Dissertations / Theses on the topic 'Metabolism'

The tricarboxylic acid (TCA) cycle is one of the central pathways in respiration and also plays an important role in a variety of metabolic processes including the synthesis of secondary metabolites and the provision of carbon skeletons for ammonium assimilation and amino acid biosynthesis. Effective regulation of these multiple demands on the TCA cycle is likely to be very important for plant fitness. One way that this regulation could be achieved is through metabolite channelling. This occurs when metabolites are transferred between enzyme active sites without diffusing into the bulk aqueous phase of the cell, and is known to be important in regulating demands in metabolic pathways. Although there is evidence that metabolite channelling exists in animals, there have been no attempts to investigate it in plant. The first aim of this thesis was therefore to investigate whether metabolite channelling exists in the plant TCA cycle. Isotope dilution experiments were developed to investigate metabolite channelling, and were able to show that metabolite channelling was present between certain enzymes of the TCA cycle in both S. tuberosum and A. thaliana mitochondria. The second aim of the thesis was investigate whether metabolite channelling is important in regulating the TCA cycle in plant mitochondria. The pattern of metabolite channelling did not change in mitochondria isolated from the light and the dark, or from mitochondria with increased or decreased TCA cycle rates, but it was not possible to say whether the metabolite channelling altered in a quantitative fashion. Overall the thesis provides the first direct evidence of channelling in the TCA cycle in plants, and further work should help to elucidate what role, if any, it plays.

Tratamos ratos por 7 dias com injeções subcutâneas de óleos de soja (S) ou de girassol (G). Verificamos o desenvolvimento de resistência insulínica, com diminuição de GLUT4 e de sua translocação no tecido adiposo de ratos tratados com estes óleos, apesar de aumento no RNAm de GLUT4 no músculo e no tecido adiposo. O tratamento com girassol diminuiu RNAm de GLUT1 no adiposo, onde também diminuiu a quantidade de RNAm de NF-<font face=\"symbol\">kB. (outros fatores transcricionais não apresentaram alteração: PPAR<font face=\"symbol\">g, MEF2A e MF2D). Dosamos ácidos graxos no plasma, fígado (sem alterações), músculo (palmitoléico aumentou no grupo G; linolênico diminuiu no S e G e araquidônico aumentou no S) e adiposo (palmítico e esteárico aumentaram e linoleico diminuiu nos grupos S e G, além de aumentar a proporção saturados/ insaturados).<br>We treated rats for 7 days with subcutaneous soybean (SB) oil or sunflower (SF)oil injections. Insulin resistance was developed, with a decrease in GLUT4 quantity and translocation in adipose tissue in the rats treated with those oils, despite GLUT4 mRNA increase in muscle and adipose tissue. Sunflower treatment led to decrease in GLUT1mRNA in adipose tissue, where NF- <font face=\"symbol\">kB mRNA was also decreased (other transcriptional factors did not change: PPAR<font face=\"symbol\">g, MEF2A e MF2D). Fatty acids were measured in the plasma, the liver (no changes), muscle (palmitoleic increased in SF, linolenic decreased in SB and SF and arachidonic increased in SB) and adipose tissue (palmitic and stearic increased and linoleico decreased in SB and SF, besides increasing in saturated/ unsaturated ratio.

