Das 1817 erstmals schriftlich erwähnte Selen galt lange Zeit nur als toxisch und sogar als procancerogen, bis es 1957 von Schwarz und Foltz als essentielles Spurenelement erkannt wurde, dessen biologische Funktionen in Säugern durch Selenoproteine vermittelt werden. Die Familie der Glutathionperoxidasen nimmt hierbei eine wichtige Stellung ein. Für diese sind konkrete Funktionen und die dazugehörigen molekularen Mechanismen, welche über die von ihnen katalysierte Hydroperoxidreduktion und damit verbundene antioxidative Kapazität hinausgehen, bislang nur unzureichend beschrieben worden. <br><br> Die Funktion der gastrointestinalen Glutathionperoxidase (GI-GPx) wird als Barriere gegen eine Hydroperoxidabsorption im Gastrointestinaltrakt definiert. Neuen Erkenntnissen zufolge wird die GI-GPx aber auch in verschiedenen Tumoren verstärkt exprimiert, was weitere, bis dato unbekannte, Funktionen dieses Enzymes wahrscheinlich macht.<br> Um mögliche neue Funktionen der GI-GPx, vor allem während der Cancerogenese, abzuleiten, wurde hier die transkriptionale Regulation der GI-GPx detaillierter untersucht. Die Sequenzanalyse des humanen GI-GPx-Promotors ergab das Vorhandensein von zwei möglichen "antioxidant response elements" (ARE), bei welchen es sich um Erkennungssequenzen des Transkriptionsfaktors Nrf2 handelt. Die meisten der bekannten Nrf2-Zielgene gehören in die Gruppe der Phase-II-Enzyme und verfügen über antioxidative und/oder detoxifizierende Eigenschaften. Sowohl auf Promotorebene als auch auf mRNA- und Proteinebene konnte die Expression der GI-GPx durch typische, in der Nahrung enthaltene, Nrf2-Aktivatoren wie z.B. Sulforaphan oder Curcumin induziert werden. Eine direkte Beteiligung von Nrf2 wurde durch Cotransfektion von Nrf2 selbst bzw. von Keap1, das Nrf2 im Cytoplasma festhält, demonstriert. Somit konnte die GI-GPx eindeutig als Nrf2-Zielgen identifiziert werden. Ob sich die GI-GPx in die Gruppe der antiinflammatorischen und anticancerogenen Phase-II-Enzyme einordnen lässt, bleibt noch zu untersuchen. <br><br> Die Phospholipidhydroperoxid Glutathionperoxidase (PHGPx) nimmt aufgrund ihres breiten Substratspektrums, ihrer hohen Lipophilie und ihrer Fähigkeit, Thiole zu modifizieren, eine Sonderstellung innerhalb der Familie der Glutathionperoxidasen ein. Mit Hilfe eines PHGPx-überexprimierenden Zellmodells wurden deshalb Beeinflussungen des zellulären Redoxstatus und daraus resultierende Veränderungen in der Aktivität redoxsensitiver Transkriptionsfaktorsysteme und in der Expression atheroskleroserelevanter Adhäsionsmoleküle untersucht. Als Transkriptionsfaktoren wurden NF-kB und Nrf2 ausgewählt. Die Bindung von NF-kB an sein entsprechendes responsives Element in der DNA erfordert das Vorhandensein freier Thiole, wohingegen Nrf2 durch Thiolmodifikation von Keap1 freigesetzt wird und in den Kern transloziert. Eine erhöhte Aktivität der PHGPx resultierte in einer Erhöhung des Verhältnisses von GSH zu GSSG, andererseits aber in einer verminderten Markierbarkeit freier Proteinthiole. PHGPx-Überexpression reduzierte die IL-1-induzierte NF-kB-Aktivität, die sich in einer verminderten NF-kB-DNA-Bindefähigkeit und Transaktivierungsaktivität ausdrückte. Auch war die Proliferationsrate der Zellen vermindert. Die Expression des NF-kB-regulierten vaskulären Zelladhäsionsmoleküls, VCAM-1, war ebenfalls deutlich verringert. Umgekehrt war in PHGPx-überexprimierenden Zellen eine erhöhte Nrf2-Aktivität und Expression der Nrf2-abhängigen Hämoxygenase-1 zu verzeichnen. Letzte kann für die meisten der beobachteten Effekte verantwortlich gemacht werden.<br><br> Die hier dargestellten Ergebnisse verdeutlichen, dass eine Modifizierung von Proteinthiolen als wichtige Determinante für die Regulation der Expression und Funktion von Glutathionperoxidasen angesehen werden kann. Entgegen früheren Vermutungen, welche oxidative Vorgänge generell mit pathologischen Veränderungen assoziierten, scheint ein moderater oxidativer Stress, bedingt durch eine transiente Thiolmodifikation, durchaus günstige Auswirkungen zu haben, da, wie hier dargelegt, verschiedene, miteinander interagierende, cytoprotektive Mechanismen ausgelöst werden. Hieran wird deutlich, dass sich "antioxidative Wirkung" oder "oxidativer Stress" keineswegs nur auf "gute" oder "schlechte" Vorgänge beschränken lassen, sondern im Zusammenhang mit den beeinflussten (patho)physiologischen Prozessen und dem Ausmaß der "Störung" des physiologischen Redoxgleichgewichtes betrachtet werden müssen. / Selenium was discovered in 1817 by the Swedish chemist Berzelius and was for a long time considered as being toxic and even procarcinogenic. In 1957, however, Schwarz and Foltz realized that selenium is an essential trace element which elicits its biological functions in mammals as a structural component of selenoproteins among which the family of glutathione peroxidases plays a dominant role. Glutathione peroxidases reduce hydroperoxides to the corresponding alcohols and contribute to the antioxidative capacity of a cell. However, other functions of glutathione peroxidases and the according molecular mechanisms have hardly been described.>br><br> The gastrointestinal glutathione peroxidase (GI-GPx) is believed to build a barrier against the absorption of foodborne hydroperoxides. In addition, GI-GPx expression is increased in different tumors. This indicates further, still unknown, functions of this enzyme.<br> In order to elucidate new possible functions of GI-GPx, especially during carcinogenesis, the transcriptional regulation of GI-GPx was analyzed in more detail. An analysis of the GI-GPx promoter sequence revealed the presence of two putative "antioxidant response elements" (ARE) which are recognition sites for the transcription factor Nrf2. Most of the known Nrf2 target genes either belong to the group of phase-II detoxification enzymes or possess antioxidative and/or detoxifying properties. On promoter level as well as on mRNA- and protein level the expression of GI-GPx was induced by typical Nrf2-activating compounds such as sulforaphane or curcumin that are contained in the diet. A direct involvement of Nrf2 was demonstrated by cotransfection of Nrf2 itself or by cotransfection of Keap1 which retains Nrf2 in the cytosol. Thus, the GI-GPx gene was unequivocally identified as a new target for Nrf2. Whether GI-GPx also belongs in the category of antiinflammatory and anticarcinogenic enzymes remains to be elucidated.<br><br> The phospholipid hydroperoxide glutathione peroxidase (PHGPx) is exceptional among the glutathione peroxidases because of its broad range of substrates, its high lipophilicity, and its ability to modify protein thiols. With PHGPx-overexpressing cells, the influence of PHGPx on the cellular redox state and on resulting changes in the activity of redox-sensitive transcription factors and on the expression of proatherogenic adhesion molecules was analyzed. For this, the redox-sensitive transcription factors NF-kB and Nrf2 were chosen. NF-kB requires free thiols for being able to bind to its responsive element within the DNA, whereas Nrf2 is released from Keap1 and translocates to the nucleus upon a modification of protein thiols. PHGPx-overexpression resulted in an increase in the ratio of GSH to GSSG, in a reduced amount of intracellular protein thiols, and in a diminished proliferation rate. Furthermore, PHGPx-overexpressing cells displayed a reduced IL-1-dependent NF-kB activity as was assessed by a reduced NF-kB DNA-binding ability and activity of a NF-kB-driven reporter gene. In addition, the expression of the NF-kB-dependent vascular cell adhesion molecule (VCAM-1) was also inhibited by overexpression of PHGPx. On the other hand, PHGPx-overexpressing cells displayed an increased activity of Nrf2 that was accompanied by an increased expression of the Nrf2-dependent heme oxygenase-1. Heme oxygenase-1 most likely is responsible for most of the aforementioned effects.