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Annesi L, Tossetta G, Borghi C, Piani F. The Role of Xanthine Oxidase in Pregnancy Complications: A Systematic Review. Antioxidants (Basel) 2024; 13:1234. [PMID: 39456486 PMCID: PMC11505381 DOI: 10.3390/antiox13101234] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2024] [Revised: 10/01/2024] [Accepted: 10/07/2024] [Indexed: 10/28/2024] Open
Abstract
Xanthine oxidoreductase (XOR) is an enzyme involved in the oxidation of hypoxanthine and xanthine to uric acid. XOR has two isoforms: xanthine dehydrogenase and xanthine oxidase (XO). XO plays a major role in oxidative stress, causing the formation of reactive oxygen species. In the present study, we aimed to summarize the evidence on the association between XO and pregnancy complications. The PRISMA checklist guided the reporting of the data. We conducted systematic searches in the PubMed and Web of Science databases to identify all human studies investigating XO in pregnancy diseases up to June 2024. A total of 195 references have been identified and 14 studies were included. Most studies focused on women with PE and GD. Overall, all the included studies found a statistically significant increase in maternal, placental, and/or fetal XO levels, activity, or tissue expression in women with pregnancy complications, compared to those with uncomplicated pregnancies. Although promising, the quality and dimension of the included studies do not allow for a definitive answer to the question of whether XO may play a crucial role in pregnancy complications. Future studies are warranted to confirm if XO could represent a prognostic and therapeutic marker in pregnancy complications and their impact on long-term maternal and offspring cardiovascular health.
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Affiliation(s)
- Lorenzo Annesi
- Hypertension and Cardiovascular Risk Research Center, Medical and Surgical Sciences Department, Alma Mater Studiorum University of Bologna, 40138 Bologna, Italy; (L.A.); (C.B.)
| | - Giovanni Tossetta
- Department of Experimental and Clinical Medicine, Università Politecnica delle Marche, 60126 Ancona, Italy;
| | - Claudio Borghi
- Hypertension and Cardiovascular Risk Research Center, Medical and Surgical Sciences Department, Alma Mater Studiorum University of Bologna, 40138 Bologna, Italy; (L.A.); (C.B.)
| | - Federica Piani
- Hypertension and Cardiovascular Risk Research Center, Medical and Surgical Sciences Department, Alma Mater Studiorum University of Bologna, 40138 Bologna, Italy; (L.A.); (C.B.)
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Mudgal R, Singh S. Xanthine Oxidoreductase in the Pathogenesis of Endothelial Dysfunction: An Update. Curr Hypertens Rev 2024; 20:10-22. [PMID: 38318826 DOI: 10.2174/0115734021277772240124075120] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2023] [Revised: 12/04/2023] [Accepted: 12/19/2023] [Indexed: 02/07/2024]
Abstract
Xanthine oxidoreductase (XOR) is a rate-limiting enzyme in the formation of uric acid (UA) and is involved in the generation of reactive oxygen species (ROS). Overproduction of ROS has been linked to the pathogenesis of hypertension, atherosclerosis, and cardiovascular disease, with multiple studies over the last 30 years demonstrating that XOR inhibition is beneficial. The involvement of XOR and its constituents in the advancement of chronic inflammation and ROS, which are responsible for endothelial dysfunction, is the focus of this evidence-based review. An overabundance of XOR products and ROS appears to drive the inflammatory response, resulting in significant endothelium damage. It has also been demonstrated that XOR activity and ED are connected. Diabetes, hypertension, and cardiovascular disease are all associated with endothelial dysfunction. ROS mainly modifies the activity of vascular cells and can be important in normal vascular physiology as well as the development of vascular disease. Suppressing XOR activity appears to decrease endothelial dysfunction, probably because it lessens the generation of reactive oxygen species and the oxidative stress brought on by XOR. Although there has long been a link between higher vascular XOR activity and worse clinical outcomes, new research suggests a different picture in which positive results are mediated by XOR enzymatic activity. Here in this study, we aimed to review the association between XOR and vascular endothelial dysfunction. The prevention and treatment approaches against vascular endothelial dysfunction in atherosclerotic disease.
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Affiliation(s)
- Rajat Mudgal
- Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research, Hajipur, Bihar, India
| | - Sanjiv Singh
- Department of Pharmacology and Toxicology, National Institute of Pharmaceutical Education and Research, Hajipur, Bihar, India
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Peleli M, Zampas P, Papapetropoulos A. Hydrogen Sulfide and the Kidney: Physiological Roles, Contribution to Pathophysiology, and Therapeutic Potential. Antioxid Redox Signal 2022; 36:220-243. [PMID: 34978847 DOI: 10.1089/ars.2021.0014] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Significance: Hydrogen sulfide (H2S), the third member of the gasotransmitter family, has a broad spectrum of biological activities, including antioxidant and cytoprotective actions, as well as vasodilatory, anti-inflammatory and antifibrotic effects. New, significant aspects of H2S biology in the kidney continue to emerge, underscoring the importance of this signaling molecule in kidney homeostasis, function, and disease. Recent Advances: H2S signals via three main mechanisms, by maintaining redox balance through its antioxidant actions, by post-translational modifications of cellular proteins (S-sulfhydration), and by binding to protein metal centers. Important renal functions such as glomerular filtration, renin release, or sodium reabsorption have been shown to be regulated by H2S, using either exogenous donors or by the endogenous-producing systems. Critical Issues: Lower H2S levels are observed in many renal pathologies, including renal ischemia-reperfusion injury and obstructive, diabetic, or hypertensive nephropathy. Unraveling the molecular targets through which H2S exerts its beneficial effects would be of great importance not only for understanding basic renal physiology, but also for identifying new pharmacological interventions for renal disease. Future Directions: Additional studies are needed to better understand the role of H2S in the kidney. Mapping the expression pattern of H2S-producing and -degrading enzymes in renal cells and generation of cell-specific knockout mice based on this information will be invaluable in the effort to unravel additional roles for H2S in kidney (patho)physiology. With this knowledge, novel targeted more effective therapeutic strategies for renal disease can be designed. Antioxid. Redox Signal. 36, 220-243.
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Affiliation(s)
- Maria Peleli
- Clinical, Experimental Surgery and Translational Research Center, Biomedical Research Foundation of the Academy of Athens, Athens, Greece.,Laboratory of Pharmacology, Department of Pharmacy, National and Kapodistrian University of Athens, Athens, Greece
| | - Paraskevas Zampas
- Clinical, Experimental Surgery and Translational Research Center, Biomedical Research Foundation of the Academy of Athens, Athens, Greece.,Laboratory of Pharmacology, Department of Pharmacy, National and Kapodistrian University of Athens, Athens, Greece
| | - Andreas Papapetropoulos
- Clinical, Experimental Surgery and Translational Research Center, Biomedical Research Foundation of the Academy of Athens, Athens, Greece.,Laboratory of Pharmacology, Department of Pharmacy, National and Kapodistrian University of Athens, Athens, Greece
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Kausar F, Yusuf Amin KM, Bashir S, Parvez A, Ahmad P. Concept of 'Ihtiraq' in Unani Medicine - A correlation with oxidative stress, and future prospects. JOURNAL OF ETHNOPHARMACOLOGY 2021; 265:113269. [PMID: 32937158 DOI: 10.1016/j.jep.2020.113269] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/27/2019] [Revised: 07/23/2020] [Accepted: 08/09/2020] [Indexed: 06/11/2023]
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE In recent years, oxidative stress (OS) and the generation of ROS have been recognized as a fundamental pathology contributing, at least partially, to a number of important diseases. However, the therapeutic application has been simplistically limited to using antioxidants with little correction of diseases, and many biomarkers of OS, although confirming and quantifying the magnitude of this pathology, are not suggestive of the underlying causes behind generation of a large amount of free radicals. Unfortunately, research has not noted the multi-implication parallel phenomenon of Ihtiraq (Combustion) in Unani Medicine, which possesses much richer etiopathological sub-typing and much more variegated selective and specific treatments (and prophylactics) corresponding to each sub-type of Ihtiraq; the identification of each sub-type's molecular counterparts can be used to develop not only sub-types of OS pathologies and corresponding selective treatments/prophylactics but also non-biomolecular factors. Eminent Unani physicians described a deteriorative phenomenon, which they termed as 'Ihtiraq' which stands for extreme metabolism or 'combustion' and is recognized as a fundamental pathology, contributing as a major factor to the development of chronic diseases. Further, Unani Medicine also possesses a pathophysiological phenomenon called 'Hararat Ghariba' (Unnatural Heat) whose diverse associations with Ihtiraq may be correlatable as upstream, parallel, or downstream associations of OS and consequent pathologies. AIM OF THE STUDY The aim of the study is to: 1. Explore the correlation of the phenomenon and etiopathology of Ihtiraq and OS and the treatment and prevention of the pathologies arising from them. 2. Extrapolate Ihtiraq, its types, causes, prevention, and treatment to OS, hitherto existing as a fundamental and monolithic pathology of increased ROS, to hypothesize its molecular-level sub-typing, as well as to propose selective interventions in these molecular sub-types of OS in place of the existing use of only basic antioxidants such as Vitamin C. MATERIAL AND METHODS This review is presented with a noteworthy insight into Unani concepts and a thorough study of classical Unani literature by Ibn Sina (10th century), Zakaria Razi (9th century), Ibn Rushd (12th century), Ibn al-Nafees (13th century), Majusi (10th century), and Jurjani (11th century), and comparative detailed study of modern concepts of OS from literature databases, as well as Google, recent researches, and review articles. RESULT The study showed very close correspondences between the phenomenon, etiopathology, and treatment and prevention of Ihtiraq in Unani Medicine and OS in contemporary biomolecular medicine. It also revealed sub-types of Ihtiraq and corresponding selective Unani treatments and prophylactics including drugs and non-drug factors. CONCLUSION After a comprehensive study and analysis of the most recent researches and classical theories, it can be stated that OS can be seen as a molecular level expression of Ihtiraq. Further, various components of Ihtiraq may be used to hypothesize molecular sub-types of OS and propose corresponding specific interventions.