Zasadnicznym celem przeprowadzonych badań była ocena skuteczności wykorzystania danych z okresowej kontroli użytkowości mlecznej w typowaniu krów z zaburzeniami metabolizmu energetycznego i wpływie opóżnienia u nich terminu pierwszej inseminacji na wskaźniki rozrodcze stada<br>The aim of the research was to assess the effectiveness of the use of data from periodic control of dairy utility in the selection of cows with energy metabolism disturbances and the impact of their delay in the first insemination on the reproductive indicators of the herd

La crema prescrita consisteix en l’ús planificat del foc en condicions ambientals adequades per tal d’aconseguir objectius de gestió prèviament definits. S’executa amb l’objectiu de reduir el risc d’incendis tot i que també s’utilitza per a la gestió de pastures i la conservació de la biodiversitat. La crema sota arbrat pot reduir el risc d’incendi de capçades, però també pot afectar el metabolisme primari i secundari de l’arbre. Tanmateix, les cremes haurien de minimitzar els efectes negatius del foc als arbres i maximitzar els positius. Així doncs, és important comprendre de quina manera els components específics del règim de foc, com són la intensitat del foc, severitat i estació de crema, afecten el funcionament de l’arbre. L’objectiu d’aquesta tesi és comprendre la influència que els factors del règim de cremes prescrites tenen en el metabolisme primari i secundari de tres espècies de pi amb tolerància al foc contrastada: Pinus nigra ssp. salzmannii (Dunal) Franco, P. sylvestris L. i P. halepensis Mill., utilitzant una combinació de tècniques dendrocronològiques, isotòpiques i de quantificació terpènica. El creixement post-crema va dependre del temps transcorregut des de la crema, l’espècie, la resistència de l’arbre, la severitat del foc i del creixement de l’arbre abans de la crema. L’any de la crema, el creixement es va reduir en P. halepensis i no es va veure afectat en P. nigra i P. sylvestris. Amb el pas del temps, el creixement es va incrementar en P. nigra, es va recuperar en P. halepensis i es va reduir en P. sylvestris. La probabilitat de morir va ser menor en P. nigra que en P. sylvestris. L’estació de crema va emergir com un factor important per explicar la mortalitat inicial: la probabilitat que un pi mori és més alta a la primavera que a la tardor per a un cert nivell de capçada socarrimada. La mortalitat tardana va ser més alta a les cremes de tardor que a les de primavera degut als majors temps de combustió registrats a la base dels troncs. Una reducció rellevant de la competència arbrada va augmentar significativament el creixement mitjançant un efecte positiu en la fusta de tardor de P. nigra i P. sylvestris, a mesura que el dany causat pel foc a la tija va disminuir. A més, vam demostrar que una crema executada just després d’un any sec no redueix la resiliència dels pins en comparació amb la dels pins no cremats. En P. halepensis, una reducció significativa de la competència va resultar en majors creixements amb el pas del temps, especialment en pins amb un menor volum de capçada socarrimada. Aquest augment va coincidir amb un any sec i es va associar amb una major conductància estomàtica, el que suggereix que la disponibilitat d’aigua va millorar després de la crema. La crema també va afectar al metabolisme secundari, específicament a la quantitat i al tipus de terpens segons l’espècie de pi i la severitat. A major volum de capçada socarrimada, la concentració de terpens a l’acícula dels pins 24 h post-crema fou major, però 1 any després es va produir una notable disminució. Aquesta reducció va ser més pronunciada en pins beneficiats per l’augment de la disponibilitat de recursos després de la crema, el que suggereix que els productes assimilats pels pins es van assignar al creixement en lloc de a mecanismes de defensa. Aquesta tesi proporciona informació valuosa per ajudar en la millora de la planificació de les cremes als boscos de pi mediterranis, en termes d’intensitat, severitat i estació de crema, oferint una nova finestra d’oportunitat per a l’ús de la crema prescrita com a eina de gestió forestal.<br>Prescribed burning is the planned use of fire to meet clear management objectives under suitable environmental conditions. It is usually executed to reduce fire hazard, but also to manage range and conserve biodiversity. Prescribed burning applied under a forest canopy can reduce crown fire hazard; however, underburning might affect the primary and secondary metabolism of trees. Planning underburning to reduced fire hazard, while minimizing the negative effects and maximizing the positive effects on trees, requires understanding how specific components of the fire regime, such as fire intensity, severity and season, affect tree performance. The goal of this doctoral thesis is to understand the influence of prescribed burning regime factors and related fire impacts on the primary and secondary metabolisms of three pine species with contrasting fire tolerances: Pinus nigra ssp. salzmannii (Dunal) Franco, P. sylvestris L. and P. halepensis Mill., using a combination of dendrochronological, isotope and terpene quantification techniques. Post-burning growth variations depended on the time since burning, the pine species, tree resistance, fire severity and tree performance before burning. In the year of burning, growth was reduced in P. halepensis and unaffected in P. nigra and P. sylvestris. However, as time passed, growth increased in P. nigra, recovered in P. halepensis and decreased in P. sylvestris. P. nigra had a lower probability of dying than P. sylvestris. Burning season emerged as an important factor for explaining initial post-burning pine mortality, since for a certain level of crown injury the probability of a pine dying was higher in spring than in fall. In contrast, delayed pine mortality was higher in fall than in spring burns probably due to the longer combustion times recorded during the fall burns at the base of the trunk. A relevant release of tree competition increased growth through a positive effect on the latewood of P. nigra and P. sylvestris as stem injury decreased. Moreover, we showed that burning just after a dry year did not reduce the growth resilience of pines in comparison with unburned pines. In P. halepensis a relevant competition release, especially in pines with lower crown volume scorched, resulted in higher growth rates as time since burning increased. This growth response coincided with a dry year and was associated with higher stomatal conductance, suggesting that water availability was enhanced after burning. Burning also affected the secondary metabolism of pines, and specifically the amount and type of terpene production depending on the pine species and fire severity. Thus, as crown injury increased, needle terpene concentration 24h post-burning also augmented. However, a remarkable decrease occurred at one year post-burning. This reduction was more pronounced in pines benefited by the increase in resource availability after burning, suggesting that pines were allocating assimilates to growth rather than to defence. From a fuel management point of view, this thesis provides valuable information that can be used to better plan prescribed burning in Mediterranean Pinus forests, in terms of required fire intensity, severity and burning season, offering a new window of opportunity for the use of prescribed burning as a forest management tool.