<br><br> The data presented here show that a modification of protein thiols can be regarded as an important determinant for the regulation and for the functions of glutathione peroxidases. In contrast to the previous assumption that oxidative processes are always linked to pathologic changes, a moderate oxidative stress seems to have beneficial effects, because it triggers different cytoprotective mechanisms. It can be concluded that the terms "antioxidative effect" or "oxidative stress" cannot simply be restricted to "good" or "bad" processes, but need to be seen in context with the modulated (patho)physiological processes and the degree of "disturbance" of the physiologic redox balance.

Women with pregestational diabetes have a 2-5 fold increased risk of giving birth to malformed babies compared with non-diabetic women. Diabetes-induced oxidative stress in maternal and embryonic tissues has been implicated in the teratogenic process. The malformations are likely to be induced before the seventh week of pregnancy, when the yolk sac is partly responsible for the transfer of metabolites to the embryo, and the uterine blood flow to the implantation site determines the net amount of nutrients available to the conceptus. We aimed to evaluate the effect on embryogenesis caused by a diabetes-induced disturbance in yolk sac morphology, uterine blood flow or altered maternal antioxidative status in conjunction with a varied severity of the maternal diabetic state. We investigated to which extent maternal diabetes with or without folic acid (FA) supplementation affects mRNA levels and protein distribution of ROS scavenging enzymes (SOD, CAT, GPX), vascular endothelial growth factor-A (Vegf-A), folate binding protein-1 (Folbp-1), and apoptosis associated proteins (Bax, Bcl-2, Caspase-3) in the yolk sacs of rat embryos on gestational days 10 and 11. We found that maternal diabetes impairs, and that FA supplementation restores, yolk sac vessel morphology, and that maternal diabetes is associated with increased apoptotic rate in embryos and yolk sacs, as well as impaired SOD gene expression. We assessed uterine blood flow with a laser-Doppler-flow-meter and found increased blood flow to implantation sites of diabetic rats compared with controls. Furthermore, resorbed and malformed offspring showed increased and decreased blood flow to their implantation sites, respectively. In mice with genetically altered CuZnSOD levels, maternal diabetes increased embryonic dysmorphogenesis irrespective of CuZnSOD expression. We thus found the maternal diabetic state to be a major determinant of diabetic embryopathy and that the CuZnSOD status exerts a partial protection for the embryo in diabetic pregnancy.

Although considered as a poison for long time, the importance of selenium as an essential trace element is now well recognized. In proteins, the redox active selenium moiety is incorportated as selenocysteine (Sec), the 21st amino acid. In mammals, selenium exerts its redox activities through several selenocysteine-containing enzymes, which include glutathione peroxidase (GPx), iodothyronine deiodinase (ID) and thioredoxin reductase (TrxR). Although these enzymes have Sec in their active sites, they catalyze completely different reactions and their substrate specificity and cofactor or co-substrate systems are significantly different. The most widely studied selenoenzyme GPx protects various organisms from oxidative stresses by catalyzing the reduction of hydroperoxides by using glutathione (GSH) as cofactor. The chemical aspects of the reduction of hydroperoxide by GPx have been extensively studied with the help of synthetic selenium and tellurium compounds. For example, 2-phenyl, 1, 2-benzoisoselenazol-3(2H)-one, commonly known as ebselen exhibits significant GPx activity by using GSH as cofactor. The anti-inflammatory, antiatherosclerotic and cytoprotective properties of ebselen have led to the design and synthesis of nex GPx mimics for potential therapeutic applications. In the first chapter, the importance of selenium in biochemistry in general and the function of selenoenzyme GPx and its synthetic mimics in particular are discussed. In the second chapter, the importance of ebselen as a GPx mimic and how thiol exchange reaction in the selenenyl sulfide intermediate deactivates its catalytic cycle and the possible ways to overcome thiol exchange reaction are described. The third chapter deals with the first synthetic chemical model that effectively mimics the unusual cyclization of sulfenic acid to a sulfenyl amide in protein Tyrosien Phosphatase 1B(PTP1B). PTP1B is a cysteine containing enzyme where the sulfenic acid (PTP1B-SOH) intermediate produced in response to its oxidation by H2O2 is rapidly converted into a sulfenyl amide species, in which sulfur atom of the catalytic cysteine is covalently bonded to the main chain nitrogen of an adjacent serine residue. This unusual protein modification in PTP1B has been proposed to protect the sulfur centre from irreversible oxidation to sulfinic acid and and sulfonic acids. In the fourth chapter, it is shown that not only the catalytic efficiency of ebselen but also its phosphatase like behavior is important for its antioxidant activity. Ebselen is regenerated from selenenic acid (R-SeOH) under a verity of conditions, which protects its selenium centre from irreversible oxidation and thus reduces its toxicity. The fifth chapter deals with spirodizaselenurane and Spirodiazatellurane. Although the chemistry of spirodioxyselenuranes and spirodiazasulfuranes has been studied extensively due to their interesting structural and stereochemical properties, there is no example of stable spirodiazaselenurane and its tellurium analogues. In the fifth chapter, the synthesis, structure and GPx-like activity of the spirodizzaselenurane and spirodiazatellurane are discussed. In summary, the synthetic sulfenic acids and seleneric acids undergo cyclization to their corresponding sulfenyl amides and selenenyl amides and thus protect their sulfur and selenium centers from irreversible inactivation. We have also observed that selenoxides and telluroxides with nearby amide moieties undergo cyclization to their corresponding cyclic spiro compounds. This unusual transformation of sulfenic acids has been recently discovered in PTP1B. As the redox regulation cycle of PTP1B and the catalytic cycle of GPx are similar we believe that GPx may involve a selenenyl amide intermediate in its catalytic cycle.

