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No. of bitstreams: 1 ESTRELA_ANDRE_LUIZ_TESE.pdf: 2000480 bytes, checksum: 0cf023323652932ee9da95b8b9900817 (MD5) Previous issue date: 2017-02-21 / INTRODUCTION: To verify if there are differences in the hormonal, immunological, redox, chronotropic and physical performance thresholds between two levels of active elderly training induced at two maximum and consecutive sessions in the short term. METHODS: Forty marathon runners were divided based on training load: a higher volume group (n = 19, training ~ 480 min / week) and a smaller group volume (n = 21, training ~ 240 min / week). Inflammatory proteins (PRC, TNF-?, IL-6) and plasma oxidative stress biomarkers (protein carbonyls, total thiols), plasma antioxidant enzyme activity (superoxide dismutase, SOD, glutathione peroxidase, GPx) and testosterone and salivary cortisol, Were evaluated at rest (baseline) and in response to the cumulative effect of the two maximal exercise tests, separated by four hours of rest. RESULTS: Most of the parameters were similar between the groups at the beginning of the study, except the lower volume group exhibited higher CRP (+ 60%, p <0.05), higher protein carbonyls (+ 23%, p = 0, 07) and lower SOD activity (-4%, p = 0.06) compared to the higher volume group. TNF-? decreased in response to the cumulative effect of similar acute exercise in both groups (ie, baseline vs. one sample collected after the second exercise session), whereas CRP increased differently (+ 60% % Higher group volume, interaction effect p <0.05). IL-6 did not change in any of the groups. Protein carbonyls decreased and total thiols increased similarly in both groups in response to exercise (p<0.05), but SOD activity differentially declined (-14% lower volume, -20% of the higher volume group, Interaction effect p <0.05). GPx did not change in either group. Salivary testosterone decreased in response to exercise was similar in both groups (p <0.05), whereas cortisol did not change. CONCLUSION: Individuals involved in higher volume training have a lower inflammatory response induced by exercise, and a greater decline in the activity of some antioxidant enzymes in post-exercise plasma, compared with individuals engaging in low volume exercise. / INTRODU??O: Conhecemos muito pouco sobre os efeitos fisiol?gicos do treinamento pesado em atletas idosos. Nesse estudo, verificamos se h? diferen?as nos limiares hormonais, imunol?gicos, redox, cronotr?picos e de desempenho f?sico entre dois n?veis de treinamento de idosos ativos induzidos a duas sess?es m?ximas e consecutivas em curto prazo. M?TODOS: Quarenta maratonistas foram divididos com base na carga de treinamento: um grupo de maior volume (n = 19, treinamento ~ 480 min / semana) e um grupo menor volume (n = 21, treinamento ~ 240 min / semana). Prote?nas inflamat?rias [prote?na C reativa (PCR), fator de necrose tumoral (TNF)-?, interleucina (IL)-6] e biomarcadores de stress oxidativo plasmaticos (carbonilos proteicos, ti?is totais), a atividade da enzima antioxidante plasm?tica (super?xido dismutase, SOD; glutationa peroxidase, GPx) e testosterona e cortisol salivar, foram avaliados em repouso (linha de base) e em resposta ao efeito acumulado dos dois testes m?ximos de exerc?cio, separados por quatro horas de descanso. RESULTADOS: A maioria dos par?metros eram semelhantes entre os grupos no in?cio do estudo, exceto o grupo de menor volume exibiu maior PCR (+ 60%, p <0,05), carbonilos de prote?nas maiores (+ 23%, p = 0,07) e menor atividade SOD (-4%, p = 0,06) em compara??o com o grupo de maior volume. O TNF-? diminuiu em resposta ao efeito acumulado de exerc?cio agudo de maneira semelhante nos dois grupos (ou seja, linha de base vs. uma amostra coletada ap?s a segunda sess?o de exerc?cio), enquanto PCR aumentou diferencialmente (+ 60% menor volume; + 24% maior volume grupo, efeito de interac??o p <0,05). A citocina IL-6 n?o se alterou em qualquer dos grupos. Os carbonilos de prote?nas diminuiram e os ti?is totais aumentaram de maneira similar em ambos os grupos em resposta ao exerc?cio (p<0,05), mas a atividade da SOD declinou diferencialmente (-14% menor volume; -20% do grupo de maior volume, efeito de intera??o p <0,05). A GPx n?o se alterou em nenhum dos grupos. A testosterona salivar diminu?da em resposta ao exerc?cio foi semelhante em ambos os grupos (p <0,05), ao passo que o cortisol n?o se alterou. CONCLUS?O: Indiv?duos idosos envolvidos em treinamento de maior volume apresentam uma resposta inflamat?ria menor induzida pelo exerc?cio, e um decl?nio maior na atividade de algumas enzimas antioxidantes no plasma p?s-exerc?cio, em compara??o com os idosos envolvidos em exerc?cios de baixo volume.