No presente trabalho analisou-se um total de 101 indivíduos, sendo 26 não infectados e 75 infectados pelo HIV e classificados de acordo com o Walter Reed Army Institute (28 pacientes WR 1, 31 pacientes WR 2 e 16 pacientes WR 3/4). 05 indivíduos infectados pelo HIV apresentaram, nos estágios iniciais, uma diminuição progressiva do peso corporal, dos níveis séricos de uréia, albumina, colesterol total, HOL colesterol e LOL colesterol. Já os níveis séricos de proteínas totais, globulinas, IgG, IgA, &#945;1 glicoproteína ácida, haptoglobina e as atividades enzimáticas da AST e da LD apresentaram elevação nos indivíduos infectados e em conseqüência da evolução da infecção. Os triglicérides demonstraram apenas tendência para aumento dos níveis séricos nos indivíduos estadiados como WR3/4. Os níveis de ferro sérico encontraram-se diminuídos nos indivíduos estadiados como WR 3/4, enquanto que a concentração de transferrina apresentou-se diminuída apenas no Grupo WR 2. Houve uma tendência para a elevação progressiva dos níveis médios de ferritina com a evolução da doença. Nenhuma alteração foi verificada nos níveis de proteína \"C\" reativa. A EC-SOO apresentou diminuição dos níveis de atividade nos indivíduos infectados pelo HIV, enquanto que em células mononucleares a SOD apresentou atividade diminuída nos indivíduos estadiados como WR 3/4. A GSH-Px não apresentou alteração de sua atividade em decorrência da infecção pelo HIV. Os níveis plasmáticos do &#945;-tocoferol e do ascorbato apresentaram tendência para diminuição, enquanto o &#946;-caroteno não apresentou alteração nos grupos estudados. Estes resultados sugerem que a haptoglobina, as globulinas e a IgA podem ser utilizadas para a avaliação da evolução da infecção pelo HIV. Por outro lado, os níveis dos constituintes do sistema de defesa antioxidante analisados indicam que os indivíduos soropositivos encontram-se menos protegidos contra a ação de espécies reativas de oxigênio, o que favoreceria a presença de um estresse oxidativo e a replicação viral.<br>A total number of 101 individuals, including 26 controls and 75 patients classified according to the Walter Reed Army Institute (28 WR 1, 31 WR 2 and 16 WR 3/4) was studied. HIV infected individuals presented, during the early stages, a progressive reduction of body weigth, as well as urea, albumin, total cholesterol, HDL cholesterol and LDL cholesterol in blood serum. However, increased serum levels of total protein, globulin, IgG, IgA, &#945;1 acid glycoprotein, haptoglobin, AST and LD were observed in HIV infected individuals during the evolution of infection. Decreased serum iron and a trend for increasing triglyceride was shown only for those individuals classified as WR 3/4. Transferrin was diminished only in the WR 2 group. A trend for enhancing serum ferritin following the progession of HIV infection was also observed. No alteration was observed on the levels of reactive \"C\" protein. Decreased EC-SOD activities were observed in HIV infected individuals as compared to controls, whereas in mononuclear cells the SOD activity was diminished only in WR 3/4 patients. HIV infection did not alter GSH-Px activity. A trend for decreasing &#945;-tocopherol and ascorbate plasma levels was shown during the evolution of HIV infected patients, while no difference was observed for &#946;-carotene levels in the studied groups. The above results suggest that haptoglobin, globulins and IgA can be used to assess the evolution of the HIV infection. Moreover, the decreased levels of the antioxidant defense system components observed in HIV infected patients may indicate that they are under an oxidative stress that could favor HIV replication.