Reactive oxygen species (ROS) such as superoxide radical anion (O2•¯), hydroxylradical (OH•), hydrogen peroxide (H2O2) and peroxynitrite (ONOO-) that are produced during the metabolism of oxygen under oxidative stress in aerobic organisms destroy several key biomolecules and lead to a number of disease states. Mammalian systems possess several effective defense mechanisms including antioxidant enzymes to detoxify these ROS. The selenocysteine-containing Glutathione peroxidase (GPx) is particularly an efficient enzyme in the detoxification of H2O2 and other hydroperoxides by using glutathione (GSH) as cofactor. The chemistry at the active siteof GPx has been extensively investigated with the help of synthetic selenium compounds. Although the anti-inflammatory compound ebselen(2-phenyl-1,2-benzoisoselenazol-3(2H)-one) is undergoing phase III clinical trial as antioxidant, the chemistry of ebselen is still not understood. The present study on a number of ebselen derivatives with various N-substitutions reveals that the substitution at the N atom is important for the antioxidant activity. This study also suggests that the nature for thiol cofactor has a dramatic effect on the GPx activity of ebselen derivatives. It has been shown that ebselen exhibits very poor catalytic activity in the presence of aromatic thiols mainly due to strong Se….O nonbonded interactions that lead to extensive thiol exchange reactions in the selenenyl sulfide intermediate. To prevent the se….O interactions, a series of tertiary amide-based diselenides have been synthesized along with their secondary amide counterparts. Detailed structure-activity correlation studies reveal that the GPx-like activity of the sec-amide-based compounds can be significantly enhanced by the substitution at the free-NH group of sec-amide functionality. The N,N-dialkylbenzylamine-based diselenides exhibit their catalytic activities via the generation of selenols which was confirmed by the reaction with anti-arthritic gold(I) compounds. Interestingly, the replacement of the hydrogen atom at the 6th position of the benzene ring of N,N-dialkylbenzylamine-based diselenides by a methoxy group prevents the thiol exchange reactions mainly be weakening the Se…N interactions and thus enhances the GPx activity. On the other hand, the catalytic activity of the tert-amine-based diselenides can also be increased by replacing the tert-amino groups with the corresponding sec-amine moieties. It has been observed that the basic amino group in the amine-based diselenides deprotonates the selenol and also the thiol cofactor, which is crucial for the higher catalytic activities of the amine-based compounds. Peroxynitrite (PN, ONOO), a strong nitrating agent, is known to inactivate a number of proteins, enzymes and other biomolecules by nitration of tyrosine residues. In this study, we have shown that the commonly used antithyroid drugs and their analogues inhibit protein tyrosine nitration. This study reveals that antithyroid agents having PN scavenging activity may be beneficial of hyperthyroidism as these compounds may protect the thyroid gland from nitrative or nitrosative stress.

Στόχος της παρούσας μελέτης ήταν να μελετήσει αν το εκχύλισμα των στιγμάτων του Crocus sativus L. αναστέλλει την επαγόμενη από σεληνιώδες νάτριο ανάπτυξη καταρράκτη σε ένα in vivo πείραμα με νεογνά επίμυων του γένους Wistar. Μέθοδοι: Τα νεογνά των επίμυων κατατάχθηκαν τυχαία σε 3 ομάδες. Ομάδα Ι (ομάδα μαρτύρων) όπου χορηγήθηκε υποδόρια φυσιολογικός ορός τη 10η ημέρα της ζωής. Ομάδα ΙΙ (ομάδα σεληνιώδους νατρίου) στην οποία χορηγήθηκε υποδόρια σεληνιώδες νάτριο (20 µmol/kg σωματικού βάρους) τη 10η ημέρα της ζωής. Ομάδα ΙΙΙ (ομάδα σεληνιώδους νατρίου και εκχυλίσματος στιγμάτων Crocus sativus L.) στην οποία εκτός από το σεληνιώδες νάτριο τη 10η ημέρα της ζωής χορηγήθηκε και εκχύλισμα στιγμάτων του Crocus sativus L. (60 mg/kg σωματικού βάρους) την 9η και 12η ημέρα της ζωής. Την 21η ημέρα της ζωής οι επίμυες θυσιάστηκαν και οι κρυσταλλοειδείς φακοί απομονώθηκαν και εξετάστηκαν για την εμφάνιση καταρράκτη. Ακολούθησε προσδιορισμός στους κρυσταλλοειδείς φακούς της δραστικότητας των αντιοξειδωτικών ενζύμων δισμουτάση του σουπεροξειδίου (SOD), της υπεροξειδάσης της γλουταθειόνης (GPx) καθώς και της καταλάσης (CAT). Προσδιορίστηκαν επίσης τα επίπεδα της γλουταθειόνης στους φακούς. Επιπλέον, μετρήθηκαν τα επίπεδα της μηλονικής διαλδεΰδης (MDA), ως δείκτη υπεροξείδωσης των λιπιδίων, καθώς και η συγκέντρωση των ελεύθερων σουλφυδρυλομάδων, ως δείκτη οξειδωτικής βλάβης των πρωτεϊνών, στους κρυσταλλοειδείς φακούς των επίμυων. Η επίδραση των χορηγούμενων παραγόντων στο πρωτεϊνικό προφίλ των φακών εκτιμήθηκε μέσω προσδιορισμού του λόγου των υδατοδιαλυτών προς τις μη υδατοδιαλυτές πρωτεΐνες του φακού. Τέλος έγινε ανάλυση των υδατοδιαλυτών πρωτεϊνών με ηλεκτροφόρηση σε πήκτωμα πολυακρυλαμιδίου. Αποτελέσματα: Το εκχύλισμα αποξηραμένων στιγμάτων του Crocus sativus L. επέδειξε σημαντική προστασία έναντι στην επαγόμενη από σεληνιώδες νάτριο καταρρακτογένεση στο in vivo πειραματικό μοντέλο που χρησιμοποιήσαμε. Οι μέσες τιμές των δραστικοτήτων των αντιοξειδωτικών ενζύμων SOD, GPx, CAT καθώς και της συγκέντρωσης της γλουταθειόνης αυξήθηκαν σημαντικά στην ομάδα που έλαβε εκχύλισμα στιγμάτων του Crocus sativus L. σε σύγκριση με την ομάδα των επίμυων που δέχθηκε μόνο την τοξική δράση του σεληνιώδους νατρίου. Το εκχύλισμα των στιγμάτων του Crocus sativus L. απέτρεψε σε σημαντικό βαθμό την υπεροξείδωση των λιπιδίων καθώς και την οξειδωτική βλάβη στις πρωτεΐνες του φακού. Επίσης απέτρεψε την πρωτεόλυση των υδατοδιαλυτών πρωτεϊνών του φακού. Συμπεράσματα: Χορήγηση εκχυλίσματος αποξηραμένων στιγμάτων του Crocus sativus L. απέτρεψε την επαγόμενη από το σεληνιώδες νάτριο καταρρακτογένεση σε νεογνά επίμυων του γένους Wistar πιθανώς μέσω ενίσχυσης της αντιοξειδωτικής άμυνας του κρυσταλλοειδούς φακού, μέσω αναστολής του βαθμού της υπεροξείδωσης των λιπιδίων, μέσω προστασίας των σουλφυδρυλομάδων των πρωτεϊνών καθώς και μέσω αναστολής της πρωτεόλυσης των υδατοδιαλυτών πρωτεϊνών του φακού. Αυτά τα ευρήματα τονίζουν τις πιθανές