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abstract: The search for highly active, inexpensive, and earth abundant replacements for existing transition metal catalysts is ongoing. Our group has utilized several redox non-innocent ligands that feature flexible arms with donor substituents. These ligands allow for coordinative flexibility about the metal centre, while the redox non-innocent core helps to overcome the one electron chemistry that is prevalent in first row transition metals. This dissertation focuses on the use of Ph2PPrDI, which can adopt a κ4-configuration when bound to a metal. One reaction that is industrially useful is hydrosilylation, which allows for the preparation of silicones that are useful in the lubrication, adhesive, and cosmetics industries. Typically, this reaction relies on highly active, platinum-based catalysts. However, the high cost of this metal has inspired the search for base metal replacements. In Chapter One, an overview of existing alkene and carbonyl hydrosilylation catalysts is presented. Chapter Two focuses on exploring the reactivity of (Ph2PPrDI)Ni towards carbonyl hydrosilylation, as well as the development of the 2nd generation catalysts, (iPr2PPrDI)Ni and (tBu2PPrDI)Ni. Chapter Three presents a new C-O bond hydrosilylation reaction for the formation of silyl esters. It was found the (Ph2PPrDI)Ni is the most active catalyst in the literature for this transformation, with turnover frequencies of up to 900 h-1. Chapter Four explores the activity and selectivity of (Ph2PPrDI)Ni for alkene hydrosilylation, including the first large scope of gem-olefins for a nickel-based catalyst. Chapter Five explores the chemistry of (Ph2PPrDI)CoH, first through electronic structure determinations and crystallography, followed by an investigation of its reactivity towards alkyne hydroboration and nitrile dihydroboration. (Ph2PPrDI)CoH is the first reported cobalt nitrile dihydroboration catalyst. / Dissertation/Thesis / Doctoral Dissertation Chemistry 2018

El primer objetivo de este trabajo consiste en poner de relevancia como la sulfonación de la polianilina (PANI) tiene implicaciones profundas que afectan al material a todos los niveles, desde niveles microscópicos: mecanismos que rigen la oxidación/reducción, el dopado/desdopado del polímero; hasta un nivel macroscópico evaluando propiedades tales como la solubilidad y la conductividad eléctrica. Para ello se realizan estudios electroquímicos con el pH y la fuera iónica, sobre PANI sulfonada (SPAN) electro-generada, y seguidamente se emplean técnicas espectro-electroquímicas (UV-Vis-NIR, FTIR y EPR in situ). Posteriormente se estudia sobre la SPAN generada químicamente la influencia del grado de sulfonación en las propiedades macroscópicas más relevantes. En los siguientes capítulos se abordan investigaciones relacionadas con potenciales aplicaciones de la PANI auto-dopada. Los temas son dispares: i) la detección de indicadores fisiológicos en biomedicina y ii) la preparación de catalizadores para aplicaciones en reactores con fines energéticos. En ambos casos la aproximación es similar, el polímero presenta propiedades óptimas de conductividad y estabilidad para constituir una matriz donde inmovilizar un componente/fase catalíticamente activa y ejercer además como eficiente colector de corriente. En el presente trabajo de investigación se ahonda en dicha posibilidad desde una premisa clara, hallar unas condiciones de preparación que respeten al máximo la integridad del polímero.