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Affiliation(s)
- Firdaus Kausar
- Dept. of Ilmul Advia, Regional Research Institute of Unani Medicine, University of Kashmir, Srinagar, 190006, India.
| | - Kunwar Mohammad Yusuf Amin
- Philosophy-Science Forum, Dept. of Ilmul Advia, Ajmal Khan Tibbiya College, Aligarh Muslim University, Aligarh, 202002, India
| | - Showkeen Bashir
- Dept. of Veterinary Biochemistry, Faculty of Veterinary Sciences and Animal Husbandry, SKAUST-K, Srinagar, India
| | - Athar Parvez
- Dept. of Ilmul Advia, Regional Research Institute of Unani Medicine, University of Kashmir, Srinagar, 190006, India
| | - Pervaiz Ahmad
- Dept. of Ilmul Advia, Regional Research Institute of Unani Medicine, University of Kashmir, Srinagar, 190006, India
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Fang C, Chen L, He M, Luo Y, Zhou M, Zhang N, Yuan J, Wang H, Xie Y. Molecular mechanistic insight into the anti-hyperuricemic effect of Eucommia ulmoides in mice and rats. PHARMACEUTICAL BIOLOGY 2019; 57:112-119. [PMID: 30843748 PMCID: PMC6419643 DOI: 10.1080/13880209.2019.1568510] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/16/2018] [Revised: 11/11/2018] [Accepted: 12/31/2018] [Indexed: 05/19/2023]
Abstract
CONTEXT Eucommia ulmoides Oliver (Eucommiaceae) has various medicinal properties. Our previous studies revealed that Eucommia ulmoides has a protective effect on hyperuricaemia. OBJECTIVE This study investigates the effect of Eucommia ulmoides cortex ethanol extract (EU) on hyperuricaemia and explores the underlying mechanism in Kunming mice and Sprague-Dawley rats. MATERIAL AND METHODS Sixty mice and sixty rats were divided into normal control, hyperuricaemia, allopurinol (10 mg/kg) and three EU groups. The EU groups received intragastric EU at 80, 160, 320 mg/kg in mice and 100, 200, 400 mg/kg in rats for 7 days. Serum uric acid (SUA) was measured using a kit. mRNA and proteins were quantified by RT-qPCR and immunohistochemical assays (IHC), respectively. RESULTS The Maximal Tolerable Dose (MTD) of EU administered intragastrically was 18 g/kg in mice. The intermediate (160 mg/kg) and high (320 mg/kg) EU treatment significantly reduced (p < 0.05) SUA levels to 130.16 μmol/L and 109.29 μmol/L, respectively, and markedly elevated the mRNA expression of organic anion transporters 1 (OAT1) and organic anion transporters 3 (OAT3), while significantly deceasing the mRNA levels of glucose transporter 9 (GLUT9) and uric acid transporter 1 (URAT1) in the mouse kidney (p < 0.05). In hyperuricemic rats, high EU (400 mg/kg) significantly reduced SUA levels to 253.85 μmol/L, and increased OAT1 and OAT3 levels, but decreased URAT1 and GLUT9, compared to the hyperuricaemia group (p < 0.05). DISCUSSION AND CONCLUSIONS This study demonstrated the potential hyperuricaemia ameliorating effect of EU. Specific active ingredients in EU should be evaluated. These results are valuable for the development of antihyperuritic agents from EU.
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Affiliation(s)
- Cong Fang
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Lanying Chen
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
- CONTACT Lanying Chen National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, 56 Yanming Road, Nanchang, China
| | - Mingzhen He
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Yingying Luo
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Mengjing Zhou
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Ni Zhang
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Jinfeng Yuan
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Huiling Wang
- National Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
| | - Yongyan Xie
- School of Basic Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, China
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Ji P, B Nonnecke E, Doan N, Lönnerdal B, Tan B. Excess Iron Enhances Purine Catabolism Through Activation of Xanthine Oxidase and Impairs Myelination in the Hippocampus of Nursing Piglets. J Nutr 2019; 149:1911-1919. [PMID: 31373370 DOI: 10.1093/jn/nxz166] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2019] [Revised: 06/17/2019] [Accepted: 06/20/2019] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Few studies have addressed the risk of nutritional iron overexposure in infancy. We previously found that excess dietary iron in nursing piglets resulted in iron overload in the liver and hippocampus and diminished socialization with novel conspecifics in a test for social novelty preference. OBJECTIVES This experiment aimed to identify metabolites and metabolic pathways affected by iron overload in the liver and hippocampus of nursing piglets. METHODS Liver and hippocampal tissues collected from 22-d-old piglets (Hampshire × Yorkshire crossbreed; 5.28 ± 0.53 kg body weight; 50% male) that received orally 0 (NI group) or 50 mg iron/(d · kg body weight) (HI group) from postnatal day (PD) 2 to PD21 were analyzed for mRNA and protein expression and enzyme activity of xanthine oxidase (XO). Untargeted metabolomics was performed using GC-MS. Expression of myelin basic protein (MBP) in the hippocampus was determined using western blot. RESULTS There were 108 and 126 metabolites identified in the hippocampus and liver, respectively. Compared with NI, HI altered 15 metabolites (P < 0.05, q < 0.2) in the hippocampus, including a reduction in myo-inositol (0.86-fold) and N-acetylaspartic acid (0.84-fold), 2 metabolites important for neuronal function and myelination. Seven metabolites involved in purine and pyrimidine metabolism (e.g., hypoxanthine, xanthine, and β-alanine) were coordinately changed in the hippocampus (P < 0.05, q < 0.2), suggesting that iron excess enhanced purine catabolism. The mRNA expression (2.3-fold) (P < 0.05) and activity of XO, a rate-limiting enzyme in purine degradation, was increased. Excess iron increased hippocampal lipid peroxidation by 74% (P < 0.05) and decreased MBP by 44% (P = 0.053). The hepatic metabolome was unaffected. CONCLUSIONS In nursing piglets, excess iron enhances hippocampal purine degradation through activation of XO, which may induce oxidative stress and alter energy metabolism in the developing brain.
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Affiliation(s)
- Peng Ji
- Department of Nutrition, University of California, Davis, Davis, CA, USA
| | - Eric B Nonnecke
- Department of Nutrition, University of California, Davis, Davis, CA, USA
| | - Nicole Doan
- Department of Nutrition, University of California, Davis, Davis, CA, USA
| | - Bo Lönnerdal
- Department of Nutrition, University of California, Davis, Davis, CA, USA
| | - Bie Tan
- Laboratory of Animal Nutritional Physiology and Metabolic Process, Key Laboratory of Agro-ecological Processes in Subtropical Region, National Engineering Laboratory for Pollution Control and Waste Utilization in Livestock and Poultry Production, Institute of Subtropical Agriculture, Chinese Academy of Sciences, Changsha, Hunan, China
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Zhang L, Wang X, Cueto R, Effi C, Zhang Y, Tan H, Qin X, Ji Y, Yang X, Wang H. Biochemical basis and metabolic interplay of redox regulation. Redox Biol 2019; 26:101284. [PMID: 31400697 PMCID: PMC6831867 DOI: 10.1016/j.redox.2019.101284] [Citation(s) in RCA: 179] [Impact Index Per Article: 29.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2019] [Revised: 07/23/2019] [Accepted: 07/25/2019] [Indexed: 12/13/2022] Open
Abstract
Accumulated evidence strongly indicates that oxidative stress, characterized by an imbalance between reactive oxygen species (ROS) production and antioxidants in favor of oxidants, plays an important role in disease pathogenesis. However, ROS can act as signaling molecules and fulfill essential physiological functions at basal levels. Each ROS would be different in the extent to stimulate and contribute to different pathophysiological effects. Importantly, multiple ROS generators can be activated either concomitantly or sequentially by relevant signaling molecules for redox biological functions. Here, we summarized the current knowledge related to chemical and biochemical features of primary ROS species and corresponding antioxidants. Metabolic pathways of five major ROS generators and five ROS clearance systems were described, including their ROS products, specific ROS enriched tissue, cell and organelle, and relevant functional implications. We provided an overview of ROS generation and induction at different levels of metabolism. We classified 11 ROS species into three types based on their reactivity and target selectivity and presented ROS homeostasis and functional implications in pathological and physiological status. This article intensively reviewed and refined biochemical basis, metabolic signaling and regulation, functional insights, and provided guidance for the identification of novel therapeutic targets.
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Affiliation(s)
- Lixiao Zhang
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA
| | - Xianwei Wang
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA
| | - Ramón Cueto
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA
| | - Comfort Effi
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA
| | - Yuling Zhang
- Cardiovascular Medicine Department, Sun Yat-sen Memorial Hospital, China
| | - Hongmei Tan
- Department of Pathophysiology, Zhongshan School of Medicine, Sun Yat-sen University, 510080, China
| | - Xuebin Qin
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA
| | - Yong Ji
- Key Laboratory of Cardiovascular Disease and Molecular Intervention, Nanjing Medical University, China
| | - Xiaofeng Yang
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA; Department of Pharmacology, Temple University School of Medicine, Philadelphia, PA, 19140, USA
| | - Hong Wang
- Center for Metabolic Disease Research, Temple University School of Medicine, Philadelphia, PA, 19140, USA; Department of Pharmacology, Temple University School of Medicine, Philadelphia, PA, 19140, USA.
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Lakatos RK, Dobolyi Á, Kovács Z. Uric acid and allopurinol aggravate absence epileptic activity in Wistar Albino Glaxo Rijswijk rats. Brain Res 2018; 1686:1-9. [PMID: 29457994 DOI: 10.1016/j.brainres.2018.02.012] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2017] [Revised: 01/05/2018] [Accepted: 02/10/2018] [Indexed: 11/18/2022]
Abstract
Uric acid has a role in several physiological and pathophysiological processes. For example, uric acid may facilitate seizure generalization while reducing uric acid level may evoke anticonvulsant/antiepileptic effects. Allopurinol blocks the activity of xanthine oxidase, by which allopurinol inhibits catabolism of hypoxanthine to xanthine and uric acid and, as a consequence, decreases the level of uric acid. Although the modulation of serum uric acid level is a widely used strategy in the treatment of certain diseases, our knowledge regarding the effects of uric acid on epileptic activity is far from complete. Thus, the main aim of this study was the investigation of the effect of uric acid on absence epileptic seizures (spike-wave discharges: SWDs) in a model of human absence epilepsy, the Wistar Albino Glaxo/Rijswijk (WAG/Rij) rat. We investigated the influence of intraperitoneally (i.p.) injected uric acid (100 mg/kg and 200 mg/kg), allopurinol (50 mg/kg and 100 mg/kg), a cyclooxygenase 1 and 2 (COX-1 and COX-2) inhibitor indomethacin (10 mg/kg) and inosine (500 mg/kg) alone and the combined application of allopurinol (50 mg/kg) with uric acid (100 mg/kg) or inosine (500 mg/kg) as well as indomethacin (10 mg/kg) with uric acid (100 mg/kg) and inosine (500 mg/kg) with uric acid (100 mg/kg) on absence epileptic activity. We demonstrated that both uric acid and allopurinol alone significantly increased the number of SWDs whereas indomethacin abolished the uric acid-evoked increase in SWD number. Our results suggest that uric acid and allopurinol have proepileptic effects in WAG/Rij rats.
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Affiliation(s)
- Renáta Krisztina Lakatos
- Institute of Biology, University of Pécs, Pécs, Hungary; Savaria Department of Biology, Savaria University Centre, ELTE Eötvös Loránd University, Szombathely, Hungary.
| | - Árpád Dobolyi
- Laboratory of Neuromorphology and Human Brain Tissue Bank, Department of Anatomy, Histology and Embryology, Semmelweis University, Budapest, Hungary; MTA-ELTE Laboratory of Molecular and Systems Neurobiology, Department of Physiology and Neurobiology, Hungarian Academy of Sciences and Eötvös Loránd University, Budapest, Hungary.
| | - Zsolt Kovács
- Savaria Department of Biology, Savaria University Centre, ELTE Eötvös Loránd University, Szombathely, Hungary.