αντικαταρρακτογενετικές δράσεις των αποξηραμένων στιγμάτων του Crocus sativus L. οι οποίες αποδίδονται στις αντιοξειδωτικές τους ιδιότητες. / The present study sought to investigate whether Crocus sativus L. stigmas extract prevents selenite-induced cataractogenesis in vivo and to study the possible protective mechanism. Methods: Wistar rat pups were randomized into 3 groups. Group I (control) received subcutaneous injection of normal saline on postnatal day 10. Groups II (selenite treated) and III (selenite and Crocus sativus L. treated) received subcutaneous injection of sodium selenite (20 µmol/kg body weight) on postnatal day 10. Group III received intraperitoneal injections of Crocus sativus L. stigmas extract (60 mg/kg body weight) on postnatal days 9 and 12. On postpartum day 21, rats were sacrificed and the lenses were isolated and examined for cataract formation. Activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) and glutathione levels, as markers of antioxidant defense, were measured in the isolated lenses. Levels of the indicator of lipid peroxidation, malondialdehyde (MDA), and protein oxidation (sulfhydryl content) in lens were also determined. Effect of the different treatments on lens’ protein profile was evaluated with the estimation of soluble to insoluble protein ratio and SDS-PAGE analysis of water-soluble fraction (WSF) of lens proteins. Results: Crocus sativus L. stigmas extract demonstrated significant protection against selenite-induced cataractogenesis in vivo. The mean activities of SOD, GPx, CAT and glutathione levels were significantly increased in group III compared to the selenite-treated group. Crocus sativus L. stigmas extract significantly prevented selenite-induced lipid peroxidation, protein oxidation, as well as proteolysis and insolubilization of the lens WSF. Conclusions: Crocus sativus L. stigmas extract prevented selenite-induced cataract formation in Wistar rats possibly by reinforcement of antioxidant status, reduction of the intensity of lipid peroxidation, protection of the sulfhydryl groups, and inhibition of proteolysis of the lens WSF. These findings highlight the anti-cataractogenic potential of Crocus sativus L. stigmas by virtue of their antioxidant properties.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The objective of the present work was to evaluate the effect of dietary linseed oil with or without injection of sodium selenite upon fatty acid profile and oxidative stability of milk from dairy cows as well as the acceptance of the milk and the increasing order of preference by the evaluators. Chapter 1 describes the experiment where fourteen cows were allocated into four treatments: Group 1 which received daily 400 mL of linseed oil (LIN); Group 2, 400 mL of linseed oil + 0.2 mg/BW sodium selenite IM (LINSe); Group 3, untreated controls (C). The oil was supplied daily after 15 days of de single application of sodium selenite. Treatments lasted 4 weeks. Linseed oil supplemented animals produced milk with higher levels of conjugated linoleic acid and omega 3 but also more susceptible to oxidation. The application of sodium selenite was effective to prevent premature oxidation of milk. Chapter 2 describes the acceptance and increasing order of preference by the evaluators. The evaluators were not able to identify differences in color, odor and flavor among samples of milk from treated and control groups in relation to a known standard. The inclusion of linseed oil on the cows diet promotes and increase in CLA and omega 3 in milk, which in turn becomes more susceptible to oxidation, requiring the use of antioxidants. Even though causing biochemicals alterations, the addition of 400 mL daily of linseed oil in the diet of dairy cows is not capable of causing sensory changes in milk. / O presente trabalho descreve as avaliações feitas no leite de vacas leiteiras submetidas a suplementação com óleo de linhaça na dieta, com ou sem injeção de selenito de sódio, quanto ao perfil de ácidos graxos, estabilidade oxidativa e em relação às propriedades organolépticas do leite. O capítulo 1 descreve o experimento em que catorze vacas foram distribuídas em três tratamentos: Grupo 1, que recebeu diariamente 400 mL de óleo de linhaça (LIN); Grupo 2, 400 mL de óleo de linhaça + 0,2 mg / Kg de selenito de sódio IM (LINSe) e Grupo 3 controles não tratados (C). O óleo foi fornecido diariamente após 15 dias da aplicação única de selenito de sódio, e o experimento teve duração de quatro semanas. Foram feitas análises do perfil de ácidos graxos e de reações ao ácido tiobarbitúrico, que mede a estabilidade oxidativa do produto. Os animais suplementados com o óleo de linhaça produziram leite com altos níveis de ácido linoleico conjugado e de ômega 3, contudo, mais suscetível à oxidação. A aplicação injetável de selenito de sódio mostrou-se eficaz ao impedir a oxidação prematura do leite. O capítulo 2 descreve o experimento onde se verificou a aceitação e a ordem crescente de preferência pelos avaliadores, de amostras de leite, através de análises de cor, odor e sabor. Catorze vacas foram distribuídas em três tratamentos: Grupo 1, que recebeu diariamente 400 mL de óleo de linhaça (LIN), Grupo 2, 400 mL de óleo de linhaça + 0,2 mg /Kg de selenito de sódio IM (LINSe) e Grupo 3 controles não tratados (C). O óleo foi fornecido diariamente após 15 dias da aplicação única de selenito de sódio, e o experimento teve duração de dez semanas. O resultado deste estudo foi de que os avaliadores não foram capazes de identificar diferenças de cor, odor e sabor entre as amostras de leite dos grupos tratados e do controle, em relação a um padrão conhecido. Assim, concluiu-se que a inclusão do óleo de linhaça na dieta de vacas leiteiras resulta em aumento do CLA, ômega 3 e consequentemente da oxidação do leite, necessitando assim o uso de substâncias antioxidantes ou promotores antioxidantes, como o selenito de sódio injetável para retardar a oxidação. Entretanto, a inclusão de 400 mL diários de óleo de linhaça na dieta de vacas leiteiras não é capaz de provocar alterações sensoriais no leite.