Al melanoma cutani, els gens que presenten una major incidència mutacional són els gens BRAF i NRAS. Alteracions en aquests gens resulten en l'activació constitutiva de la via de RAS-ERK1/2, contribuint activament així en el desenvolupament i la progressió tumoral del melanoma. Tot i que ambdues mutacions donen lloc a alteracions de la mateixa via de senyalització, està àmpliament descrit que els tumors que es generen d&#8217;aquestes constitueixen dues entitats diferents tant a nivell molecular com des del punt de vista de la clínica. Una qüestió rellevant al voltant rau en el fet que mentre els melanomes mutats en BRAF disposen de teràpies específiques dirigides contra aquest oncogèn, els melanomes que presenten mutacions en NRAS no tenen tractaments específics. Com a conseqüència, aquests pacients són tractats amb teràpies antitumorals més genèriques, amb taxes de resposta molt menors i a més amb una elevada toxicitat. En aquest context, desemmascar les diferències moleculars existents entre els tumors amb mutacions en BRAF i en NRAS és essencial per a l'establiment de noves estratègies terapèutiques dirigides a pacients que presenten mutacions en NRAS. Resultats obtinguts amb anterioritat al nostre grup de recerca, juntament amb els d'altres investigacions, han confirmat la presència de diferents patrons metabòlics subjectes a la regulació per BRAFV600E. No obstant això, gairebé no existeix evidència al voltant del paper de les mutacions en NRAS a la regulació metabòlica. L'establiment de característiques metabòliques específiques de melanomes amb mutacions en NRAS podria contribuir al desenvolupament de nous enfocaments terapèutics dirigits contra aquest tipus de tumor. Durant el desenvolupament d'aquest estudi hem investigat les implicacions moleculars derivades de la manca de glucosa en cèl·lules de melanoma mutades en NRASQ61 i BRAFV600E, per tal d'establir si la presència de característiques metabòliques depenent de NRAS podria ser explotada per al desenvolupament de noves teràpies contra aquest tipus de tumor. En aquest estudi, hem demostrat la presència de patrons metabòlics sota el control d'NRASQ61. Les cèl·lules que presenten mutacions en NRASQ61 mostren una resposta diferencial a l'estrès metabòlic en comparació amb les cèl·lules mutades en BRAFV600E, donant com a resultat la hiperactivació de la via de RAS-ERK1/2 i a la sensibilització d'aquestes cèl·lules a l'inhibidor multi-cinasa Sorafenib. PFKFB2, PFKFB3 i PFK-1 són elements clau en la regulació d'aquest procés. Amb això, proposem una nova aproximació terapèutica per al tractament dirigit dels melanomes mutats en NRASQ61, establerta per la combinació de 2-deoxi-D-glucosa (2DG) i Sorafenib. Després dels resultats obtinguts, podem concloure que els tumors que presenten mutacions en NRAS i BRAF són entitats diferents a diferents nivells, no només a nivell clínic i molecular, sinó també a nivell metabòlic, el que implica l'existència de noves finestres terapèutiques per al tractament de tumors que presenten mutacions en NRAS.<br>Los genes BRAF y NRAS presentan una mayor incidencia mutacional en melanoma cutáneo. Alteraciones en estos genes resultan en la activación constitutiva de la vía de RAS-ERK1/2, lo que contribuye activamente al desarrollo y la progresión tumoral del melanoma. Aunque ambas mutaciones dan lugar a alteraciones de la misma vía de señalización, ha sido ampliamente descrito que los tumores que se generan de las mismas, constituyen dos entidades diferentes tanto a nivel molecular como desde el punto de vista clínico. Una cuestión relevante reside en el hecho de que mientras los melanomas mutados en BRAF disponen de terapias específicas dirigidas contra el oncogén, los melanomas que presentan mutaciones en NRAS carecen de tratamientos específicos. Como consecuencia, estos pacientes son tratados con tratamientos antitumorales más genéricos, que desembocan en tasas de respuesta mucho menores y en una elevada toxicidad. En este contexto, el desenmascaramiento de las diferencias moleculares existentes entre los tumores con mutaciones en BRAF y en NRAS es esencial para el establecimiento de nuevas estrategias terapéuticas dirigidas a pacientes que presentan mutaciones en NRAS. Resultados obtenidos previamente en nuestro grupo de investigación, sumados a los de otras investigaciones, han confirmado la presencia de diferentes patrones metabólicos sujetos a la regulación por BRAFV600E. Sin embargo, apenas existe evidencia sobre el papel de las mutaciones en NRAS en la regulación metabólica. El establecimiento de características metabólicas específicas de