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Yong T, Zhang M, Chen D, Shuai O, Chen S, Su J, Jiao C, Feng D, Xie Y. Actions of water extract from Cordyceps militaris in hyperuricemic mice induced by potassium oxonate combined with hypoxanthine. JOURNAL OF ETHNOPHARMACOLOGY 2016; 194:403-411. [PMID: 27717908 DOI: 10.1016/j.jep.2016.10.001] [Citation(s) in RCA: 48] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/29/2016] [Revised: 07/13/2016] [Accepted: 10/03/2016] [Indexed: 06/06/2023]
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE Cordyceps militaris was recorded in the classic traditional Chinese medicine book with the main functions of "protecting liver and enhancing kidney functions", influencing serum uric acid levels. AIM OF STUDY The aim is to investigate the hypouricemic effects and possible mechanism of C. militaris in hyperuricemic mice. MATERIALS AND METHODS A water extract (WECM) was prepared by decocting C. militaris directly at 80 °C in water bath, followed by lyophilization. WECM at 50, 100 and 200mg/kg was orally administered to hyperuricemic mice induced by potassium oxonate and hypoxanthine combinedly and allopurinol (5mg/kg) was served as a positive control. RESULTS WECM exhibited excellent hypouricemic activity, which could decrease the serum uric acid levels of the hyperuricemic mice (306μmol/L) to 189, 184 and 162μmol/L at different doses respectively (P<0.01), approaching the levels of normal mice (184μmol/L). The urate transporter 1 (URAT1) protein levels of kidney at different doses of WECM were 28.15, 17.43, 9.03pg/mL respectively, much lower than that in the hyperuricemia group (93.45pg/mL, P<0.01); and suggested WECM may interact with URAT1. Docking simulations using modeled structure of URAT1 suggested that LYS145, ARG325, ARG477 and ASP168 of URAT1 are key functional residues of URAT1. Four active compounds in C. militaris were identified and their interaction energies with target were estimated between -200 and -400kcal/mol. CONCLUSIONS These findings suggested that C. militaris produced significant hypouricemic actions and the hypouricemic effects of WECM may be attributed to the inhibitive effect of WECM on URAT1 protein levels. The results of blood urine nitrogen and serum creatinine levels and liver, kidney and spleen coefficients showed that WECM have no negative impacts on liver, renal and spleen functions. The screened four active compounds using molecular docking method deserve further investigation in other work.
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Affiliation(s)
- Tianqiao Yong
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China; Guangdong Yuewei Edible Fungi Technology Co., Guangzhou 510663, China.
| | - Minglong Zhang
- Guangdong Yuewei Edible Fungi Technology Co., Guangzhou 510663, China
| | - Diling Chen
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China
| | - Ou Shuai
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China
| | - Shaodan Chen
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China
| | - Jiyan Su
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China
| | - Chunwei Jiao
- Guangdong Yuewei Edible Fungi Technology Co., Guangzhou 510663, China
| | - Delong Feng
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China
| | - Yizhen Xie
- State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application and Guangdong Open Laboratory of Applied Microbiology, Guangdong Institute of Microbiology, Guangzhou 510070, China; Guangdong Yuewei Edible Fungi Technology Co., Guangzhou 510663, China
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Xanthine Oxidoreductase-Derived Reactive Species: Physiological and Pathological Effects. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2015; 2016:3527579. [PMID: 26823950 PMCID: PMC4707389 DOI: 10.1155/2016/3527579] [Citation(s) in RCA: 177] [Impact Index Per Article: 17.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/25/2015] [Accepted: 11/01/2015] [Indexed: 12/20/2022]
Abstract
Xanthine oxidoreductase (XOR) is the enzyme that catalyzes the oxidation of hypoxanthine to xanthine and xanthine to uric acid and is widely distributed among species. In addition to this housekeeping function, mammalian XOR is a physiological source of superoxide ion, hydrogen peroxide, and nitric oxide, which can function as second messengers in the activation of various pathways. This review intends to address the physiological and pathological roles of XOR-derived oxidant molecules. The cytocidal action of XOR products has been claimed in relation to tissue damage, in particular damage induced by hypoxia and ischemia. Attempts to exploit this activity to eliminate unwanted cells via the construction of conjugates have also been reported. Moreover, different aspects of XOR activity related to phlogosis, endothelial activation, leukocyte activation, and vascular tone regulation, have been taken into consideration. Finally, the positive and negative outcomes concerning cancer pathology have been analyzed because XOR products may induce mutagenesis, cell proliferation, and tumor progression, but they are also associated with apoptosis and cell differentiation. In conclusion, XOR activity generates free radicals and other oxidant reactive species that may result in either harmful or beneficial outcomes.
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Nishino T, Okamoto K, Kawaguchi Y, Matsumura T, Eger BT, Pai EF, Nishino T. The C-terminal peptide plays a role in the formation of an intermediate form during the transition between xanthine dehydrogenase and xanthine oxidase. FEBS J 2015; 282:3075-90. [PMID: 25817260 PMCID: PMC4832347 DOI: 10.1111/febs.13277] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2014] [Revised: 03/09/2015] [Accepted: 03/21/2015] [Indexed: 01/24/2023]
Abstract
UNLABELLED Mammalian xanthine oxidoreductase can exist in both dehydrogenase and oxidase forms. Conversion between the two is implicated in such diverse processes as lactation, anti-bacterial activity, reperfusion injury and a growing number of diseases. We have constructed a variant of the rat liver enzyme that lacks the carboxy-terminal amino acids 1316-1331; it appears to assume an intermediate form, exhibiting a mixture of dehydrogenase and oxidase activities. The purified variant protein retained ~ 50-70% of oxidase activity even after prolonged dithiothreitol treatment, supporting a previous prediction that the C-terminal region plays a role in the dehydrogenase to oxidase conversion. In the crystal structure of the protein variant, most of the enzyme stays in an oxidase conformation. After 15 min of incubation with a high concentration of NADH, however, the corresponding X-ray structures showed a dehydrogenase-type conformation. On the other hand, disulfide formation between Cys535 and Cys992, which can clearly be seen in the electron density map of the crystal structure of the variant after removal of dithiothreitol, goes in parallel with the complete conversion to oxidase, resulting in structural changes identical to those observed upon proteolytic cleavage of the linker peptide. These results indicate that the dehydrogenase-oxidase transformation occurs rather readily and the insertion of the C-terminal peptide into the active site cavity of its subunit stabilizes the dehydrogenase form. We propose that the intermediate form can be generated (e.g. in endothelial cells) upon interaction of the C-terminal peptide portion of the enzyme with other proteins or the cell membrane. DATABASE Coordinate sets and structure factors for the four crystal structures reported in the present study have been deposited in the Protein Data Bank under the identification numbers 4YRW, 4YTZ, 4YSW, and 4YTY.
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Affiliation(s)
- Tomoko Nishino
- Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan
| | - Ken Okamoto
- Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan
| | - Yuko Kawaguchi
- Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan
| | - Tomohiro Matsumura
- Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan
| | - Bryan T Eger
- Department of Biochemistry, University of Toronto, ON, Canada
| | - Emil F Pai
- Department of Biochemistry, University of Toronto, ON, Canada
- Departments of Medical Biophysics and Molecular Genetics, University of Toronto, ON, Canada
- Campbell Family Institute for Cancer Research, Ontario Cancer Institute/University Health Network, Toronto, ON, Canada
| | - Takeshi Nishino
- Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo, Japan
- Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Japan
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Kelley EE. Dispelling dogma and misconceptions regarding the most pharmacologically targetable source of reactive species in inflammatory disease, xanthine oxidoreductase. Arch Toxicol 2015; 89:1193-207. [PMID: 25995007 DOI: 10.1007/s00204-015-1523-8] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2015] [Accepted: 04/27/2015] [Indexed: 01/04/2023]
Abstract
Xanthine oxidoreductase (XOR), the molybdoflavin enzyme responsible for the terminal steps of purine degradation in humans, is also recognized as a significant source of reactive species contributory to inflammatory disease. In animal models and clinical studies, inhibition of XOR has resulted in diminution of symptoms and enhancement of function in a number of pathologies including heart failure, diabetes, sickle cell anemia, hypertension and ischemia-reperfusion injury. For decades, XOR involvement in pathologic processes has been established by salutary outcomes attained from treatment with the XOR inhibitor allopurinol. This has served to frame a working dogma that elevation of XOR-specific activity is associated with enhanced rates of reactive species generation that mediate negative outcomes. While adherence to this narrowly focused practice of designating elevated XOR activity to be "bad" has produced some benefit, it has also led to significant underdevelopment of the processes mediating XOR regulation, identification of alternative reactants and products as well as micro-environmental factors that alter enzymatic activity. This is exemplified by recent reports: (1) identifying XOR as a nitrite reductase and thus a source of beneficial nitric oxide ((•)NO) under in vivo conditions similar to those where XOR inhibition has been assumed an optimal treatment choice, (2) describing XOR-derived uric acid (UA) as a critical pro-inflammatory mediator in vascular and metabolic disease and (3) ascribing an antioxidant/protective role for XOR-derived UA. When taken together, these proposed and countervailing functions of XOR affirm the need for a more comprehensive evaluation of product formation as well as the factors that govern product identity. As such, this review will critically evaluate XOR-catalyzed oxidant, (•)NO and UA formation as well as identify factors that mediate their production, inhibition and the resultant impact on inflammatory disease.
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Affiliation(s)
- Eric E Kelley
- Department of Anesthesiology and Vascular Medicine Institute, School of Medicine, University of Pittsburgh, W1357 BST, 200 Lothrop Street, Pittsburgh, PA, 15213, USA,
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Pathophysiology of circulating xanthine oxidoreductase: New emerging roles for a multi-tasking enzyme. Biochim Biophys Acta Mol Basis Dis 2014; 1842:1502-17. [DOI: 10.1016/j.bbadis.2014.05.022] [Citation(s) in RCA: 146] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2014] [Revised: 05/21/2014] [Accepted: 05/22/2014] [Indexed: 02/07/2023]
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Kaya MO, Kaya Y, Çelik G, Kurtuluş F, Arslan O, Güler ÖÖ. Differential in vitro inhibition studies of some cerium vanadate derivatives on xanthine oxidase. J Enzyme Inhib Med Chem 2014; 30:286-9. [PMID: 24964345 DOI: 10.3109/14756366.2014.920837] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
In this preliminary study, a new series of some cerium vanadate derivatives have been investigated as new type of inhibitors of xanthine oxidase (XO; E.C 1.17.3.2). XO is a superoxide-producing enzyme found normally in serum and the lungs, and its activity is concerned with several important health problems such as gout, severe liver damage, vascular dysfunction and injury, oxidative eye injury and renal failure. In this study, we present a critical overview of the effects of these novel type agents on XO with comparing the efficacy and safety profiles of allopurinol, the efficient classical inhibitor of XO.
