Key Redox Enzymes Research Articles

The condition of the intracellular metabolism of peripheral blood leukocytes in arterial hypertension (experimental study)

Objective: the comparative analysis of the features of intracellular metabolism of polymorphonuclear leukocytes (PNL) in the peripheral blood of rats with the normal blood pressure and hereditary stress-induced arterial hypertension (ISIAH). Material and methods. Indicators of carbohydrate and lipid metabolism, activity of some key redox enzymes in leukocytes of rats as well as a level of PNL functional activity were examined by histochemical methods. Results. The study determined that the stable increased blood pressure is accompanied by hypertrophy of the left ventricle of the heart, degenerative changes in cardiomyocytes and the decreased microcirculatory bed density. In this case amount of intracellular glycogen in leukocytes was decreased in 23% at unchanged amount of intracellular lipids. Activities of intracellular ATP-ase and myeloperoxidase were reduced in 22% with non-significant increasing of succinate dehydrogenase activity (in 13%). Amount of PNL with positive NBT-test was increased in 35% in the rats of ISIAH. Conclusions. Analysis of the studied metabolic indicators of PNL shows disturbance of the energy-supplying and enzymatic processes in the leukocytes on the background of arterial hypertension.

Tropospheric ozone rapidly decreases root growth by altering carbon metabolism and detoxification capability in growing soybean roots

High tropospheric ozone (O3) concentrations lead to significant global soybean (Glycine max) yield reductions. Research concerning O3 impacts on soybean has focused on the contributions of above-ground tissues. In this study, Mandarin (Ottawa) (O3-sensitive) and Fiskeby III (O3-tolerant) soybean genotypes provide contrasting materials to investigate O3 effects on root growth. We compared root morphological and proteomic changes when 16-day-old plants were treated with charcoal-filtered (CF) air or elevated O3 (80 ppb O3 for 7 h/day) in continuously stirred-tank reactors (CSTR) for 7 days. Our results showed that in Mandarin (Ottawa), decreased expression of enzymes involved in the tricarboxylic acid (TCA) cycle contributes to reduction of root biomass and diameter under elevated O3. In contrast, O3 tolerance in Fiskeby III roots was associated with O3-dependent induction of enzymes involved in glycolysis and O3-independent expression of enzymes involved in the ascorbate-glutathione cycle. We conclude that a decreased abundance of key redox enzymes in roots due to limited carbon availability rapidly alters root growth under O3 stress. However, maintaining a high abundance of enzymes associated with redox status and detoxification capability contributes to overall O3 tolerance in roots.

Talcum powder induces malignant transformation of human primary normal ovarian epithelial cells but not human primary normal peritoneal fibroblasts

Objective: The aim of this study was to demonstrate whether exposure to talcum powder induces malignant transformation in human primary normal ovarian epithelial cells and human primary normal peritoneal fibroblasts. Epidemiological and molecular studies have linked perineal use of talcum powder to increased risk of ovarian cancer. Exposure to talcum powder was shown to induce specific point mutations in key redox enzymes that altered their activities in both normal and epithelial ovarian cancer cells.

Molecular Basis Supporting the Association of Talcum Powder Use With Increased Risk of Ovarian Cancer.

Genital use of talcum powder and its associated risk of ovarian cancer is an important controversial topic. Epithelial ovarian cancer (EOC) cells are known to manifest a persistent prooxidant state. Here we demonstrated that talc induces significant changes in key redox enzymes and enhances the prooxidant state in normal and EOC cells. Using real-time reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay, levels of CA-125, caspase-3, nitrate/nitrite, and selected key redox enzymes, including myeloperoxidase (MPO), inducible nitric oxide synthase (iNOS), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), and glutathione reductase (GSR), were determined. TaqMan genotype analysis utilizing the QuantStudio 12K Flex was used to assess single-nucleotide polymorphisms in genes corresponding to target enzymes. Cell proliferation was determined by MTT proliferation assay. In all talc-treated cells, there was a significant dose-dependent increase in prooxidant iNOS, nitrate/nitrite, and MPO with a concomitant decrease in antioxidants CAT, SOD, GSR, and GPX (P < .05). Remarkably, talc exposure induced specific point mutations that are known to alter the activity in some of these key enzymes. Talc exposure also resulted in a significant increase in inflammation as determined by increased tumor marker CA-125 (P < .05). More importantly, talc exposure significantly induced cell proliferation and decreased apoptosis in cancer cells and to a greater degree in normal cells (P < .05). These findings are the first to confirm the cellular effect of talc and provide a molecular mechanism to previous reports linking genital use to increased ovarian cancer risk.