melanomas con mutaciones en NRAS podría contribuir al desarrollo de nuevos enfoques terapéuticos dirigidos contra este tipo de tumor. Durante el desarrollo de este estudio hemos investigado las implicaciones moleculares derivadas de la falta de glucosa en células de melanoma mutadas en NRASQ61 y BRAFV600E, con el fin de establecer si la presencia de características metabólicas dependientes de NRAS podría ser explotada para el desarrollo de nuevas terapias contra este tipo de tumor. En este estudio, hemos demostrado la presencia de patrones metabólicos bajo el control de NRASQ61. Las células que presentan mutaciones en NRASQ61 muestran una respuesta diferencial al estrés metabólico, en comparación con las células mutadas en BRAFV600E, que desemboca en la hiperactivación de la vía de RAS-ERK1/2 y en la sensibilización de estas células al inhibidor multi-quinasa Sorafenib. PFKFB2, PFKFB3 y PFK-1 son elementos clave en la regulación de este proceso. Adicionalmente, proponemos una nueva aproximación terapéutica para el tratamiento dirigido de los melanomas mutados en NRASQ61, basada en la combinación de 2-deoxi-D-glucosa (2DG) y Sorafenib. Tras los resultados obtenidos, podemos concluir que los tumores que presentan mutaciones en NRAS y BRAF son entidades diferentes a distintos niveles, no solo a nivel clínico y molecular, sino también a nivel metabólico, lo que implica la existencia de nuevas ventanas terapéuticas para el tratamiento de tumores que presentan mutaciones en NRAS.<br>BRAF and NRAS are the most commonly found mutated genes in cutaneous melanoma. Alterations in these genes result in the constitutive activation of the RAS-ERK1/2 pathway, contributing to tumor development and progression. Beside both genes are consecutive located in the same signaling cascade, BRAF and NRAS mutated tumors are considered two different entities at clinical and molecular levels, resulting in distinct signaling patterns and different biological behavior. Furthermore, while there is a first line of treatment using targeted therapy against BRAF mutant melanomas, NRAS mutant tumors remain without specific line of treatment, showing low response rates and high toxicity to the currently applied therapies. Thus, the understanding of the molecular differences between BRAF and NRAS mutant tumors is essential to improve therapeutic opportunities for the treatment of patients carrying NRAS mutations. Previous results in our group, together with additional investigations, have highlighted the presence of different metabolic settings subjected to BRAFV600E oncogene regulation. However, little is known about the role of NRAS mutations in metabolic rewiring. Deciphering metabolic settings in NRAS mutant melanomas could provide new avenues for the establishment of specific therapeutic approaches against these, until now, untargetable tumors. In this study, we have investigated the molecular implications of glucose starvation in NRASQ61 and BRAFV600E mutant cells in order to establish whether the presence of NRAS-dependent metabolic settings can be exploited for the development of targeted therapies against NRAS mutant melanomas. Overall, in this study we have demonstrated the presence of NRASQ61 oncogene-dependent metabolic settings. NRASQ61 mutant cells show a differential response to metabolic stress when compared to BRAFV600E mutant cells, which results in the hyperactivation of the RAS-ERK1/2 pathway and the sensitization to the multikinase inhibitor Sorafenib. PFKFB2, PFKFB3 and PFK-1 are key players in the regulation of this process. We also propose a novel approach for the specific targeting of NRASQ61 mutant melanomas based on the combination of 2-deoxy-D-glucose (2DG) and Sorafenib. We conclude that NRAS and BRAF mutant tumors are different entities at different levels, not only at molecular and clinical levels but also at metabolic level and this fact provides a new therapeutic window for the targeting of NRAS mutant tumors.<br>Universitat Autònoma de Barcelona. Programa de Doctorat en Bioquímica, Biologia Molecular i Biomedicina