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Affiliation(s)
- Mustafa Oğuzhan Kaya
- Department of Chemistry, Science and Art Faculty, Balikesir University , Balikesir , Turkey
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Agarwal A, Banerjee A, Banerjee UC. Xanthine oxidoreductase: a journey from purine metabolism to cardiovascular excitation-contraction coupling. Crit Rev Biotechnol 2011; 31:264-80. [PMID: 21774633 DOI: 10.3109/07388551.2010.527823] [Citation(s) in RCA: 63] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023]
Abstract
Xanthine oxidoreductase (XOR) is a ubiquitous complex cytosolic molybdoflavoprotein which controls the rate limiting step of purine catabolism by converting xanthine to uric acid. It is known that optimum concentrations of uric acid (UA) and reactive oxygen species (ROS) are necessary for normal functioning of the body. The ability of XOR to perform detoxification reactions, and to synthesize UA and reactive oxygen species (ROS) makes it a versatile intra- and extra-cellular protective "housekeeping enzyme". It is also an important component of the innate immune system. The enzyme is a target of drugs against gout and hyperuricemia and the protein is of major interest as it is associated with ischemia reperfusion (I/R) injury, vascular disorders in diabetes, cardiovascular disorders, adipogenesis, metabolic syndrome, cancer, and many other disease conditions. Xanthine oxidoreductase in conjugation with antibodies has been shown to have an anti-tumor effect due to its ability to produce ROS, which in turn reduces the growth of cancer tissues. Apart from this, XOR in association with nitric oxide synthase also participates in myocardial excitation-contraction coupling. Although XOR was discovered over 100 years ago, its physiological and pathophysiological roles are still not clearly elucidated. In this review, various physiological and pathophysiological functional aspects of XOR and its association with various forms of cancer are discussed in detail.
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Affiliation(s)
- Amit Agarwal
- Department of Pharmaceutical Technology (Biotechnology), National Institute of Pharmaceutical Education and Research, Punjab, India
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Jiang Y, Guo C, Fishel ML, Wang ZY, Vasko MR, Kelley MR. Role of APE1 in differentiated neuroblastoma SH-SY5Y cells in response to oxidative stress: use of APE1 small molecule inhibitors to delineate APE1 functions. DNA Repair (Amst) 2009; 8:1273-82. [PMID: 19726241 DOI: 10.1016/j.dnarep.2009.08.003] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2009] [Revised: 07/20/2009] [Accepted: 08/10/2009] [Indexed: 11/25/2022]
Abstract
Oxidative DNA damage has been implicated in a number of central nervous system pathologies. The base excision repair (BER) pathway is one of the most important cellular protection mechanisms that respond to oxidative DNA damage. Human apurinic (apyrimidinic) endonuclease/redox effector factor (APE1/Ref-1 or APE1) is an essential enzyme in the BER pathway and is expressed in both mitotic and post-mitotic cells in humans. In neurons, a reduction of APE1 expression increases chemotherapy-induced cytotoxicity, while overexpression of APE1 protects cells against the cytotoxicity. However, given the multiple functions of APE1, knockdown of total APE1 is not completely informative of whether it is the redox or DNA repair activity, or interactions with other proteins. Therefore, the use of selective small molecules that can block each function independent of the other is of great benefit in ascertaining APE1 function in post-mitotic cells. In this study, we chose differentiated SH-SY5Y cells as our post-mitotic cell line model to investigate whether a drug-induced decrease in APE1 DNA repair or redox activity contributes to the growth and survival of post-mitotic cells under oxidative DNA damaging conditions. Here, we demonstrate that overexpression of WT-APE1 or C65-APE1 (repair competent) results in significant increase in cell viability after exposure to H(2)O(2). However, the 177/226-APE1 (repair deficient) did not show a protective effect. This phenomenon was further confirmed by the use of methoxyamine (MX), which blocks the repair activity of APE1 that results in enhanced cell killing and apoptosis in differentiated SH-SY5Y cells and in neuronal cultures after oxidative DNA damaging treatments. Blocking APE1 redox function by a small molecule inhibitor, BQP did not decrease viability of SH-SY5Y cells or neuronal cultures following oxidative DNA damaging treatments. Our results demonstrate that the DNA repair function of APE1 contributes to the survival of nondividing post-mitotic cells following oxidative DNA damage.
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Affiliation(s)
- Yanlin Jiang
- Department of Pediatrics (Section of Hematology/Oncology), Herman B Wells Center for Pediatric Research, United States
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Concentraciones de ácido úrico en pacientes con preeclampsia y eclampsia. CLINICA E INVESTIGACION EN GINECOLOGIA Y OBSTETRICIA 2008. [DOI: 10.1016/s0210-573x(08)73069-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
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Tasci I, Deveci S, Isik AT, Comert B, Akay C, Mas N, Inal V, Yamanel L, Mas MR. Allopurinol in rat chronic pancreatitis: effects on pancreatic stellate cell activation. Pancreas 2007; 35:366-371. [PMID: 18090245 DOI: 10.1097/mpa.0b013e31806dbaaa] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
OBJECTIVES Activation of pancreatic stellate cells (PSCs) is a key event in pancreatic fibrosis. Xanthine oxidase-derived free radicals are involved in the mechanism of chronic pancreatitis (CP). We here searched the in vivo effects of allopurinol on PSC activation and its relation to tissue oxidative stress and histological findings in rat CP. METHODS Rat CP was induced with intraductal trinitrobenzene sulfonic acid in groups 1 (n = 16) and 2 (n = 10). Group 3 (n = 10) received intraductal saline. Four weeks after induction, group 1 received allopurinol (200 mg/kg, s.c.), and groups 2 and 3 received saline. After 4 weeks, oxidative stress parameters, histological evaluation, and immunostaining for alpha-smooth muscle actin (+) PSCs were performed in the pancreata. RESULTS Oxidative stress parameters improved significantly in group 1 compared with groups 2 and 3. Collagen deposition and lobular/sublobular atrophy were significantly lower in group 1 than in group 2. Alpha-smooth muscle actin (+) PSCs counts in group 1 were significantly lower than in group 2, and were in correlation with the degree of fibrosis and atrophy. CONCLUSIONS Allopurinol inhibits PSC activation in vivo. Pancreatic fibrosis can be prevented, at least in part, by antioxidant treatment through xanthine oxidase metabolism. Long-term use of allopurinol and its analogs may be considered in clinical trials with CP.
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Affiliation(s)
- Ilker Tasci
- Department of Internal Medicine, Gulhane School of Medicine, Ankara, Turkey
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Nagasaka S, Katoh H, Niu CF, Matsui S, Urushida T, Satoh H, Watanabe Y, Hayashi H. Protein kinase A catalytic subunit alters cardiac mitochondrial redox state and membrane potential via the formation of reactive oxygen species. Circ J 2007; 71:429-36. [PMID: 17322647 DOI: 10.1253/circj.71.429] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
BACKGROUND The identification of protein kinase A (PKA) anchoring proteins on mitochondria implies a direct effect of PKA on mitochondrial function. However, little is known about the relationship between PKA and mitochondrial metabolism. METHODS AND RESULTS The effects of PKA on the mitochondrial redox state (flavin adenine dinucleotide (FAD)), mitochondrial membrane potential (DeltaPsi(m)) and reactive oxygen species (ROS) production were investigated in saponin-permeabilized rat cardiomyocytes. The PKA catalytic subunit (PKAcat; 50 unit/ml) increased FAD intensities by 56.6+/-7.9% (p<0.01), 2'7'-dichlorofluorescin diacetate (DCF) intensities by 10.5+/-3.3 fold (p<0.01) and depolarized DeltaPsi(m) to 48.1+/-9.5% of the control (p<0.01). Trolox (a ROS scavenger; 100 micromol/L) inhibited PKAcat-induced DeltaPsi(m), FAD and DCF alteration. PKAcat-induced DeltaPsi(m) depolarization was inhibited by an inhibitor of the inner membrane anion channel (IMAC), 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS: 1 micromol/L) but not by an inhibitor of mitochondrial permeability transition pore (mPTP), cyclosporine A (100 nmol/L). CONCLUSIONS PKAcat alters FAD and DeltaPsi(m) via mitochodrial ROS generation, and PKAcat-induced DeltaPsi(m) depolarization was not caused by mPTP but rather by DIDS-sensitive mechanisms, which could be caused by opening of the IMAC. The effects of PKA on mitochondrial function could be related to myocardial function under the condition of extensive beta-adrenergic stimulation.
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Affiliation(s)
- Shiro Nagasaka
- Division of Cardiology, Internal Medicine III, Hamamatsu University School of Medicine, Hamamatsu, Japan
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Papezíková I, Lojek A, Cízová H, Cíz M. Alterations in plasma antioxidants during reperfusion of the ischemic small intestine in rats. Res Vet Sci 2006; 81:140-7. [PMID: 16297418 DOI: 10.1016/j.rvsc.2005.09.010] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2005] [Revised: 09/12/2005] [Accepted: 09/19/2005] [Indexed: 02/07/2023]
Abstract
The aim of this study was to evaluate the contribution of three plasma antioxidants (albumin, uric acid, SH groups) to the plasma total peroxyl radical-trapping antioxidant capacity (TRAP) in 2 and 4 h of intestinal reperfusion in rats. TRAP increased significantly both after 2 and 4 h of reperfusion. Neither albumin nor SH groups contributed significantly to this increase. TRAP was strongly influenced by the increase in uric acid concentration and also probably by the cell destruction caused by oxidative stress. Since the TRAP increase was accompanied by an increase in the level of thiobarbituric acid reactive substances (TBARS, a marker of lipid peroxidation), we can conclude that even such a large increase in TRAP is not sufficient to prevent the progression of lipid peroxidation and oxidative stress.
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Affiliation(s)
- Ivana Papezíková
- Laboratory of free radical pathophysiology, Institute of Biophysics, Academy of Sciences of The Czech Republic, Královopolská 135, 612 65 Brno, Czech Republic
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Bilbao E, de Cerio OD, Cajaraville MP, Cancio I. Cloning and expression pattern of peroxisomal enzymes in the mussel Mytilus galloprovincialis and in the thicklip grey mullet Chelon labrosus: generation of new tools to study peroxisome proliferation. MARINE ENVIRONMENTAL RESEARCH 2006; 62 Suppl:S109-12. [PMID: 16709435 DOI: 10.1016/j.marenvres.2006.04.004] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/09/2023]
Abstract
Aquatic organisms living in coastal/estuarine areas show peroxisome proliferation after exposure to different environmentally relevant pollutants. In order to generate new tools to assess peroxisome proliferation in aquatic animals, peroxisomal enzymes were cloned using degenerate primers in the mussel Mytilus galloprovincialis and in the thicklip grey mullet Chelon labrosus. Fragments of catalase (CAT), thiolase (THIO), polyamine oxidase (POX) and xanthine oxidoreductase (XOR) were cloned and their expression pattern studied in different tissues by semi-quantitative RT-PCR. In mussels, CAT, THIO, POX and XOR were expressed in digestive gland, mantle and gills while in mullets CAT, THIO and POX were expressed in liver, spleen, brain, heart, muscle and gills. XOR was mainly expressed in liver and heart. Mature mullets showed the highest expression of peroxisomal enzymes in liver, spleen and brain, while in juveniles expression was mainly found in muscle tissues, liver and gills. Laboratory experiments of exposure to organic pollutants are being performed to study the usefulness of these tools to study peroxisome proliferation in pollution biomonitoring programmes.