Backround: Oxidative stress (OS) has been implicated in pathogenesis of human disorders such as depressive disorder, sepsis, cardiovascular disease, acute and chronic pancreatitis, and cancer. Increased OS is result of imbalance between increased reactive oxygen and nitrogen species (RONS) production and / or insufficient activity of antioxidant defence system. Antioxidant system, which is composed of antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidases (GPX), glutathione reductase (GR) and non- enzymatic antioxidant reduced glutathione (GSH) plays an important role in the protection of cells against enhanced OS. The aim of this study was to assess the OS markers and antioxidant enzymes in different pathophysiological states. Materials and methods: Activities of erythrocyte glutathione peroxidase (GPX1), GR and concentration of GSH as well as levels of OS markers were analysed in six different pathophysiologic states. These parameters were measured in 35 women with depressive disorder (DD), 40 patients with metabolic syndrome (MetS), 30 septic patients (S) followed up in the course of sepsis; 15 non-septic critically ill patients (NC), 13 patients with acute pancreatitis (AP), 50 with chronic pancreatitis (CP) and 50 patients with pancreatic cancer (PC), compared to...

Introdução: As neuropatias periférica (NP) e autonômica cardiovascular (NAC) são complicações prevalentes do diabetes mellitus (DM). Há indícios de que seu desenvolvimento se deve não somente ao controle metabólico. Neste sentido, a busca por preditores genéticos faz-se imperativa. Diversos genes relacionados às vias bioquímicas que levam ao dano celular induzido pela hiperglicemia têm sido investigados, destacando-se os genes relacionados às vias do extresse oxidativo. O balanço entre os sistemas antioxidantes (como glutationa e tiorredoxina) e pró-oxidantes (como o NADPH-oxidase) é um importante fator na defesa celular contra o estresse oxidativo. Objetivo primário: avaliar a associação entre os polimorfismos de um único nucleotídeo (SNP) pertencentes às vias anti- e pró-oxidantes mencionadas e a NP e NAC em pacientes DM tipo 1: 718C/T na região 3\' UTR (untranslated region) (rs713041) no gene da glutationa peroxidase 4 (GPX4); -129 C/T (rs1788390) no gene da glutamato cisteína ligase (GLCL); -1365 C/T (rs7211) no gene da proteína de interação com a tiorredoxina (TXNIP); -2810 A/G (rs6610650) no gene do CYBB; - 675 T/A (não registrado) no gene do CYBA. Objetivo secundário: avaliar a relação entre as diferentes complicações microvasculares entre si (neuropatia, doença renal diabética [DRD] e retinopatia diabética [RD]). Material e métodos: foram selecionados 378 pacientes com DM tipo 1 do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo e do Hospital das Clínicas da Universidade Estadual de Campinas com mais de 10 anos de doença e controle inadequado (HbA1c >8% em algum período da vida), e examinados para NP e NAC. A genotipagem dos polimorfismos foi realizada pela técnica de reação em cadeia da polimerase em tempo real (Sistema Taqman ®). Foram avaliadas variáveis clínicas, laboratoriais do metabolismo glicêmico e lipídico e a presença de DRD e de RD. Foram avaliados 257 pacientes retrospectivamente quanto à evolução da taxa de filtração glomerular estimada (TFGe) em relação à NP e NAC. O teste de Pearson foi usado para comparar as frequências dos genótipos e a magnitude de associação foi estimada pelo cálculo do odds ratios (OR), com respectivo intervalo de confiança (IC) ajustada por regressão logística para possíveis fatores de confusão. Resultados: A presença de pelo menos um alelo T do SNP +718C/T no gene GPX4 conferiu proteção para NAC (OR=0,39; IC 95% 0,17 - 0,84; P = 0,0165). Na análise de associação entre as complicações, observou-se que: (1) na presença de NP há aumento na probabilidade de NAC (OR = 3,38; IC95% 2,01 - 5,73; P < 0,0001), de albuminúria A3 (OR = 7,17; IC95% 3,68 - 14,53; P < 0,0001) e de RD proliferativa (OR = 9,58; IC95% 5,04 - 19,09; P < 0,0001) e (2) na presença de NAC há aumento na probabilidade de NP (OR = 3,72; IC95% 2,14 - 6,53; P < 0,0001), de albuminuria A3 (OR = 9,37; IC95% 4,68 - 19,65; P < 0,0001) e de RD proliferativa (OR = 3,30; IC95% 1,81 - 6,18, P < 0,0001). No subgrupo avaliado retrospectivamente, a presença de NP e de NAC associou-se com queda de TFGe anual maior (-4,74 mL/min/ano vs. -1,22 mL/min/ano; P < 0,0001 e -3,74 mL/min/ano vs. -1,54 mL/min/ano; P = 0,04, respectivamente). Conclusões: (1) a presença do alelo T no SNP +718C/T (rs713041) no gene GPX4 confere proteção para NAC na população com DM tipo 1 estudada e (2) existe associação de risco para NP e NAC entre si e para as formas graves de DRD e RD / Introduction: Peripheral (PN) and cardiovascular autonomic neuropathies (CAN) are prevalent complications of diabetes mellitus (DM). There are indications that their development occurs not only secondary to metabolic control. Thus, search for genetic predictors is very important. Several genes related to the biochemical pathways that lead to cellular damage induced by hyperglycemia have been investigated, emphasizing the genes related to the pathways of oxidative stress. The balance between antioxidant systems (such as glutathione and thioredoxin) and pro-oxidants (such as NADPH oxidase) is an important factor in cell defense against oxidative stress. Primary objective: to evaluate the association between the single nucleotide polymorphisms (SNP) of the mentioned anti-and pro-oxidant pathways and NP and NAC in type 1 DM patients: 718C / T in the 3 \'UTR (untranslated region) (rs713041) of the glutathione peroxidase 4 gene (GPX4); -129 C / T (rs1788390) in the glutamate cysteine ligase gene (GLCL); -1365 C / T (rs7211) in the thioredoxin interaction protein gene (TXNIP); -2810 A / G (rs6610650) in the CYBB gene; - 675 T / A (unregistered) in the CYBA gene. Secondary objective: to evaluate the relationship between different microvascular complications (neuropathy, diabetic renal disease [DRD] and