Talc induces a pro-oxidant state in normal and ovarian cancer cells through gene point mutations in key redox enzymes

Talc induces a pro-oxidant state in normal and ovarian cancer cells through gene point mutations in key redox enzymes

Insight into the molecular mechanism of the sulfur oxidation process by reverse sulfite reductase (rSiR) from sulfur oxidizer Allochromatium vinosum.

Sulfur metabolism is one of the oldest known biochemical processes. Chemotrophic or phototrophic proteobacteria, through the dissimilatory pathway, use sulfate, sulfide, sulfite, thiosulfate or elementary sulfur by either reductive or oxidative mechanisms. During anoxygenic photosynthesis, anaerobic sulfur oxidizer Allochromatium vinosum forms sulfur globules that are further oxidized by dsr operon. One of the key redox enzymes in reductive or oxidative sulfur metabolic pathways is the DsrAB protein complex. However, there are practically no reports to elucidate the molecular mechanism of the sulfur oxidation process by the DsrAB protein complex from sulfur oxidizer Allochromatium vinosum. In the present context, we tried to analyze the structural details of the DsrAB protein complex from sulfur oxidizer Allochromatium vinosum by molecular dynamics simulations. The molecular dynamics simulation results revealed the various types of molecular interactions between DsrA and DsrB proteins during the formation of DsrAB protein complex. We, for the first time, predicted the mode of binding interactions between the co-factor and DsrAB protein complex from Allochromatium vinosum. We also compared the binding interfaces of DsrAB from sulfur oxidizer Allochromatium vinosum and sulfate reducer Desulfovibrio vulgaris. This study is the first to provide a comparative aspect of binding modes of sulfur oxidizer Allochromatium vinosum and sulfate reducer Desulfovibrio vulgaris.

Synthesis and antiproliferative activities of thioxoflavonoids on three human cancer cells

Two series of fifteen novel thioxoflavonoids 2a-2h and 4a-4g were synthesized from corresponding flavonoids 1a-1h and 3a-3g by reacting with Lawesson’s reagent, respectively. Their in vitro antiproliferative activities were evaluated on a panel of three human cancer cell lines (Hela, HCC1954 and SK-OV-3) using cell counting kit-8 (CCK-8) assay. The results showed that most of the target compounds exhibited moderate to good antiproliferative activities against the three human cancer cell lines. In particular, thioxoflavonoids 2f and 2g showed the strongest antiproliferative activity on all three human cancer cell lines with IC50 values ranging from 3.34 to 4.67 μM, 4f showed the best antiproliferative activity on Hela cells (IC50 2.30 μM), 2e showed the best antiproliferative activity on HCC1954 cells (IC50 2.13 μM) and SK-OV-3 cells (IC50 2.33 μM). The antiproliferative activities may be involved in their antioxidant activity, which can be speculated by their ability to scavenge free radicals and by their capacity of affecting key redox enzymes.