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Affiliation(s)
- Eider Bilbao
- Laboratory of Cell Biology and Histology, Department of Zoology and Animal Cell Biology, School of Science and Technology, University of the Basque Country, E-48080 Bilbo, Basque Country, Spain
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Ko JKS, Lam FYL, Cheung APL. Amelioration of experimental colitis by Astragalus membranaceus through anti-oxidation and inhibition of adhesion molecule synthesis. World J Gastroenterol 2005; 11:5787-94. [PMID: 16270386 PMCID: PMC4479677 DOI: 10.3748/wjg.v11.i37.5787] [Citation(s) in RCA: 67] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the protective effects of Astragalus membranaceus (Am) against hapten-induced colitis in male Sprague-Dawley rats as well as its underlying mechanism.
METHODS: Experimental colitis was induced in rats by enema administration of 2,4-dinitrobenzene sulfonic acid (DNBS). Rats were either pretreated with Am extract (2 or 4 g/kg, p.o. once daily) starting from 10 d before DNBS enema, or received Am post-treatment (2 or 4 g/kg, p.o. twice daily) on the three consecutive days following DNBS administration. Colonic lesion area and histological damage were determined, while the activities of myeloperoxidase (MPO) and xanthine oxidase, as well as reduced glutathione (GSH) content were measured in the excised colonic tissues. Besides, protein expression of inducible nitrite oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1) and P-selectin was also detected by Western blot analysis.
RESULTS: Our findings had shown that both macroscopic lesion area and histological colonic damage induced by DNBS were significantly reduced by both Am pre- and post-treatments. These were accompanied by attenuation of the elevated colonic MPO activity and downregulation of the iNOS, P-selectin, and ICAM-1 protein expression. Besides, deprivation of colonic GSH level under colitis condition was also preserved.
CONCLUSION: These results demonstrate that Am possesses both preventive and therapeutic potential in experimental colitis. The anti-inflammatory actions involve anti-oxidation along with inhibition of adhesion molecule synthesis in the colonic tissues.
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Affiliation(s)
- Joshua-Ka-Shun Ko
- School of Chinese Medicine, Hong Kong Baptist University, Kowloon Tong, Hong Kong, China.
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Bianciardi P, Scorza R, Ghilardi G, Samaja M. Xanthine oxido-reductase activity in ischemic human and rat intestine. Free Radic Res 2005; 38:919-25. [PMID: 15621709 DOI: 10.1080/10715760412331273430] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/26/2022]
Abstract
We measured time course and extent of xanthine dehydrogenase (XD) to xanthine oxidase (XO) conversion in ischemic human and rat intestine. To model normothermic no-flow ischemia, we incubated fresh biopsies for 0, 2, 4, 8 and 16h. At t = 0h, XO was less in humans than in rats (P < 0.0004), while XD was essentially the same (P = NS). After 16h incubation at 37 degrees C, there was no appreciable XD-to-XO conversion and no change in neither XO nor XD activity in human intestine. In contrast, the rat intestine had XO/(XO + XD) ratio doubled in the first 2h and then maintained that value until t = 16 h. In conclusion, no XO-to-XD conversion was appreciable after 16 h no-flow normothermic ischemia in human intestine; in contrast, XO activity in rats increased sharply after the onset of ischemia. An immunohistochemical labelling study shows that, whereas XO + XD expression in liver tissue is localised in both hepatocytes and endothelial cells, in the intestine that expression is mostly localised in epithelial cells. We conclude that XO may be considered as a major source of reactive oxygen species in rats but not in humans.
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Affiliation(s)
- Paola Bianciardi
- Dipartimento di Medicina, Chirurgia e Odontoiatria, University of Milan, San Paolo Hospital, via di Rudini' 8-20142 Milano, Italy
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Kozlov AV, Szalay L, Umar F, Kropik K, Staniek K, Niedermüller H, Bahrami S, Nohl H. Skeletal muscles, heart, and lung are the main sources of oxygen radicals in old rats. Biochim Biophys Acta Mol Basis Dis 2005; 1740:382-9. [PMID: 15949706 DOI: 10.1016/j.bbadis.2004.11.004] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2004] [Revised: 09/30/2004] [Accepted: 11/09/2004] [Indexed: 11/25/2022]
Abstract
The aim of this study was to compare rat tissues with respect to their reactive oxygen and nitrogen species (RONS) generating activities as a function of age. We quantified the RONS generation in vivo in young (6 months) and in old (30 months) male Sprague-Dawley rats using the recently developed spin trap 1-hydroxy-3-carboxy-pyrrolidine, applied intravenously. This spin trap reacts with superoxide radical and peroxynitrite yielding a stable spin adduct which is detectable by means of electron paramagnetic resonance (EPR) spectroscopy in frozen tissues. In old rats RONS generation was significantly increased compared to their young counterparts in the following order: blood<skeletal muscle<lung<heart, but did not change in intestine, brain, liver, and kidney. Experiments with isolated heart mitochondria showed a significant rate of RONS generation in succinate-supplemented mitochondria from old rats while no RONS were detected in mitochondria from young rats. This study identifies heart, lung, and skeletal muscle as the tissues with increased RONS formation as a function of age.
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Affiliation(s)
- Andrey V Kozlov
- Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Vienna, Austria.
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Nohl H, Gille L, Staniek K. Intracellular generation of reactive oxygen species by mitochondria. Biochem Pharmacol 2005; 69:719-23. [PMID: 15710349 DOI: 10.1016/j.bcp.2004.12.002] [Citation(s) in RCA: 168] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022]
Abstract
Mitochondria have bioenergetic properties that strongly suggest their involvement in the cellular formation of reactive oxygen species (ROS). Apparent confirmation of this process has come from work with isolated mitochondria, which have been shown to produce H(2)O(2) from dismutating superoxide radicals. Two different sites were reported to shuttle single electrons to oxygen out of the normal respiratory sequence. However, the mechanisms for ROS formation at these two sites are controversial. Arguments against mitochondrial ROS formation in the living cell are based on the fact that bioenergetic alterations may result from the mechanical removal of mitochondria from their natural environment. Furthermore, the invasive detection methods that are generally used may be inappropriate because of the possible interaction of the detection system with mitochondrial constituents. The use of non-invasive detection methods has proved that ROS formation does not occur unless changes in the physical state of the membrane are established. The aim of this commentary is to discuss critically the arguments in favor of mitochondria as the main intracellular source of ROS. The pros and cons of working with isolated mitochondria, as well as the detection methodology are carefully analyzed to judge whether or not the above assumption is correct. The conclusion that mitochondria are the main ROS generators in the cell contradicts the fact that ROS release was not observed. However, if electron flow from ubiquinol to the bc(1) complex is hindered due to changes in lipid fluidity, single electrons may transfer to dioxygen and produce H(2)O(2) via superoxide radicals.
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Affiliation(s)
- Hans Nohl
- Research Institute of Pharmacology and Toxicology, University of Veterinary Medicine Vienna, Veterinärplatz 1, A-1210 Vienna, Austria.
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Prusis P, Dambrova M, Andrianov V, Rozhkov E, Semenikhina V, Piskunova I, Ongwae E, Lundstedt T, Kalvinsh I, Wikberg JES. Synthesis and quantitative structure-activity relationship of hydrazones of N-amino-N'-hydroxyguanidine as electron acceptors for xanthine oxidase. J Med Chem 2004; 47:3105-10. [PMID: 15163191 DOI: 10.1021/jm031127c] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
A series of new N-hydroxyguanidines were synthesized and tested for electron acceptor activity on bovine milk xanthine oxidase using xanthine as reducing substrate. Manual inspection of the structure-activity data revealed that molecules containing nitro groups ("set A") show a different structure-activity relationship pattern compared to non-nitro compounds ("set B"). Accordingly separate QSAR models were built and validated for the two sets. Substantial differences were found in properties governing acceptor activity for the models, the only common property being sterical access to the imino nitrogen atom of the hydroxyguanidinimines. For set A molecules the presence of a nitro substituent at a certain distance range from the hydroxuguanidino group was most important. In addition, the presence of a nitro group in the ortho position interacting with NH(2) of the hydroxyguanidino group, and the mutual geometry of the phenyl ring, hydroxyguanidine, and imine groups was important for this set. By contrast, for set B molecules the acceptor activity was most influenced by the geometry of methoxy groups and the size and geometry of meta and para substituents of the phenyl ring.
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Affiliation(s)
- Peteris Prusis
- Department of Pharmaceutical Biosciences, Division of Pharmacology, Uppsala University, SE 751 24 Uppsala, Sweden
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27
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Martí R, Múrio E, Varela E, Bilbao I, Pascual C, Margarit C, Segura RM. Xanthine oxidoreductase and preservation injury in human liver transplantation. Transplantation 2004; 77:1239-45. [PMID: 15114092 DOI: 10.1097/01.tp.0000120384.52033.bc] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
BACKGROUND Preservation injury is a major cause of primary graft dysfunction in liver transplantation (LT). Oxidative damage is considered to be the first event leading to graft damage. Xanthine oxidoreductase (XOR) and neutrophil activation, two sources of reactive oxygen species, could play a role in the development of graft dysfunction. METHODS We determined activities of XOR forms, polymorphonuclear elastase (PMN-E), aminotransferases, and hyaluronic acid in plasma of 20 patients undergoing LT. Samples were taken from the radial artery (RA) before the anhepatic phase; from the portal vein (PV) before reperfusion; from graft caval effluent (CE) at reperfusion; and from RA, PV, and the hepatic vein (HV) 10 and 90 min postreperfusion. RESULTS The graft, but not recipient bowel, released XOR into blood (XOR in CE, median, 61.2 mU/g protein [range, 1.9-160.4 vs. undetectable in PV before reperfusion). Circulating XOR was transformed from dehydrogenase to reversible oxidase (XOrev) (XOrev-to-XOR ratio, 48.1% in CE and 65.1% in HV 90 min postreperfusion). Neutrophil activation was detected in the recipients before reperfusion, and in liver at early post-reperfusion (median PMN-E was 0.85 microg/g protein [range, 0.01-1.58] in RA before the anhepatic phase; 2.22 microg/g protein [range, 0.20-5.88] in PV prereperfu-sion; and 3.60 microg/g protein [range, 0.48-6.78] in HV 10 min postreperfusion). XOR, but none of the other markers, was higher in the CE of patients with moderate primary graft dysfunction than in those with slight primary graft dysfunction. CONCLUSIONS XOR release and neutrophil activation are produced during LT, and they are potentially injurious mechanisms associated with this therapy.