Les espècies més longeves han evolucionat disminuint la producció endògena d’espècies reactives d’oxigen i proveint-se d’estructures resistents a la oxidació. Per tant, aquelles espècies que viuen més gaudeixen de mitocòndries metabòlicament més eficients i estructuralment més estables. De fet, característiques fenotípiques de la longevitat inclouen la reducció del contingut del complex I i dels aminoàcids sulfurats. Aleshores, l’activitat de determinades vies de senyalització intracel·lulars juga un paper clau regulant l’expressió de gens associats a un fenotip longeu. En aquest context, aquesta tesi pretén determinar i) la modulació de determinades subunitats del complex I associada a la longevitat; ii) els canvis en el contingut dels aminoàcids sulfurats i els seus intermediaris metabòlics en teixits post-mitòtics i iii) plasma d’espècies més longeves; iv) la regulació del contingut dels diferents elements específics del complex 1 de mTOR en termes de longevitat; i v) l’existència un perfil metabòlic associat a humans de longevitat extrema. Els resultats obtinguts mostren l’existència de perfils metabòlics associats a la longevitat de les espècies que, en alguns casos, són diferents a aquells perfils associats a la longevitat individual. A més, les espècies més longeves han evolucionat disminuint el contingut de determinades subunitats del complex I que podrien ésser responsables de la menor producció d’espècies reactives d’oxigen. Per altra banda, existeixen factors genètics que podrien determinar l’activitat basal de mTORC1, i que podrien, almenys en part, explicar el fenotip associat a la longevitat. Per tant, sembla que l’assoliment d’una major longevitat implica una adaptació metabòlica i estructural.<br>Las especies más longevas han evolucionado disminuyendo la producción endógena de especies reactivas de oxígeno y proveyéndose de estructuras resistentes a la oxidación. Por lo tanto, aquellas especies que viven más disfrutan de mitocondrias metabólicamente más eficientes y estructuralmente más estables. De hecho, características fenotípicas de la longevidad incluyen la reducción del contenido del complejo I y de amino ácidos sulfurados. Por lo tanto, la activad de determinadas vías de señalización intracelular juegan un papel clave regulando la expresión de genes asociados a un fenotipo longevo. En este contexto, esta tesis pretende determinar i) la modulación de determinadas subunidades del complejo I asociada a la longevidad; ii) los cambios en el contenido de amino acido sulfurados y de sus intermediarios metabólicos en tejidos post-mitóticos y iii) plasma de especies más longevas; iv) la regulación del contenido de distintos elementos específicos del complejo 1 de mTOR en términos de longevidad; y v) la existencia de un perfil metabólico asociado a humanos de longevidad extrema. Los resultados obtenidos muestran la existencia de perfiles metabólicos asociados a la longevidad de las especies que, en algunos casos, son diferentes a aquellos perfiles asociados a la longevidad individual. Además, las especies más longevas han evolucionado disminuyendo el contenido de determinadas subunidades del complejo I que podrían ser responsables de la menor producción de especies reactivas de oxígeno. Por otra parte, existen factores genéticos que podrían determinar la actividad basal de mTOR, y que podrían, al menos en parte, explicar el fenotipo asociado a la longevidad. Por lo tanto, parece que lograr una mayor longevidad implica una adaptación metabólica y estructural.<br>Long-lived species have evolved by decreasing the rate of endogenous reactive oxygen species production and providing them of oxidation-resistant structures. Hence, species that live longer benefit from metabolically efficient and structurally stable mitochondria. In fact, phenotypic traits of longevity include reduced content of complex I and sulphur-containing amino acids. Then, the activity of selected intracellular signalling pathways plays a key role regulating the expression of genes associated to a longevity phenotype. In this context, this thesis aims to determine i) the modulation of specific complex I subunits associated to longevity; ii) the changes on sulphur amino acids content and its metabolic intermediates in post-mitotic tissues and ii) plasma from long-lived species; iv) the content regulation of the different mTOR complex 1 specific forming elements in terms of longevity; and v) the existence of a metabolic profile associated to human extreme longevity. The obtained results reveal the existence of metabolic profiles associated to species longevity that, in some cases, differ from those profile associated to individual longevity. Furthermore, longer lived species have evolved reducing the content of specific complex 1 subunits that might be responsible for the limited reactive oxygen species production. Otherwise, genetic factors that might determine the basal activity of mTORC1 exist, and that could, at least In part, explain the longevity associated phenotype. Thus, it seems that the achievement of an extended longevity implies a metabolic and structural adaptation.