Oxidative stress: a key regulator of leiomyoma cell survival

To determine the effects of attenuating oxidative stress with the use of dichloroacetate (DCA) on the expression of key redox enzymes myeloperoxidase (MPO) and inducible nitric oxide synthase (iNOS) as well as on apoptosis. Prospective experimental study. University medical center. Cells established from myometrium and uterine fibroid from the same patients. Cells were exposed to normal (20% O2) or hypoxic (2% O2) conditions for 24hours with or without DCA (20μg/mL), a metabolic modulator that shifts anaerobic to aerobic metabolism. Nitrate/nitrite (iNOS activity indicator), iNOS, Bcl-2/Bax ratio, MPO, and caspase-3 activities and levels were determined by means of Greiss assay, real-time reverse-transcription polymerase chain reaction, and ELISA. Data were analyzed with the use of SPSS by means of one-way analysis of variance with Tukey post hoc analysis and independent t tests. MPO, iNOS, and nitrate/nitrite expression were higher in leiomyoma than in myometrial cells, and they were further enhanced by hypoxia in myometrial cells. Treatment with the use of DCA decreased MPO, iNOS, and nitrate/nitrite levels and negated the effect of hypoxia in both types of cells. Leiomyoma cells showed less apoptosis, as indicated by both caspase-3 activity and the Bcl-2/Bax ratio, than myometrial cells. Hypoxia further decreased apoptosis in myometrial cells with no further effect on leiomyoma cells. Treatment with DCA resulted in increased apoptosis in both types of cells, even in the presence of hypoxia. Shifting anaerobic to aerobic metabolism with the use of DCA resulted in an increase in apoptosis in leiomyoma cells and protected myometrial cells from the acquisition of the leiomyoma-like phenotype.

Specific point mutations in key redox enzymes are associated with chemoresistance in epithelial ovarian cancer

Oxidative stress plays an important role in the pathophysiology of ovarian cancer. Resistance to chemotherapy presents a significant challenge for ovarian cancer treatment. Specific single nucleotide polymorphisms (SNPs) in key redox enzymes have been associated with ovarian cancer survival and progression. The objective of this study was to determine whether chemotherapy induces point mutations in key redox enzymes that lead to the acquisition of chemoresistance in epithelial ovarian cancer (EOC). Human EOC cell lines and their chemoresistant counterpart were utilized for this study. Specific SNPs in key redox enzymes were analyzed by TaqMan SNP Genotyping. Activities and levels of key redox enzymes were determined by real-time RT-PCR, ELISA and a greiss assay. Point mutations in key redox enzymes were introduced into sensitive EOC cells via the CRISPR/Cas9 system. Cell viability and IC50 for cisplatin were determined by the MTT Cell Proliferation Assay. Data was analyzed with SPSS using Student's two-tailed t-tests and One-way ANOVA followed by Dunnett's or Tukey's post hoc tests, p<0.05. Here, we demonstrate that chemoresistant EOC cells are characterized by a further enhancement in oxidative stress as compared to sensitive counterparts. Additionally, chemoresistant EOC cells manifested specific point mutations, which are associated with altered enzymatic activity, in key redox enzymes that are not detected in sensitive counterparts. Supplementation of an antioxidant was able to successfully sensitize EOC cells to chemotherapeutics. Causality was established by the induction of these point mutations in sensitive EOC cells, which resulted in a significant increase in the level of chemoresistance. These findings indicate that chemotherapy induces specific point mutations in key redox enzymes that contribute to the acquisition of chemoresistance in EOC cells, highlighting a potential novel mechanism. Identification of targets for chemoresistance with either biomarker and/or screening potential will have a significant impact for the treatment of this disease.

Molecular mechanism underlying antileishmanial effect of oxabicyclo[3.3.1]nonanones: Inhibition of key redox enzymes of the pathogen

We report oxabicyclo[3.3.1]nonanones as inhibitors of key redox enzymes, trypanothione synthetase (TryS), and trypanothione reductase (TryR) of Leishmania. Further, detailed cellular effects of 4-(4,4,8-Trimethyl-7-oxo-3-oxabicyclo[3.3.1]non-2-yl)-benzoic acid methyl ester, a oxabicyclo[3.3.1]nonanones, on the parasite were investigated. As these compounds inhibit key redox enzymes (TryR amd TryS), treatment of these compounds resulted in increased reactive oxygen species (ROS), mitochondrial membrane damage, activation of caspase like proteases, and DNA damage that finally leads to apoptosis. Although the compound has modest IC50 value against parasite (4.9±0.4μM), they identify a novel chemical space to design and develop drugs based on these compounds against the Leishmania parasite. This is first report of oxabicyclo[3.3.1]nonanones as antileishmanial.