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Affiliation(s)
- Ramon Martí
- Biochemistry Department, Hospital Universitari Vall d'Hebron, Barcelona, Spain
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Ardan T, Kovaceva J, Cejková J. Comparative histochemical and immunohistochemical study on xanthine oxidoreductase/xanthine oxidase in mammalian corneal epithelium. Acta Histochem 2004; 106:69-75. [PMID: 15032331 DOI: 10.1016/j.acthis.2003.08.001] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022]
Abstract
We have previously found that xanthine oxidase (one form of xanthine oxidoreductase that generates reactive oxygen species, such as superoxide radicals and hydrogen peroxide) is present in corneal epithelium of normal rabbit eye. It was suggested that the reactive oxygen species contribute to additional eye damage related to prolonged continuous contact lens wear and irradiation of the eye with UV-B light. To further explore the potential danger of xanthine oxidase as a source of reactive oxygen species, we have examined in the present paper whether xanthine oxidoreductase and xanthine oxidase are present in corneal epithelium of other mammalian species, employing immunohistochemical and enzyme histochemical methods. In corneal epithelium of normal eyes of ox, pig, guinea-pig, and rat xanthine oxidoreductase activity was detected by the tetrazolium salt reduction method and xanthine oxidase activity was localized by a method based on cerium ions capturing hydrogen peroxide. For the immunohistochemical demonstration of the enzymes, rabbit anti-bovine xanthine oxidase antibody, rabbit anti-human xanthine oxidase antibody and monoclonal mouse anti-human xanthine oxidase/xanthine dehydrogenase/aldehyde oxidase antibody were used. The immunohistochemical and enzyme histochemical results show that xanthine oxidoreductase and xanthine oxidase are present both as proteins and as active enzymes in the corneal epithelium of all animals studied. It is hypothesized that under various pathological states, xanthine oxidase-generated reactive oxygen species might contribute to eye damage.
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Affiliation(s)
- Taras Ardan
- Department of Eye Histochemistry and Pharmacology, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Vídenská 1083, 14220, Prague 4, Czech Republic
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29
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Kruidenier L, Verspaget HW. Review article: oxidative stress as a pathogenic factor in inflammatory bowel disease--radicals or ridiculous? Aliment Pharmacol Ther 2002; 16:1997-2015. [PMID: 12452933 DOI: 10.1046/j.1365-2036.2002.01378.x] [Citation(s) in RCA: 283] [Impact Index Per Article: 12.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Virtually all inflammatory mediators investigated to date seem to be dysregulated in the inflamed intestinal mucosa of patients with inflammatory bowel disease. However, which of these are actually involved in the initiation and perpetuation of intestinal tissue damage is still not fully understood. Amongst these mediators are the reactive oxygen metabolites, produced in large amounts by the massively infiltrating leucocytes. These reactive oxygen metabolites are believed to constitute a major tissue-destructive force and may contribute significantly to the pathogenesis of inflammatory bowel disease. This paper provides a concise overview of reactive oxygen metabolite biochemistry, the types of cell and tissue damage potentially inflicted by them, and the endogenous antioxidants which should prevent these harmful effects. An up-to-date summary of the available human experimental data suggests that reactive oxygen metabolite-mediated injury is important in both the primary and downstream secondary pathophysiological mechanisms underlying intestinal inflammation. Nonetheless, how the individual components of the mucosal antioxidant enzymatic cascade respond to inflammatory conditions is a neglected area of research. This particular aspect of intestinal mucosal oxidative stress therefore merits further study, in order to provide a sound, scientific basis for the design of antioxidant-directed treatment strategies for inflammatory bowel disease patients.
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Affiliation(s)
- L Kruidenier
- Department of Gastroenterology and Hepatology, Leiden University Medical Center, Leiden, The Netherlands.
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30
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Martelin E, Lapatto R, Raivio KO. Regulation of xanthine oxidoreductase by intracellular iron. Am J Physiol Cell Physiol 2002; 283:C1722-8. [PMID: 12388055 DOI: 10.1152/ajpcell.00280.2002] [Citation(s) in RCA: 21] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Xanthine oxidoreductase (XOR) may produce reactive oxygen species and play a role in ischemia-reperfusion injury. Because tissue iron levels increase after ischemia, and because XOR contains functionally critical iron-sulfur clusters, we studied the effects of intracellular iron on XOR expression. Ferric ammonium citrate and FeSO(4) elevated intracellular iron levels and increased XOR activity up to twofold in mouse fibroblast and human bronchial epithelial cells. Iron increased XOR protein and mRNA levels, whereas protein and RNA synthesis inhibitors abolished the induction of XOR activity. A human XOR promoter construct (nucleotides +42 to -1937) was not induced by iron in human embryonic kidney cells. Hydroxyl radical scavengers did not block induction of XOR activity by iron. Iron chelation by deferoxamine (DFO) decreased XOR activity but did not lower endogenous XOR protein or mRNA levels. Furthermore, DFO reduced the activity of overexpressed human XOR but not the amount of immunoreactive protein. Our data show that XOR activity is transcriptionally induced by iron but posttranslationally inactivated by iron chelation.
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Affiliation(s)
- Eeva Martelin
- Hospital for Children and Adolescents, Research Laboratory, University of Helsinki, Biomedicum Helsinki, 00014 University of Helsinki, Finland.
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31
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Frederiks WM, Vreeling-Sindelárová H. Ultrastructural localization of xanthine oxidoreductase activity in isolated rat liver cells. Acta Histochem 2002; 104:29-37. [PMID: 11993848 DOI: 10.1078/0065-1281-00629] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
Xanthine oxidoreductase (XOR) can exist in a dehydrogenase form (XD) and an oxidase form (XO). The D-form uses NAD as cofactor and the O-form uses oxygen as second substrate and produces oxygen radicals. Both enzymes have a high affinity for hypoxanthine and xanthine as substrate and produce uric acid, a potent antioxidant. In the present study, XOR activity was demonstrated with the ferricyanide method in permeabilized isolated rat liver cells at the electron microscopical level. Moreover, ultrastructural localization of XO activity in these cells was studied with the cerium salt method. Activity of both XOR and XO was found in matrix and core of peroxisomes of rat liver parenchymal cells. Only XOR activity was present as well in the cytoplasm of rat liver parenchymal cells. In Kupffer cells and sinusoidal endothelial cells, XOR activity was demonstrated in vesicles and occasionally on granular endoplasmic reticulum. XO activity was not found in Kupffer cells and sinusoidal endothelial cells. The presence of uric acid oxidase activity in matrix and core of peroxisomes as was found previously suggests further breakdown of purines to allantoin in peroxisomes. It is suggested that the major function of XOR activity in the cytoplasm of rat liver parenchymal cells and in sinusoidal cells is not the production of oxygen radicals, but rather the production of uric acid which can act as a potent antioxidant.
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Affiliation(s)
- Wilma M Frederiks
- Academic Medical Center, University of Amsterdam, Department of Cell Biology and Histology, The Netherlands.
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32
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Abstract
Xanthine oxidoreductase (XOR) is a complex molybdoflavoenzyme, present in milk and many other tissues, which has been studied for over 100 years. While it is generally recognized as a key enzyme in purine catabolism, its structural complexity and specialized tissue distribution suggest other functions that have never been fully identified. The publication, just over 20 years ago, of a hypothesis implicating XOR in ischemia-reperfusion injury focused research attention on the enzyme and its ability to generate reactive oxygen species (ROS). Since that time a great deal more information has been obtained concerning the tissue distribution, structure, and enzymology of XOR, particularly the human enzyme. XOR is subject to both pre- and post-translational control by a range of mechanisms in response to hormones, cytokines, and oxygen tension. Of special interest has been the finding that XOR can catalyze the reduction of nitrates and nitrites to nitric oxide (NO), acting as a source of both NO and peroxynitrite. The concept of a widely distributed and highly regulated enzyme capable of generating both ROS and NO is intriguing in both physiological and pathological contexts. The details of these recent findings, their pathophysiological implications, and the requirements for future research are addressed in this review.
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Affiliation(s)
- Roger Harrison
- Department of Biology and Biochemistry, University of Bath, Bath, UK.
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Abstract
OBJECTIVES The aim of this study was to investigate the level and the form of xanthine oxidoreductase (XOR) in severely diseased human livers, to ascertain whether the modifications of the enzyme activity reported in experimental pathology also occur in human liver disease. METHODS Total, dehydrogenase, and oxidase activities of XOR were measured in samples of human liver removed for transplantation or partial hepatectomy. Samples included four groups: 1) histologically normal liver tissue, adjacent to metastases from extrahepatic tumors (controls), 2) liver with virus-related cirrhosis; 3) liver with virus-negative cirrhosis, and 4) hepatocellular carcinoma tissue (HCC). RESULTS The level of total XOR was significantly higher in liver with virus-related cirrhosis, but not in virus-negative cirrhosis, than in controls. In virus-positive cirrhosis, the total XOR activity correlated positively with the level of ALT. The percentage of XOR oxidase activity in cirrhotic liver, regardless of virus infection, correlated positively with aspartate amino-transferase, bilirubin concentration, and partial thromboplastin time, and negatively with prothrombin time. The activity of XOR was significantly lower in HCC than in control tissue or in a nonneoplastic area of the same liver. CONCLUSIONS Consistent with previous reports in experimental pathology, the level of XOR was increased in cirrhotic liver, in association with viral infection. This increment correlated with ALT, suggesting a relationship between XOR activity and the extent of liver injury caused by viral replication. The percentage of oxidase activity seems to be correlated with tissue damage and consequent liver impairment. The low XOR activity observed in HCC is consistent with reported experimental pathology.
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Affiliation(s)
- Fiorenzo Stirpe
- Department of Experimental Pathology, University of Bologna, Italy
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34
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Cighetti G, Bortone L, Sala S, Allevi P. Mechanisms of action of malondialdehyde and 4-hydroxynonenal on xanthine oxidoreductase. Arch Biochem Biophys 2001; 389:195-200. [PMID: 11339808 DOI: 10.1006/abbi.2001.2328] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Studies have been made on the possible involvement of malondialdehyde (MDA) and (E)-4-hydroxynon-2-enal (HNE), two terminal compounds of lipid peroxidation, in modifying xanthine oxidoreductase activity through interaction with the oxidase (XO) and/or dehydrogenase (XDH) forms. The effect of the two aldehydes on XO (reversible, XO(rev), and irreversible, XO(irr)) and XDH was studied using xanthine oxidase from milk and xanthine oxidoreductase partially purified from rat liver. The incubation of milk xanthine oxidase with these aldehydes resulted in the inactivation of the enzyme following pseudo-first-order kinetics: enzyme activity was completely abolished by MDA (0.5-4 mM), while residual activity (5% of the starting value) associated with an XO(irr) form was always observed when the enzyme was incubated in the presence of HNE (0.5-4 mM). The addition of glutathione to the incubation mixtures prevented enzyme inactivation by HNE. The study on the xanthine oxidoreductase partially purified from rat liver showed that MDA decreases the total enzyme activity, acting only with the XO forms. On the contrary HNE leaves the same level of total activity but causes the conversion of XDH into an XO(irr) form.
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Affiliation(s)
- G Cighetti
- Department of Medical Chemistry and Biochemistry, Faculty of Medicine, University of Milan, Via Saldini 50, 20133 Milan, Italy.