Magnetite (Fe3O4) is a commonly found in the environment and can form via several pathways, including biotic and abiotic reduction of Fe3+ oxides and the oxidation of Fe2+ and Fe0. Despite extensive research, the redox behavior of magnetite is poorly understood. In previous work, the extent and kinetics of contaminant reduction by magnetite varied by several orders of magnitude between studies, two fundamentally different models are used to explain magnetite oxidation (i.e., core-shell diffusion and redox-driven), and reported reduction potentials vary by almost 1 V. In other fields of science (e.g., physics), magnetite stoichiometry (x = Fe2+/Fe3+) is a commonly measured property, however, in environmental studies, the stoichiometry is rarely measured. The stoichiometry of magnetite can range from 0.5 (stoichiometric) to 0 (completely oxidized), with intermediate values (0 < x < 0.5) referred to as nonstoichiometric or partially oxidized magnetite. To determine the relationship between magnetite stoichiometry and contaminant fate, the reduction rates of three substituted nitrobenzenes (ArNO2) were measured. The kinetic rates varied over five orders of magnitude as the particle stoichiometry increased from x = 0.31 to 0.50. Apparent 15N kinetic isotope effects (15N-AKIE) values for ArNO2 were greater than unity for all magnetite stoichiometries investigated, and indicated that mass transfer processes are not controlling the reaction rate. To determine if the reaction kinetics were redox-driven, magnetite open circuit potentials (EOCP) were measured. EOCP values were linearly related to the stoichiometry, with more stoichiometric magnetite having a lower potential, in good agreement with redox-driven models. The reaction of aqueous Fe2+ and magnetite was investigated. Similar to previous findings for other Fe3+ oxides, the formation of a stable sorbed Fe2+ species was not observed; instead, the sorbed Fe2+ underwent interfacial electron transfer to form a partially oxidized magnetite phase, which was accompanied by reduction of the underlying magnetite. The lack of a stable sorbed Fe2+ species on magnetite indicated that the traditional surface complexation model was incorrect; instead, the uptake of Fe2+ by magnetite appeared to be limited by the whole particle (i.e., the sorbed and underlying phases combined) reaching a stoichiometry of 0.5.

Eukaryotic gene expression is a complex process that can be controlled at the level of transcription, post-transcription, translation, or post-translation. In recent years there has been growing interest in understanding the role of the 3'-untranslated region (UTR) in post-transcriptional regulation. The 3'-UTR contains many regulatory sequences, including microRNA (miR) target sites and AU-rich elements (AREs). Although a relatively recent discovery, miRs have been shown to downregulate target gene expression and have important roles in regulating many cellular processes, including cellular growth. Cellular growth consists of two distinct states, proliferation and quiescence. The proliferative state consists of G1, S, G2, and M phases while quiescence is the G0 phase. In response to mitogenic stimuli, quiescent cells enter the proliferative cycle and may transit back to the quiescent state. Reentry into quiescence is essential to prevent aberrant proliferation as well as to protect the cellular life span. Cells that remain in quiescence for an extended period of time lose their ability to proliferate. It has been shown that the redox status of the cells may regulate quiescence and proliferative capacity since overexpression of SOD2 protects the proliferative capacity of quiescent cells. We hypothesized that the redox environment regulates proliferative capacity through miR expression and regulation of miR targets. Early results showed treatment with hydroxytyrosol (HT), an olive-derived catechol, was able to protect the proliferative capacity of quiescent normal human fibroblasts. HT was shown to use hydrogen peroxide and produce superoxide in a catechol-semiquinone-quinone redox cycle. Interestingly, HT also induced SOD2 expression. Further results from microRNA PCR arrays and Taqman PCR assays showed a significant decrease (4-fold) in miR-302 levels during quiescence compared to proliferating normal human fibroblasts, suggesting that miR-302 could regulate cellular growth states. Results from a Q-RT-PCR and dual luciferase-3'-UTR reporter assays identified ARID4a (AT-Rich Interacting Domain 4a, also known as RBP1) and CCL5 (C-C motif Ligand 1) as targets for miR-302. Ionizing radiation, that is well known to induce oxidative stress and delay cell cycle progression, decreased miR-302 levels, which was associated with an increase in its target mRNA levels, ARID4a and CCL5. Such an inverse correlation was also observed in cells treated with hydrogen peroxide, SOD2 overexpressing cells, and HT treated cells. Overexpression of miR-302 suppresses ARID4a and CCL5 mRNA levels, and increased the percentage of S-phase cells. These results identified miR-302 as a redox-sensitive regulator of ARID4a and CCL5 mRNAs as well as demonstrate a regulatory role of miR-302 during quiescence and proliferation.