Orientador: Carmen Silvia Fernandes Boaro<br>Resumo: Espécies vegetais são capazes de produzir diversidade de substâncias, que desempenham funções importantes para sua sobrevivência e adaptação ao ecossistema. O metabolismo primário, é essencial para o crescimento, desenvolvimento, maturação e reprodução de qualquer espécie. O metabolismo especializado, dependente do primário, é responsável por originar o óleo essencial, que são misturas de metabólitos especializados voláteis, representados principalmente por monoterpenos e sesquiterpenos. Cada espécie vegetal produz um óleo essencial de composição característica específica, podendo ser influenciado por fatores bióticos e abióticos. A fenologia pode influenciar processos bioquímicos e rotas metabólicas capazes de modificar a formação de substâncias biologicamente ativas, alterando diretamente o conteúdo e a qualidade dos óleos essenciais. Sendo assim, o objetivo deste trabalho foi avaliar se as fases fenológicas, vegetativa e reprodutiva modificam o desempenho fotossintético e o perfil do óleo essencial de Xylopia aromatica (Lam.) Mart., influenciando sua atividade biológica na defesa antioxidante e ação antifúngica. As variáveis, fluorescência da clorofila a, trocas gasosas, carboidratos, atividade enzimática e peroxidação lipídica, potencial água, conteúdo relativo de água das folhas, extração, rendimento, caracterização química e atividade antifúngica do óleo essencial de Xylopia aromatica foram avaliadas em 24 plantas, 12 no estádio vegetativo e 12 no reprodutivo, coletadas... (Resumo completo, clicar acesso eletrônico abaixo)<br>Abstract: Research aimed at the knowledge of plant species allows the elaboration of projects that aim at the understanding of development, conservation of biodiversity and sustainable exploitation of natural resources. The primary metabolism, represented by photosynthesis and the specialized one, that synthesizes the essential oil, can be influenced by the environmental and phenological conditions, which can influence the chemical profile of the essential oil and the biological activity in the vegetal defense, including against fungi, bacteria and virus. Compounds from the specialized metabolism present biological activity and potential for the production of bactericides and fungicides. Therefore, it is necessary to know the stage of development of plant species in which the substances of interest, with economic potential, are more concentrated, thus orienting, if appropriate, the collection period, aiming at the conservation and sustainable use. There are scientific studies that reveal biological activity of essential oils, as observed for the genus Xylopia, but none of them relates the primary and specialized metabolism to the stage of development in which the species is found. In this way, the objective of this research was to evaluate if the phenological, vegetative and reproductive phases of Xylopia aromatica (Lam.) Mart. modify the photosynthetic performance and the profile of the essential oil, which may influence its biological activity in the antioxidant defense and antifunga... (Complete abstract click electronic access below)<br>Mestre

La GK és clau en la regulació del metabolisme glucídic tant per la seva funció al pàncrees en què determina la secreció d’insulina, com per la seva funció al fetge on controla les vies d’utilització i d’emmagatzematge de glucosa. Per aquest motiu és una bona diana per a la teràpia de la diabetis, i diverses companyies farmacèutiques estan desenvolupant activadors sintètics de la GK (GKA) pel tractament de la diabetis de tipus 2. Els estudis de GKAs existents avaluen l’activació de la GK a nivell sistèmic i no es coneixen els seus efectes específis sobre la GK al fetge. Per indagar-los, com que no disposàvem d’un GKA específic per a la GK del fetge, ens vam proposar la sobrexpressió d’una forma de GK amb una mutació activadora (GK-A456V) que confereix característiques cinètiques a l’enzim idèntiques a les obtingudes amb els GKAs. L’introduírem al fetge perivenós mitjançant transfecció gènica hidrodinàmica (perquè és la zona on s’expressa majoritàriament l’enzim salvatge en condicions fisiològiques). Un dels controls emprats en l’estudi fou la sobrexpressió de la GK salvatge també a la