Natural Terpenes Prevent Mitochondrial Dysfunction, Oxidative Stress and Release of Apoptotic Proteins during Nimesulide-Hepatotoxicity in Rats

Nimesulide, an anti-inflammatory and analgesic drug, is reported to cause severe hepatotoxicity. In this study, molecular mechanisms involved in deranged oxidant-antioxidant homeostasis and mitochondrial dysfunction during nimesulide-induced hepatotoxicity and its attenuation by plant derived terpenes, camphene and geraniol has been explored in male Sprague-Dawley rats. Hepatotoxicity due to nimesulide (80 mg/kg BW) was evident from elevated SGPT, SGOT, bilirubin and histo-pathological changes. Antioxidants and key redox enzymes (iNOS, mtNOS, Cu/Zn-SOD, Mn-SOD, GPx and GR) were altered significantly as assessed by their mRNA expression, Immunoblot analysis and enzyme activities. Redox imbalance along with oxidative stress was evident from decreased NAD(P)H and GSH (56% and 74% respectively; P<0.001), increased superoxide and secondary ROS/RNS generation along with oxidative damage to cellular macromolecules. Nimesulide reduced mitochondrial activity, depolarized mitochondria and caused membrane permeability transition (MPT) followed by release of apoptotic proteins (AIF; apoptosis inducing factor, EndoG; endonuclease G, and Cyto c; cytochrome c). It also significantly activated caspase-9 and caspase-3 and increased oxidative DNA damage (level of 8-Oxoguanine glycosylase; P<0.05). A combination of camphene and geraniol (CG; 1∶1), when pre-administered in rats (10 mg/kg BW), accorded protection against nimesulide hepatotoxicity in vivo, as evident from normalized serum biomarkers and histopathology. mRNA expression and activity of key antioxidant and redox enzymes along with oxidative stress were also normalized due to CG pre-treatment. Downstream effects like decreased mitochondrial swelling, inhibition in release of apoptotic proteins, prevention of mitochondrial depolarization along with reduction in oxidized NAD(P)H and increased mitochondrial electron flow further supported protective action of selected terpenes against nimesulide toxicity. Therefore CG, a combination of natural terpenes prevented nimesulide induced cellular damage and ensuing hepatotoxicity.

The Suppressive Effects of Bursopentine (BP5) on Oxidative Stress and NF-ĸB Activation in Lipopolysaccharide-activated Murine Peritoneal Macrophages

Background/Aim: Bursopentine (BP5) is a novel thiol-containing pentapeptide isolated from chicken bursa of Fabricius, and is reported to exert immunomodulatory effects on B and T lymphocytes. It has been found that some thiol compounds, such as glutathione (GSH) and N-acetylcysteine (NAC) protect living cells from oxidative stress. This led us to investigate whether BP5 had any ability to protect macrophages from oxidative stress as well as any mechanism that might underlie this process. Methods: Murine peritoneal macrophages activated by lipopolysaccharide (LPS) (2 µg/ml) were treated with single bouts (0, 25, 50, and 100 µM) of BP5. Results: BP5 potently suppressed the markers for oxidative stress, including nitric oxide (NO), reactive oxygen species (ROS), lipid peroxidation, and protein oxidation. It also decreased the expression and activity of inducible nitric oxide synthase (iNOS) and promoted a protective antioxidant state by elevating GSH content and by activating the expression and activity of certain key antioxidant and redox enzymes, including glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD) and catalase (CAT). This suppressive effect on oxidative stress was accompanied by down-regulated expression and activity of nuclear factor kappa B (NF-ĸB). Conclusion: These findings demonstrate that BP5 can protect LPS-activated murine peritoneal macrophages from oxidative stress. BP5 may have applications as an anti-oxidative stress reagent.