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35
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Martí R, Varela E, Pascual C, Segura RM. Determination of xanthine oxidoreductase forms: influence of reaction conditions. Clin Chim Acta 2001; 303:117-25. [PMID: 11163031 DOI: 10.1016/s0009-8981(00)00390-9] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
Xanthine oxidoreductase (XOR) has been implicated in ischaemia-reperfusion injury, and increases in this enzyme have been found in plasma of patients with different illnesses. The catalytic concentrations of the XOR forms found in plasma, using various reaction conditions, greatly differ in the related literature. We studied the effect of the assay conditions on the xanthine oxidation rate catalysed by the XOR forms. Our results demonstrate inhibition of XOR by the reaction products and a time-dependent decrease in the reaction rates of XOR forms. Substrate consumption and inhibition by the products did not account for this decrease. Determination at 60 min incubation leads to catalytic concentrations up to 80% lower for the XOR forms than those obtained at 10 min. We conclude that elimination of the reaction products (NADH, H(2)O(2) and O(2)) from the reaction mixture, and short incubation times, are necessary for accurate measurement of the XOR activities.
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Affiliation(s)
- R Martí
- Servei de Bioquímica, Hospital General Universitari Vall d'Hebron, 119-129, 08035 Barcelona, Spain
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36
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Martelin E, Palvimo JJ, Lapatto R, Raivio KO. Nuclear factor Y activates the human xanthine oxidoreductase gene promoter. FEBS Lett 2000; 480:84-8. [PMID: 11034305 DOI: 10.1016/s0014-5793(00)01909-8] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
To study the regulation of the human xanthine oxidoreductase (XOR) gene, we cloned 1.9 kb of the promoter region. In reporter gene assays, a construct encompassing nucleotides between 142 to +42 conferred maximal basal activity of the XOR promoter in 293T cells, in comparison with shorter (-92 to +42) or longer (up to -1937 to +42) constructs. The promoter activity was low in NIH-3T3 cells. The most active construct contained a putative CCAAT motif at -119 to -123. Electrophoretic mobility shift assays showed that this sequence binds the ubiquitous nuclear factor Y (NF-Y). Mutation of the CCAAT motif (CTGAT) abolished the NF-Y binding and considerably reduced the promoter activity. Our data suggest an important functional role for NF-Y in the transcriptional activation of the human XOR gene.
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Affiliation(s)
- E Martelin
- Hospital for Children and Adolescents, Research Laboratory, University of Helsinki, Finland.
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37
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Dambrova M, Baumane L, Kiuru A, Kalvinsh I, Wikberg JE. N-Hydroxyguanidine compound 1-(3,4-dimethoxy- 2-chlorobenzylideneamino)-3-hydroxyguanidine inhibits the xanthine oxidase mediated generation of superoxide radical. Arch Biochem Biophys 2000; 377:101-8. [PMID: 10775447 DOI: 10.1006/abbi.2000.1745] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
We here show that the novel N-hydroxyguanidine derivative PR5 (1-(3, 4-dimethoxy-2-chlorobenzylideneamino)-3-hydroxyguanidine) is acting as an alternative electron acceptor in xanthine oxidase catalyzed oxidation of xanthine. The reduction product is the corresponding guanidine derivative 1-(3, 4-dimethoxy-2-chlorobenzylideneamino)guanidine (PR9). The reaction occurs under both anaerobic and aerobic conditions. Moreover, EPR measurements show that the action of PR5 is associated with the inhibition of superoxide radical formation seen under aerobic conditions. PR5 also supports xanthine oxidase catalyzed anaerobic oxidation of NADH. Kinetic studies indicate that increasing xanthine concentration significantly increases the apparent K(m) of PR5, but it remains unaltered by changing NADH concentration. Moreover, the molybdenum center inhibitor allopurinol inhibits the PR5-sustained oxidation of xanthine and NADH equally well, whereas the flavin adenine dinucleotide site inhibitor diphenyliodonium (DPI) markedly inhibits only the PR5-sustained oxidation of NADH. We suggest that PR5 binds and becomes reduced at the molybdenum center of the xanthine oxidase. We also found that both PR5 and its reduction product PR9 can inhibit the oxygen-sustained xanthine oxidase reaction. The properties of PR5 suggest that it is a member of a novel class of compounds which we have termed xanthine oxidase electron acceptor-inhibitor drugs. The potential use of xanthine oxidase electron acceptor-inhibitors in the prevention of free radical mediated tissue damage in organ ischemia-reperfusion diseases is discussed.
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Affiliation(s)
- M Dambrova
- Department of Pharmaceutical Biosciences, Uppsala University, Uppsala, S-75124, Sweden.
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38
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Colpaert EE, Lefebvre RA. Interaction of hypoxanthine/xanthine oxidase with nitrergic relaxation in the porcine gastric fundus. Br J Pharmacol 2000; 130:359-66. [PMID: 10807674 PMCID: PMC1572077 DOI: 10.1038/sj.bjp.0703317] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022] Open
Abstract
The influence of hypoxanthine (HX)/xanthine oxidase (XO) on short-term [electrical field stimulation (EFS; 4 Hz) for 10 s and 3 min; bolus of exogenous NO (10(-5) M)] and long-term [EFS (4 Hz) and continuous NO-infusion for 20 min] nitrergic relaxations was investigated in circular muscle strips of the pig gastric fundus. HX (3x10(-4) M) / XO (64 mu ml(-1)) did not affect EFS for 10 s and 3 min; the short-lasting relaxation in response to a bolus of exogenous NO (10(-5) M) was changed into a biphasic relaxation with a small and short first phase followed by a larger and prolonged second phase. Cu/Zn superoxide dismutase (Cu/Zn SOD; 1000 u ml(-1)) and uricase (100 mu ml(-1)) respectively enhanced the amplitude of the first phase and diminished the amplitude of the second phase. Ascorbate (5x10(-4) M) and bilirubin (2x10(-4) M) prevented the prolonged component. Exposure to HX/XO during long-term EFS elicited a complete, stable reversal of relaxation starting after a delay. During continuous NO-infusion, HX/XO induced an immediate, complete but transient reversal. The antioxidants bilirubin, ascorbate, alpha-tocopherol, urate, glutathione and Cu/Zn SOD, the hydrogen peroxide degrading enzyme catalase, the hydroxyl radical scavengers dimethylsulphoxide and mannitol, and the cofactor flavin adenine dinucleotide did not influence the reversal induced by HX/XO during either EFS or NO-infusion. The cell-permeable manganese SOD mimetic EUK-8 modified the stable reversal during long-term EFS into a transient one. The results suggest that a nitrated uric acid derivative is responsible for the prolonged second phase in the relaxation to a bolus of exogenous NO in the presence of HX/XO. The exact underlying mechanism of the reversal induced by HX/XO during sustained relaxation remains unclear.
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Affiliation(s)
- E E Colpaert
- Heymans Institute of Pharmacology, Ghent University Medical School, De Pintelaan 185, B-9000 Ghent, Belgium
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Rasmussen JT, Rasmussen MS, Petersen TE. Cysteines involved in the interconversion between dehydrogenase and oxidase forms of bovine xanthine oxidoreductase. J Dairy Sci 2000; 83:499-506. [PMID: 10750108 DOI: 10.3168/jds.s0022-0302(00)74909-5] [Citation(s) in RCA: 20] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Mammalian xanthine oxidoreductase exists intracellularly in its dehydrogenase form. However, outside of this reducing milieu the enzyme quickly transforms into an oxidase form. Interconversion can be controlled by sulfhydryl reactive reagents, suggesting that disulfide bridging is linked to this phenomenon. The present work identified cysteines involved in the interconversion process. Purified enzyme was subjected to mild reduction with 1,4-dithioerythriol to regain dehydrogenase activity, and the accessible cysteines were labeled with specific radioactive alkylation reagents, iodoacetic acid. This partial alkylation stabilizes the dehydrogenase form, presumable by hindering formation of disulfide bond(s). Six of 38 cysteines were found to be labeled (residues 169, 170, 535, 992, 1317, and 1325). The significance of this labeling of bovine xanthine oxidoreductase is discussed in relation to structural knowledge about the enzyme, and especially by comparison with the AA sequences of avian and invertebrate enzymes, which do not undergo conversion. Cysteines 535 and 992 are the most likely marked residues to be involved in the interconversion, whereas the other cysteines are located too far from the cofactorbinding areas in xanthine oxidoreductase.
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Affiliation(s)
- J T Rasmussen
- Protein Chemistry Laboratory, University of Aarhus, Denmark.
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40
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Dillaman RM, Greenaway P, Linton SM. Role of the midgut gland in purine excretion in the robber crab, Birgus latro (Anomura: Coenobitidae). J Morphol 1999; 241:227-35. [PMID: 10461133 DOI: 10.1002/(sici)1097-4687(199909)241:3<227::aid-jmor5>3.0.co;2-g] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
White fecal strands of Birgus latro are composed of small spherules of uric acid with a mean diameter of 1.6 +/- 0.6 microm. Large numbers of membrane-bound spherules with concentric lamellae are present in the R cells of the midgut gland, so we suggest that lengths of white feces are produced by coordinated secretion of these spherules into the lumen of the midgut gland tubules. There are four cell types in the tubules with embryonic (E) cells at the distal tip, B cells in a narrow band at the distal end and R cells making up the bulk of the tubules and gland. F cells are sparsely scattered among the R cells. Midgut gland tissue was assayed for activities of xanthine dehydrogenase and xanthine oxidase, the two forms of xanthine oxidoreductase. Contrary to previous reports, we found that the midgut gland of B. latro contains only high activities of xanthine dehydrogenase. If proteinase inhibitors were omitted from the assays, however, significant activity of xanthine oxidase was measured, a result we regard as an artifact attributable to the partial conversion of xanthine dehydrogenase to xanthine oxidase by endogenous proteinases. R cells were demonstrated to contain peroxisomes, which may be involved in lipid metabolism rather than synthesis of uric acid.
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Affiliation(s)
- R M Dillaman
- School of Biological Sciences, University of New South Wales, Sydney, Australia
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The Effect of Nitecapone, a New Antioxidant, on Myocardial Function After Aortic Cross-clamping in Experimental Heart Ischemia. Int J Angiol 1999; 8:16-21. [PMID: 9826401 DOI: 10.1007/bf01616836] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022] Open
Abstract
During aortic cross-clamping, the myocardium suffers from global ischemia, which is followed by reperfusion after declamping. The generation of free oxygen radicals increases during reperfusion, resulting in arrhythmias and impaired cardiac function. This study was conducted to evaluate the effect of a novel antioxidant nitecapone (NC) on cardiac reperfusion injury in vivo. Twelve pigs were anesthetized and after sternotomy the aorta and the right atrium were cannulated for cardiopulmonary bypass. The heart was arrested with either +4 degreesC crystalloid cardioplegia alone in the control group (n = 6) or cardioplegia with NC (50 µM) added in the NC group (n = 6). Cardioplegia was added every 20 minutes. After 1 hour of aortic cross-clamping, blood samples for oxidative stress analysis were taken, and hemodynamic profile surveillance continued for 90 minutes. Heart rate (p = 0.04) and left ventricular end diastolic pressure (LVEDP) (p = 0.04) were significantly lower in the NC group than in the C group after aortic declamping. Cardiac output and myocardial contractility (dP/dtmax) were also enhanced in the group receiving NC, but the difference was not statistically significant. At 30 minutes after reperfusion, the coronary production (coronary sinus-aorta) of thiobarbituric acid reactive substances correlated inversely with cardiac output (r = -0.90, p = 0.001) and stroke volume (r = -0.82, p = 0.007). The effect of NC on lipid peroxidation seems to be modest and therefore the target of NC is unclear. NC would appear, however, to be a beneficial additive in the crystalloid cardioplegia in terms of functional recovery.
