshyna1,2, H. V. Deneha1, O. Y. Osadchuk1, O. B. Stoliar1

1Volodymyr Hnatiuk Ternopil National Pedagogical University, Ukraine;
e-mail: halynka.f@gmail.com;
2I.Ya. Horbachevsky Ternopil State Medical University, Ukraine

Indices of oxidative stress are recognized molecular markers and prognostic criteria for malignant transformation of tissue, but their value depends on the type of tumor and the stage of its development. The goal of this study was to clarify the relationship between the characteristics of the oxidative stress system including­ metal-associated ones and the cytotoxicity manifestations in neoplastically transformed human ovarian tissue. The highest level of Mn-superoxide dismutase activity (by 630%) and metallothionein protein (MT, 100%) has been estimated for the first time in malignant ovarian tissue compared to normal ovarian tissue. The researchers have also found a much higher level of oxy-radical formation (by 332%), a lower activity of catalase (by 49%) and a lower level of reduced glutathione (by 46%) and its redox index (0.84 versus 0.89 in the control) in tumor tissue. Under the relatively stable content of zinc, copper and cadmium in MTs, the content of zinc and especially copper in a form non-binding with MTs was significantly lower in the malignant tissue compared to normal one while the content of cadmium was higher. A discriminant analysis of all definable parameters revealed that the higher content of the products of oxidative destruction of proteins, lipids, fragmented DNA and the activity of cathepsin D, especially in its free form (by 235%), are the main characteristic signs of malignant ovarian tissue.

Transcriptional regulation of the Hansenula polymorpha GSH2 gene in the response to cadmium ion treatment

O. V. Blazhenko, А. B. Kotlyarchuk, V. M. Ubiyvovk

Institute of Cell Biology, National Academy of Science of Ukraine, Lviv;
e-mail: Oleksandra.Blazhenko@googlemail.com

In a previous study we cloned GSH2 gene, encoding γ-glutamylcysteine synthetase (γGCS) in the yeast Hansenula рolymorpha. In this study an analysis of molecular organisation of the H. рolymorpha GSH2 gene promoter was conducted and the potential binding sites of Yap1, Skn7, Creb/Atf1, and Cbf1 transcription factors were detected. It was established that full regulation of GSH2 gene expression in the response to cadmium and oxidative stress requires the length of GSH2 promoter to be longer than 450 bp from the start of translation initiation. To study the transcriptional regulation of H. polymorpha GSH2 gene recombinant strain, harbouring­ a reporter system, in which 1.832 kb regulatory region of GSH2 gene was fused to structural and terminatory regions of alcohol oxidase gene, was constructed. It was shown that maximum increase in H. polymorpha GSH2 gene transcription by 33% occurs in the rich medium under four-hour incubation with 1 μM concentration of cadmium ions. In the minimal medium the GSH2 gene expression does not correlate with the increased total cellular glutathione levels under cadmium ion treatment. We assume that the increased content of total cellular glutathione under cadmium stress in the yeast H. polymorpha probably is not controlled on the level of GSH2 gene transcription.

Quantum chemical modeling of antioxidant activity of glutathione interacting with hydroxyl- and superoxide anion radicals

N. V. Solovyova1, T. Y. Kuznetsova2

1Higher State Medical Educational Institution of Ukraine,
Ukrainian Medical Stomatological Academy, Poltava, Ukraine;
2Yu. Kondratyuk Poltava National Technical University, Ukraine;
e-mail: KZT7@yandex.ru

Following the analysis of the results of quantum chemical simulation of interaction between a GSH molecule and oxygen radicals •ОН and •ООˉ, it was found that it takes place through the acid-base mechanism, where GSH acts as a base towards •ОН, and as an acid towards •ООˉ. The results of quantum chemical calculations (electron density redistribution, energy characteristics) were correlated at the time of interaction of a GSH molecule with •ОН and •ООˉ with a change of macroscopic parameters of the process of free oxygen radical electroreduction in the presence of GSH (potential and maximum current of reduction waves), which is a direct experimental macroscale evidence of results of the conducted nanoscale theoretical simulation.