diabetic retinopathy [DR]). Material and methods: 378 type 1 patients DM were selected from the Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo and from Hospital das Clínicas da Universidade Estadual de Campinas; they had more than 10 years of disease and inadequate control (HbA1c > 8% at some period of life), and were examined for NP and NAC. Polymorphism genotyping was performed by real-time polymerase chain reaction (Taqman System®). Clinical, glycemic and lipid metabolism laboratorial variables and the presence of DRD and DR were evaluated. Also 257 patients were retrospectively evaluated regarding the evolution of the estimated glomerular filtration rate (eGFR) in relation to PN and CAN. The Pearson test was used to compare the frequencies of the genotypes and the magnitude of association was estimated by the odds ratios (OR), with the respective confidence interval (CI) adjusted by logistic regression for possible confounding factors. Results: The presence of at least one T allele at the SNP + 718C / T of the GPX4 gene protected for CAN (OR = 0.39; 95% CI 0.17-0.84; P = 0.0165). In the analysis of the association between complications, it was observed that: (1) in the presence of NP, there was an increase in risk of NAC (OR = 3.38; 95% CI 2.01 - 5.73; P < 0.0001), of albuminuria A3 (OR = 7.17; 95% CI 3.68 - 14.53; P < 0.0001) and of proliferative DR (OR = 9.58; 95% CI, 5.04 - 19.09; P < 0.0001) (2) in the presence of CAN, there was an increase in risk of PN (OR = 3.72; 95% CI 2.14 - 6.53; P < 0.0001), albuminuria A3 (OR = 9.37; CI95% 4.68 - 19.65; P<0,0001) and proliferative DR (OR = 3.30; CI95% 1.81 - 6.18; P < 0,0001).) In the subgroup evaluated retrospectively, the presence of PN and CAN was associated with a greater annual decrease of eGFR (-4.74 mL / min / year vs. -1.22 mL / min / year, P < 0.0001 and - 3.74 mL / min / yr vs. -1.54 mL / min / yr, P < 0.04, respectively). Conclusions: (1) the presence of the T allele at the SNP + 718C / T (rs713041) in the GPX4 gene confers protection for the presence of CAN in the studied type 1 DM population and (2) there is a risk association for PN and CAN among themselves and for severe forms of DRD and RD

A nefropatia diabética (ND) é uma das principais causas de nefropatia crônica, o que torna o diabetes mellitus (DM) responsável por 44% da prevalência de doença renal crônica (DRC) no mundo. O papel do estresse oxidativo na patogênese da ND está bem estabelecido e genes pertencentes a vias pró- e antioxidantes são possíveis candidatos a conferirem susceptibilidade genética a essa e a outras complicações crônicas. Além do estresse oxidativo, o transporte intracelular de glicose, mediado por transportadores específicos, também parece exercer influência sobre a ND e outras complicações. O objetivo deste trabalho foi avaliar a associação entre ND e alguns polimorfismos de um único nucleotídeo (SNPs) em genes que codificam proteínas transportadoras de glicose (GLUT2 [SLC2A2]), proteínas pró-oxidantes (p22phox [CYBA] e NOX-2 [CYBB]) e proteínas antioxidantes (glutationa peroxidase-4 [GPX4] e catalase [CAT]) em uma coorte brasileira (n=453; 45,8% de pacientes com ND) e três coortes francesas (SURGENE [n=340; 17,7% de pacientes com ND na fase basal], GENEDIAB [n=313; 66,7% de pacientes com ND] e GENESIS [n=636; 49,7% de pacientes com ND]) de pacientes portadores de DM tipo 1. Os SNPs foram genotipados com o uso da técnica de reação em cadeia da polimerase (PCR) em tempo real e os resultados expressos em odds ratio (OR) ou hazard ratio (HR), com seus respectivos intervalos de confiança (IC), determinados em modelos ajustados de regressão logística politômica ou regressão de risco proporcional de Cox, respectivamente. A razão albumina/creatinina urinária (ACR) ou a taxa de excreção urinária de albumina (EUA) foram utilizadas para definir os estágios de ND e os pacientes foram classificados de acordo com a presença ou ausência de ND incipiente (ACR 30 - 300 mg/g de creatinina ou EUA 20 - 200 ?g/min ou 20 - 200 mg/L) e creatinina plasmática <1,7 mg/dL), ND estabilizada (ACR >300 mg/g de creatinina ou EUA > 200 ug/min ou > 200 mg/L e creatinina plasmática < 1,7 mg/dL ) ou ND avançada (ACR > 300 mg/g de creatinina ou EUA > 200 ug/min ou > 200 mg/L e creatinina plasmática > 1,7 mg/dL ou qualquer terapia de reposição renal) e também foram avaliadas associações dos SNPs com a taxa de filtração glomerular estimada (TFGe). O alelo raro A do SNP rs6610650 no gene CYBB foi associado com valores baixos de TFGe em mulheres na coorte brasileira e com a prevalência de ND estabilizada/avançada em mulheres da coorte francesa (OR 1,75; IC 95% 1,11 - 2,78; p=0,016). O alelo raro T do SNP rs713041 no gene GPX4 foi inversamente associado com a prevalência de ND estabilizada/avançada em homens na coorte brasileira (OR 0,30, IC95% 0,13 - 0,68, p=0,004) e com valores elevados de TFGe em homens na coorte francesa. O alelo raro A do SNP rs7947841 no gene CAT foi associado com a prevalência de ND incipiente (OR 2,79; IC95% 1,21 - 6,24; p=0,01) e ND estabilizada/avançada (OR 5,72; IC95% 1,62 - 22,03; p=0,007), bem como com a incidência de eventos renais, definidos como novos casos de microalbuminúria ou progressão para um estágio mais grave de ND durante o seguimento de estudo, na coorte SURGENE (HR 1,82; IC95% 1,13 - 2,81; p=0,01). O mesmo alelo de risco associou-se com a prevalência de ND incipiente (OR 3,13; IC95% 1,42 - 7,24; p=0,004) e com a incidência de insuficiência renal crônica terminal (IRCT) na coorte GENEDIAB (HR 2,11; IC95% 1,23 - 3,60; p=0,008) e com a prevalência de ND incipiente (OR 2,16; IC95% 1,14 - 4,10, p=0,02) e ND estabilizada/avançada (OR 2,71; IC95% 1,38 - 5,42; p=0,004) na coorte brasileira. O alelo raro T do SNP rs9932581 no gene CYBA foi inversamente associado com a prevalência de ND estabilizada/avançada (OR 0,60; IC95% 0,46 - 0,78; p=0,0001) e com valores mais baixos de TFGe nos pacientes de descendência europeia da coorte GENESIS/GENEDIAB. Este mesmo alelo foi associado