New medical treatments, compounds that affect human health, nutritional supplements, and other substances, are introduced to society every day. The accurate determination of the potential toxicity from these substances is of critical importance to our society. Goals of the modern toxicologist not only involve the determination of the toxic potential of new substances but also: the elucidation of mechanisms; improving existing assays; and developing new assays to study toxicity. This thesis addresses these goals in two topics fundamental to toxicology. Re-evaluating the expression of dose and susceptibility of cells in culture The exposure of cells in culture to drugs, xenobiotics, and other compounds is one of the first tools used to determine the potential for toxicity. Problems can arise when results of these experiments are translated to next-level toxicity experiments (e.g. animals and humans). I hypothesized that "dose" in cell culture can be improved by designing and reporting experiments based on dose in moles per cell. When experiments were compared on an extracellular concentration basis, a large apparent variability in toxicity was observed. However, if these same exposures were expressed as moles per cell, all experiments yielded the same toxicity. In addition to the evaluation of mole per cell, I investigated the susceptibility of various cells to 1,4-benzoquinone. I hypothesized that upon exposure to toxins that bind covalently, larger cells would require more molecules per cell of toxin versus a smaller cell to achieve identical toxicities. I found a linear correlation between cell volume(pL) and ED50 (mole per cell where 50 % cell viability is lost), supporting my hypothesis. This work could improve current cell culture protocols and allow for better and less expensive determination of toxicities. Heritability of the red blood cell storage lesion Blood transfusions are an integral part of modern medicine with 5 million people receiving blood each year in the United States. There is growing evidence that red blood cells (RBCs) stored for longer periods are less therapeutically beneficial and could even be harmful to patients. This phenomenon of diminished RBC function with increased time in storage is called the storage lesion. However, there is great variation between different donors in the severity of the storage lesion in their donated RBCs. I hypothesized that part of this variability in the RBC storage lesion is determined by heritable genetic differences. To test this hypothesis, a study using mono- and di-zygotic twins was performed to determine the heritability of adenosine triphosphate (ATP), glutathione (GSH), glutathione disulfide (GSSG) and hemolysis in stored blood. Major discoveries in this study include: GSH, GSSG, and the half-cell reduction potential (Ehc) are heritable (57 %, 51 %, and 70 %, respectively) in non-stored RBCs. In addition, ATP was found to be heritable in two different storage solutions (62 % in AS-3, 71 % in CP2D); as well as GSH, GSSG, Ehc and hemolysis (59 %, 48 %, 64 %, and 53 %, respectively). These discoveries could eventually be used to develop new genetic tests that would predict the rate of deterioration in stored blood quality on an individual basis.