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Laakso J, Mervaala E, Himberg JJ, Teräväinen TL, Karppanen H, Vapaatalo H, Lapatto R. Increased kidney xanthine oxidoreductase activity in salt-induced experimental hypertension. Hypertension 1998; 32:902-6. [PMID: 9822451 DOI: 10.1161/01.hyp.32.5.902] [Citation(s) in RCA: 51] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Clinical and experimental studies have established an association between high sodium intake and arterial hypertension. The renal mechanisms resulting in impaired sodium excretion in hypertension-prone subjects are not clear. In hypertension-prone rats, high blood pressure results in increased renal mass and hemodynamic changes, both of which may alter renal oxygen distribution. Xanthine oxidoreductase (XOR) oxidizes ATP metabolites hypoxanthine and xanthine to urate. Because XOR is induced by hypoxia, we assessed kidney XOR activity in 2 models of salt-sensitive hypertension, spontaneously hypertensive rats (SHR) and Dahl salt-sensitive (Dahl S) rats. Increasing sodium intake from basal (0.08%) to high (2.56% wt/dry wt in the diet) increased renal XOR activity dose-dependently from 68+/-8 to 143+/-21 microU/mg protein in the Dahl S (P<0.05) but not in Dahl salt-resistant (Dahl R) rats. On basal and high sodium diets, SHR had higher renal XOR activity (101+/-10 and 134+/-26 microU/mg protein, respectively) than normotensive Wistar-Kyoto rats (55+/-2 and 58+/-6 microU/mg protein, P<0.05). Sodium restriction (0.02% wt/wt) downregulated kidney XOR activity in both Dahl S and R rats by nearly 40%. In SHR, allopurinol treatment totally inhibited renal XOR activity, but neither systolic blood pressure nor renal mass changed. The results suggest that renal XOR induction is a consequence of increased salt intake or the resulting hypertension. However, further studies on renal XOR activity during the development of hypertension are needed to assess the importance of XOR in the pathophysiology of arterial hypertension.
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Affiliation(s)
- J Laakso
- Institute of Biomedicine, Departments of Medical Chemistry, Pharmacology and Toxicology, University of Helsinki, Finland.
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Slavíková H, Lojek A, Hamar J, Dusková M, Kubala L, Vondrácek J, Cíz M. Total antioxidant capacity of serum increased in early but not late period after intestinal ischemia in rats. Free Radic Biol Med 1998; 25:9-18. [PMID: 9655516 DOI: 10.1016/s0891-5849(98)00030-6] [Citation(s) in RCA: 39] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
The ischemia of small intestine was induced in anesthetized Wistar rats by occluding the superior mesenteric artery for 45 min and then the reperfusion was set. Serum samples were obtained at the end of the ischemic period and also during early (1 to 4 h) and late postischemic period (1 to 4 d). The total antioxidant capacity (TRAP) of serum samples was evaluated using luminol enhanced chemiluminescence. The increased mobilization of phagocytic cells and the release of reactive oxygen species into the circulation was observed from the first and second hour of the postischemic period, respectively. Nevertheless, the activity of natural antioxidant mechanisms of serum was already elicited at the end of the ischemic period. Furthermore, the TRAP of serum increased with the increasing duration of early postischemic period. Among the antioxidants studied, urate and ascorbate concentrations exerted the highest correlation with TRAP, but 31.6% of the total antioxidant capacity remained for the activity of an unidentified antioxidant(s). After being exhausted, the TRAP of serum oscillated around the preoperation level at days 1-4 of the postischemic period. The increase in total antioxidant capacity of serum induced by oxidative stress was sufficient to prevent lipoperoxidation both in serum and intestinal tissue.
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Affiliation(s)
- H Slavíková
- Institute of Biophysics, Academy of Sciences of the Czech Republic, Brno.
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Parks DA, Skinner KA, Skinner HB, Tan S. Multiple organ dysfunction syndrome: Role of xanthine oxidase and nitric oxide. PATHOPHYSIOLOGY 1998. [DOI: 10.1016/s0928-4680(98)00008-x] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
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Pesonen EJ, Linder N, Raivio KO, Sarnesto A, Lapatto R, Höckerstedt K, Mäkisalo H, Andersson S. Circulating xanthine oxidase and neutrophil activation during human liver transplantation. Gastroenterology 1998; 114:1009-15. [PMID: 9558291 DOI: 10.1016/s0016-5085(98)70321-x] [Citation(s) in RCA: 47] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
BACKGROUND & AIMS Oxygen free radicals, generated by xanthine oxidase (XO) and activated leukocytes, are involved in reperfusion injury in experimental liver transplantation. The roles of XO and neutrophil activation during reperfusion in clinical liver transplantation were studied. METHODS In 10 patients undergoing liver transplantation, we assessed plasma concentrations of circulating XO by enzyme-linked immunosorbent assay (ELISA), the purine metabolites hypoxanthine, xanthine, and urate by high-performance liquid chromatography, lactoferrin by ELISA, and malondialdehyde fluorometrically up to 48 hours postoperatively. RESULTS During reperfusion after portal vein declamping, elevated plasma concentrations of XO (52.1 ng/mL [range, 8.0-440.1]), hypoxanthine (81.62 micromol/L [48.2-108.7]), xanthine (21.01 micromol/L [8.7-22.3]), and lactoferrin (532.6 ng/mL [370.4-1326.6]) were observed compared with the preoperative levels (0 ng/mL [0-12], 1.88 micromol/L [0.62-3.15], 0.95 micromol/L [0-0.41], and 164.3 ng/mL [73.7-334.1], respectively; all P < 0.05). No changes occurred in urate or malondialdehyde. After portal vein declamping, XO, hypoxanthine, and xanthine levels were substantially greater in the hepatic than portal vein (all P < 0.05). Marginal transhepatic differences occurred in lactoferrin. CONCLUSIONS Reperfusion during liver transplantation is associated with liberation of xanthine oxidase, hypoxanthine, and xanthine from the liver into the circulation. During reperfusion, intravascular neutrophil activation takes place in the hepatic circulation.
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Affiliation(s)
- E J Pesonen
- Children's Hospital, University of Helsinki, Helsinki, Finland.
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Skinner KA, Crow JP, Skinner HB, Chandler RT, Thompson JA, Parks DA. Free and protein-associated nitrotyrosine formation following rat liver preservation and transplantation. Arch Biochem Biophys 1997; 342:282-8. [PMID: 9186489 DOI: 10.1006/abbi.1997.0114] [Citation(s) in RCA: 49] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
Nitrotyrosine in human and animal tissues has been associated with pathologic conditions such as atherosclerosis, renal failure, and acute lung disease. In this study, free and protein-associated nitrotyrosine were determined in plasma and tissue samples using a dual-channel electrochemical detection method. Free nitrotyrosine was quantified in acetonitrile-extracted samples while protein-associated nitrotyrosine was determined in proteinase K-digested samples. In human plasma, total nitrotyrosine increased from 2.3 to 4.3 and 13.2 mumol/mol Tyr following addition of 0, 0.5, and 1 mM ONOO-. To determine if nitrotyrosine was produced during ex vivo hypothermic preservation, rat livers were stored in University of Wisconsin solution (UW) for 0, 6, or 8 h and reperfused for 3 h. Total nitro-tyrosine increased 359 and 908% after 6 and 8 h preservation compared to 0 h. To determine if nitrotyrosine was produced in vivo following hepatic ischemia, a rat preservation-transplantation model was utilized in which livers were flushed with cold UW (0-h group) or transplanted following 6 h hypothermic preservation in UW. Free nitrotyrosine increased from 15.7 +/- 0.3 in the 0-h group to 23.6 +/- 2.5 mumol/mol Tyr, 24 h posttransplant of 6-h preserved livers. Protein-associated nitrotyrosine increased from 9.5 +/- 1.1 in the 0-h group to 27.5 +/- 0.7 mumol/mol Tyr in the 6-h preservation-transplantation group. Protein-associated nitrotyrosine provides an integrative determination of nitration. Detection of free and protein-associated nitrotyrosine in biologic samples may allow insight into the role of .NO-derived oxidants in tissue injury associated with various pathologic conditions.
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Affiliation(s)
- K A Skinner
- Department of Anesthesiology, University of Alabama at Birmingham 35233, USA
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Abstract
Superoxide anion radicals have been implicated in a variety of pathological processes. Under physiological conditions, superoxide dismutase (SOD) is effectively able to disproportionate superoxide anions into hydrogen peroxide and dioxygen. Until now, no techniques have been available to localize SOD activity within tissues. In the present study, SOD activity was detected in different rat tissues using a thin film of xanthine oxidase between the glass slide and the unfixed cryostat section and a medium containing hypoxanthine as a source of electrons for the production of superoxide anions. The incubation medium also contained cerium ions to precipitate the hydrogen peroxide product and polyvinyl alcohol to prevent leakage of soluble and/or loosely bound enzymes from the sections into the incubation medium. The cerium perhydroxides that are formed were visualized for the light microscope in a second step using an incubation medium consisting of 3,3'-diaminobenzidine, cobalt ions, and hydrogen peroxide, which results in oxidation of the diaminobenzidine to the final insoluble blue reaction product. By this methodology, high enzyme activity was found not only in endothelial cells of liver and kidney but also in hepatocytes of liver, myocytes of heart, smooth and striated cells of muscle, acinar cells of pancreas, epithelial cells of kidney ducts, and epithelial cells of the small intestine and colon. These findings were largely in agreement with immunohistochemical data obtained using antibodies against the Cu/Zn- and Mn-SODs. However, high activity was also detected extra-cellularly at the surface of epithelia of trachea, esophagus, small intestine, and colon and at the extracellular matrices, cartilage, and connective tissues. We conclude from these latter data that the activity of the extracellular form of the dismutase is localized. The present method allows the analysis of all three types of known SOD activity (Cu/Zn, Mn, and extracellular) in different tissues and cell compartments.
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Affiliation(s)
- W M Frederiks
- University of Amsterdam, Department of Cell Biology and Histology, The Netherlands
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Ultrastructural localization of xanthine oxidase activity in the digestive tract of the rat. ACTA ACUST UNITED AC 1995. [DOI: 10.1007/bf00173844] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/26/2022]
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