com a incidência de eventos renais e de IRCT nas coortes SURGENE (HR 0,63; IC95% 0,46 - 0,86; p=0,003) e GENESIS/GENEDIAB (HR 0,51; IC95% 0,31 - 0,78; p=0,002), respectivamente. Entretanto estes resultados não foram replicados na coorte brasileira. O alelo raro T do SNP rs11924032 no gene SLC2A2 foi inversamente associado com a perda da TFGe ao logo do tempo (0,02%/ano vs 2,18%/ano para os pacientes portadores do genótipo GG; p=0,005), na coorte SURGENE. Este mesmo alelo foi inversamente associado com a incidência de IRCT nas coortes GENESIS/GENEDIAB (HR 0,53; IC95% 0,29 - 0,89; p=0,01). Os resultados observados para o gene SLC2A2 não forneceram fortes indícios para afirmarmos que este gene exerça um papel relevante no desenvolvimento da ND nos pacientes com DM tipo 1 nas coortes francesas estudadas. Em contrapartida, os SNPs nos genes que codificam as proteínas pró-oxidantes CYBA e CYBB e as proteínas antioxidantes GPX-4 e CAT foram capazes de modular o risco para doença renal em pacientes portadores de DM tipo 1, sendo que os SNPs presentes nos genes CYBB, GPX4 e CAT tiveram seus resultados replicados em coortes independentes, o que corrobora a importância destes genes e, consequentemente, do estresse oxidativo, na patogênese da ND / Diabetic nephropathy (DN) is a major cause of chronic nephropathy, with diabetes mellitus (DM) accounting for 44% of the prevalence of chronic kidney disease (CKD) in the world. The role of oxidative stress in the pathogenesis of DN is well established and genes belonging to pro- and antioxidant pathways are possible candidates to confer genetic susceptibility to this and other chronic complications. Besides oxidative stress, intracellular glucose transport mediated by specific transporters, also appears to influence DN and other complications. The aim of this study was to evaluate the association between DN and some single nucleotide polymorphisms (SNPs) present in genes encoding glucose transport proteins (GLUT2 [SLC2A2]), pro- (p22phox [CYBA] and NOX-2 [CYBB]) and antioxidants (glutathione peroxidase-4 [GPX4] and catalase [CAT]) proteins, in a Brazilian cohort [n= 453; 45.8% f patients with DN], and three French cohorts (SURGENE [n=340; 17.7% of patients with DN at baseline], GENEDIAB [n=313; 66.7% of patients with DN], and GENESIS [n=636; 49.7% of patients with DN]) of patients with type 1 DM. The SNPs were genotyped using the technique of real time polymerase chain reaction (PCR) and results expressed as odds ratio (OR) and hazard ratio (HR), with their respectively 95% confidence intervals (CI), determined by adjusted models of polytomic logistic regression and Cox proportional hazard regression, respectively. The albumin/creatinine ratio (ACR) or the urinary albumin excretion (UAE) rate were used to define the DN stages and the patients were classified according to the presence or absence of incipient DN (ACR 30 - 300 mg/g of creatinine or UAE 20 - 200 ug/min or 20 - 200 mg/L) and plasmatic creatinine < 1,7 mg/dL), established DN (ACR > 300 mg/g of creatinine or EUA > 200 ug/min or > 200 mg/L and plasmatic creatinine <1,7 mg/dL) or advanced DN (ACR >300 mg/g of creatinine or UAE > 200 ug/min or > 200 mg/L and plasmatic creatinine > 1,7 mg/dL or any renal replacement therapy). Associations for the estimated glomerular filtration rate (eGFR) were also evaluated. The rare allele A of the SNP rs6610650 in CYBB gene was associated with low values of eGFR in women in the Brazilian cohort and with the prevalence of established/advanced DN in women in the French cohort (OR 1.75, 95%CI 1.11 - 2.78, p=0.016). The rare allele T of the SNP rs713041 in GPX4 gene was inversely associated with the prevalence of established/advanced DN in men in the Brazilian cohort (OR 0.30, 95%CI 0.13 - 0.68, p=0.004) and with higher values of eGFR in men in the French cohort. The rare allele A of the SNP rs7947841 in CAT gene was associated with the prevalence of incipient DN (OR 2.79, 95%CI 1.21 - 6.24, p=0.01) and established/advanced DN (OR 5.72; 95%CI 1.62 - 22.03, p=0.007) as well as the incidence of renal events, defined as new cases of microalbuminuria or progression to a more severe stage during the follow-up study, in SURGENE cohort (HR 1.82, 95%CI 1.13 - 2.81, p=0.01). The same risk allele was associated with the prevalence of incipient DN (OR 3.13, 95%CI 1.42 - 7.24, p=0.004), the incidence of end-stage renal disease (ESRD) in the cohort GENEDIAB (HR 2.11, 95%CI 1.23 - 3.60, p=0.008) and with the prevalence of incipient DN (OR 2.16, 95%CI 1.14 - 4.10, p=0.02) and established/advanced DN (OR 2.71, 95%CI 1.38 - 5.42, p=0.004) in the Brazilian cohort. The rare T allele of the SNP rs9932581 in CYBA gene was inversely associated with the prevalence of established/advanced DN (OR: 0.60, 95%CI: 0.46 - .78, p=0.0001) and associated with lower values of eGFR in patients of GENESIS/GENEDIAB cohort. The same allele was inversely associated with the incidence of renal events and ESRD in SURGENE (HR 0.63, 95%CI 0.46 - 0.86, p=0.003) and GENESIS/GENEDIAB (HR 0.51, 95%CI 0.31 - 0.78, p=0.002) cohorts. However, these results were not replicated in the Brazilian cohort. The rare T allele of the SNP rs11924032 in SLC2A2 gene was inversely associated with the loss of eGFR during the follow-up (0.02%/year vs. 2.18%/year for patients with the GG genotype, p=0.005) in the SURGENE cohort. The same allele was inversely associated with the incidence of ESRD in the GENESIS/GENEDIAB cohorts (HR 0.53, 95%CI 0.29 - 0.89, p=0.01). The results observed for the SLC2A2 gene, in this study, did not provide strong evidence to state that this gene exerts a relevant role in the development of DN in patients with type 1 DM in the studied cohorts. However, SNPs in genes encoding the pro-oxidant proteins CYBA and CYBB, and the antioxidants proteins GPX-4 and CAT were able to modulate the risk of renal disease in patients with type 1 DM. The studied SNPs in CYBB, GPX4 and CAT genes had their results replicated in independent cohorts, which confirms the importance of these genes and, hence, of the oxidative stress in the pathogenesis of DN