The reaction between aqueous Fe(II) and Fe(III) oxides is extremely complex, and can catalyze Fe(II)-Fe(III) electron transfer, exchange of Fe atoms between the aqueous and solid phases, mineral transformation, and contaminant reduction. Together, these processes represent a phenomenon referred to as Fe(II)-catalyzed Fe oxide recrystallization, which has been observed under controlled conditions in the laboratory for numerous Fe oxides. In the environment, Fe oxides are likely surrounded by organic carbon in various forms, but their potential to interfere with Fe(II)-catalyzed Fe oxide recrystallization, and its subsequent environmental relevance has not been well studied. The Fe(II)-catalyzed recrystallization of stable Fe oxides goethite and magnetite was studied in the presence of several environmentally relevant classes of organic carbon. For both goethite and magnetite, Fe(II)-catalyzed recrystallization continued relatively undeterred in the presence of electron shuttling compounds, natural organic matter isolates, and extracellular polysaccharides. Slight inhibition was observed when spent media from dissimilatory iron-reducing cultures was present, but only by sorbing a long-chain phospholipid to the oxides was significant inhibition observed. The lack of interference by organic carbon indicates that Fe(II)-catalyzed Fe oxide recrystallization is likely to be relevant throughout a wide range of environments, and represents a significant process with regards to the geochemical cycling of Fe atoms, a claim supported by evidence of Fe(II)-driven isotope mixing in real soils. The movement of atoms during Fe(II)-catalyzed Fe oxide recrystallization is not limited to just Fe however. Multiple trace elements have been shown to exchange between the aqueous and solid phases along with Fe during the Fe(II)-catalyzed recrystallization of Fe oxides. The effect of organic carbon, both sorbed to the oxide surface and coprecipitated with the oxide, on Fe(II)-catalyzed atom exchange and transformation of ferrihydrite was studied. Again, the presence of organic carbon did not appear to influence Fe atom exchange kinetics. It also did not appear to influence the rapid transformation of ferrihydrite to lepidocrocite. The presence of organic carbon does appear to ultimately have implications for mineral transformation, as over longer time periods it stabilized lepidocrocite, preventing its subsequent transformation to magnetite or goethite.

Hematopoietic stem cells (HSCs) that function to maintain the hematopoietic compartment through self-renewal and differentiation capacities, as well as their downstream progeny, are susceptible to transformation resulting in the generation of the leukemic stem cell (LSC). Chief in the factors that control HSC regulation and protection of the HSC compartment is the cellular redox environment. Deregulation of the Hematopoietic Stem/Progenitor Cell (HSPC) redox environment results in loss of HSPC function and exhaustion. The characteristic developments of HSPC exhaustion via exposure to redox stress closely mirror phenotypic traits of hematopoietic malignancies, presenting the HSPC/LSC redox environment as a potential therapeutic target. While myelosuppression and HSPC exhaustion are detrimental side effects of classical chemotherapies, new approaches that differentially modify the HSPC/LSC redox environment may demonstrate LSC cytotoxicity while offering protection of normal HSPC function via differential activation of internal signaling pathways. Precisely how the redox environment and downstream signaling events are affected by these treatments remains unclear; thus highlighting the need for robust methods that evaluate the HSPC/LSC redox state. Because the glutathione (GSH), glutathione disulfide (GSSG) redox couple heavily contributes to the management of HSPC function and redox environment, characterizing the GSH/GSSG redox potential at the HSPC level would provide great insight for therapeutic opportunities. However, accurate measurement the GSH/GSSG redox potential within HSPCs/LSCs has been difficult due to their inherently low numbers. Here, we describe the development and validation of a sensitive method used for the direct and simultaneous quantitation of both oxidized and reduced GSH via LC-MS/MS. We use these methodologies to establish a difference in GSH-GSSG half-cell reduction potentials between normal and malignant HSPCs and examine the therapeutic effect of a redox active MnSOD mimetic, Mn(III) mesotetrakis (N-n-butoxyethylpyridinium-2yl) porphyrin, MnTnBuOE-2-PyP5+ (MnP), within these populations in vitro as well as within a human xenograft model in vivo. MnP demonstrates significant cytotoxic effects in several malignant models, while inducing an opposite cytoprotective effect in normal HSPC populations. The GSH/GSSG redox balance, specifically managed by glutathione reductase activity, is identified as a determining factor of MnP efficacy in various malignant populations. Treatment of the human myelodysplastic cell line (MDSL) offers mechanistic insights into MnP efficacy through hydrogen peroxide mediated activation of activator protein 1 (AP-1) signaling. We identify the redox dependent activation of JunB, a known regulator of normal myeloid lineage HSC proliferation, as a transcriptional mechanistic mediator of MnP treatment induced AP-1 signaling resulting in malignant cytotoxicity. The development of this novel method allowing for the identification of targetable differences between normal and malignant cell populations has provided insight to the underpinnings of potential redox based therapies. Additionally, the finding that MnP can target varying cellular redox states and exert selective cytotoxicity in malignant over normal populations by re-gaining lost control of AP-1 signaling demonstrates the potential for development of safe therapeutics within a variety of clinical applications.