INTRODUÇÃO: acredita-se que o controle glicêmico inadequado, a duração do diabetes melito (DM) e a presença de hipertensão arterial e dislipidemia sejam os fatores de risco mais importantes para o desenvolvimento das complicações microvasculares no DM, contudo, existem inúmeras evidências sugerindo que uma predisposição genética participe da suscetibilidade para o desenvolvimento dessas complicações. Vários genes relacionados aos mecanismos dos danos induzidos pela hiperglicemia têm sido investigados. O papel do estresse oxidativo na patogênese das complicações crônicas do DM vem sendo demonstrado e os genes que codificam enzimas que participam dos mecanismos antioxidantes são candidatos a conferirem suscetibilidade ou proteção contra as complicações crônicas. A glutationa é um dos mais importantes antioxidantes endógenos; no entanto, a associação entre polimorfismos em genes que codificam enzimas que participam desse sistema e complicações crônicas do DM foi pouco explorada na literatura. OBJETIVOS: avaliar a associação de polimorfismos em três genes que codificam enzimas relacionadas ao metabolismo da glutationa com o desenvolvimento de nefropatia e retinopatia em pacientes diabéticos tipo 1. Foram estudados: o polimorfismo -129C/T do gene GCLC, o número de repetições do trinucleotídeo GCG no exon 1 do gene GPX1 e o polimorfismo -65T/C do gene GPX3. CASUÍSTICA E MÉTODOS: 299 pacientes (139 do gênero masculino e 160 do gênero feminino) com DM tipo 1 com mais de 15 anos de diagnóstico e mau controle glicêmico foram divididos conforme presença ou ausência das seguintes complicações: nefropatia diabética (ND) avançada, ND, doença renal crônica (DRC) estágios 3 a 5 e retinopatia diabética proliferativa (RDP). Em cada grupo foram avaliadas as freqüências das variantes alélicas dos três genes estudados. RESULTADOS: a distribuição dos genótipos na população estudada foi consistente com o equilíbrio de Hardy-Weinberg para os três genes analisados. A presença de pelo menos um alelo T do polimorfismo -129C/T do gene GCLC conferiu risco independente para a presença de ND avançada (OR = 2,82 ; IC 95% = 1,13 - 7,05; p = 0,026), para ND (OR = 3,64; IC 95% = 1,27 10,36; p = 0,016) e para DRC estágios 3 a 5 (OR = 5,74; IC 95% = 2,17 15,1; p < 0,001) e a presença de pelo menos um alelo C do polimorfismo -65 T/C conferiu risco independente para a presença de ND avançada (OR = 2,62; IC 95% = 1,19 -5,72, p = 0,022) na população estudada. Não houve associação do número de repetições do trinucleotídeo GCG do gene GPX1 com nenhuma das complicações estudadas. O haplótipo CC_TT, composto pelos alelos selvagens dos genes GCLC e GPX3, foi negativamente associado com ND avançada (OR = 0,32, IC 95% = 0,15 0,66; p = 0,002) e DRC (OR = 0,25; IC 95% = 0,11 - 0,55; p = 0,001). CONCLUSÕES: a presença de pelo menos um alelo T do polimorfismo -129 C/T do gene GCLC e de pelo menos um alelo C do polimorfismo -65 T/C do gene GPX3, ambos associados a uma menor atividade transcricional do respectivo gene, conferiram risco para a presença de complicações renais na população de pacientes estudada. / INTRODUCTION: glycemic control, diabetes duration, systemic hypertension and dyslipidemia have been implicated as main risk factors for the development of diabetic microangiopathy, however there is evidence suggesting that genetic predisposition plays a role in the susceptibility to microvascular complications. Based on underlying pathogenesis, polymorphisms of several genes belonging to multiple pathways have been investigated, like the genes related to mechanisms of hyperglycemia-induced damage. The role of oxidative stress in the pathogenesis of diabetic complication has been increasingly demonstrated and genes coding enzymes involved in antioxidant defense are candidates to confer susceptibility or protection against these complications. Glutathione is one the most important endogen antioxidants, however, the association between polymorphisms in genes related to glutathione metabolism and diabetic complications has not been deeply investigated. OBJECTIVES: to study the association between polymorphisms in three genes which code enzymes related to glutathione metabolism and the development of nephropathy and retinopathy in type 1 diabetic patients: the polymorphism -129 C/T of GCLC, the number of trinucleotide GCG repeats at exon 1 of GPX1 and the polymorphism -65 T/C of GPX3. CASUISTIC AND METHODS: 299 type 1 diabetic patients (139 male and 160 female) with at least 15 years from diagnosis and poor glycemic control were studied. The patients were divided in two groups according to the presence or absence of diabetic complications: with and without diabetic nephropathy (DN), advanced DN, chronic kidney disease (CKD) stages 3 to 5 and proliferative diabetic retinopathy (PDR). RESULTS: The allelic distribution of the three studied polymorphisms was consistent with Hardy-Weinberg equilibrium. The presence of at least one T allele of GCLC 129 C/T was an independent risk factor for advanced DN (OR = 2.82 ; CI 95% = 1.13 -7.05; p = 0.026), for DN (OR = 3.64; CI 95% = 1.27 10.36; p = 0.016) and for CKD stages 3 to 5 (OR = 5.74; CI 95% = 2.17 15.1; p < 0.001) and the presence of at least one C allele of GPX3 -65 T/C was an independent risk factor for advanced DN (OR = 2.62; IC 95% = 1.19 -5.72, p = 0.022) in the studied population. There were no associations between GCG trinucleotide repeats of GPX1 and diabetic complications. The haplotype CC_TT, composed by GCLC and GPX3 wild type alleles, was negatively related to advanced DN (OR = 0.32, CI 95% = 0.15 0.66; p = 0.002) and CKD (OR = 0.25; CI 95% = 0.11 0.55; p = 0.001). CONCLUSIONS: the presence of at least one T allele of -129C/T polymorphism of GCLC and one C allele of -65 T/C polymorphism of GPX3, both associated to a lower transcriptional activity of its genes, conferred risk for renal complications in the studied population.