zona perivenosa, que ens permetria comparar els resultats d’aquesta sobrexpressió local de la GK amb la sobrexpressió no zonal descrita a la bibliografia. Per una banda, avaluàrem els efectes de la sobrexpressió i de l’activació de la GK hepàtica en el context d’un animal sa a llarg termini, per determinar si hi havia risc de resistència a insulina (com s’esdevé en alguns models de sobrexpressió no zonal de GK al fetge). Tant l’activació com la sobrexpressió de GK al fetge perivenós a curt termini comportaren una disminució de la glicèmia i de la insulinèmia. No obstant, en avaluar-ho a llarg termini, la sobrexpressió de GK generà un fenotip de resistència a insulina exclusivament al fetge. En canvi, la GK-A456V comportà un fenotip amb un perfil metabòlic similar als animals controls, sense alteracions dels nivells de glúcids i lípids (sèrics i hepàtics). Al fetge, malgrat que no provocava canvis en el metabolisme glucídic i lipídic, l’activació de la GK va promoure la desregulació per GKRP i la inducció de la glucosa-6-fosfatasa. Per altra banda, la presència de la GK activada al fetge resultà en una activació del catabolisme al teixit adipós. Per altra banda, vam emprar un model de diabetis de tipus 1 per determinar els efectes de la GK-A456V sobre el fenotip diabètic independentment de la insulina, amb la finalitat de valorar l’activació de la GK hepàtica com a tractament alternatiu als GKAs sistèmics. La sobrexpressió perivenosa de GK, tot i que comportà un increment del metabolisme de glucosa al fetge (increment del glicogen hepàtic, de ub-PFK-2, de L-PK, c-myc) i una disminució de la gluconeogènesi (reducció dels nivells de PEPCK), va provocar dislipidèmia com a resultat de la disminució de la β-oxidació d’àcids grassos (reducció dels nivells de Cpt-1) i una inducció de la lipogènesi hepàtica (augment de FAS, ACC, ChREBP). Altres models de sobrexpressió de GK, descrits a la literatura, que no tenien en compte el concepte de zonació hepàtica, també presentaven alteracions en el metabolisme lipídic. En canvi, l’activació de la GK comportà una rellevant disminució de la hiperglicèmia diabètica tot i tenir un lleu efecte sobre l’activació del metabolisme de glucosa i sobre la inhibició de la gluconeogènesi. Sobretot és interessant que la millora de la hiperglicèmia diabètica no va acompanyada d’alteracions del metabolisme lipídic. En conjunt, aquest treball suggereix que l’activació de la GK exclusivament al fetge és una estratègia terapèutica millor per a la diabetis que la sobrexpressió de la GK.<br>Synthetic glucokinase activators have been used in the context of type 2 diabetes therapy, mainly for their insulin secretagogue activity. However, the impact of these drugs on liver GK has not been studied in vivo. Since GK activators and activating GK mutations confer identical kinetic properties to GK, we hypothesize that hepatic overexpression of a mutated form of GK, GKA456V, described in a patient with Persistent Hyperinsulinemic Hypoglycemia of Infancy (PHHI), shall mimic the liver-specific effects of GK-activating drugs. GKA456V was overexpressed in the liver of streptozotocin diabetic mice and also in healthy mice. Metabolite profiling in serum and liver extracts, together with key components of glucose and lipid homeostasis, were analyzed and compared to GK wild-type transfected animals. Cell compartmentalization of mutant and wild-type GK was also examined in vivo. In the type 1 diabetic mice, GKA456V overexpression markedly reduced blood glucose in the absence of dislipidemia, in contrast to wild-type GK-overexpressing mice. Enhanced glucose utilization did not correlate with glycogen synthesis or lactate production. PEPCK mRNA was not affected, whereas the mRNA for the catalytic subunit of glucose-6-phosphatase was upregulated ~4-fold in the liver of GKA456V treated animals. Moreover, GKA456V was not translocated to the nucleus after a short fast, confirming that this activating mutation disrupted GKRP regulation. In healthy mice, the overexpression of hepatic GK resulted in insulin resistance. Otherwise, GKA456V overepxressing animals were not insulin resistant. They showed increased mRNA and protein content of the catalytic subunit of glucose-6-phosphatase in the liver, and an idnuction of catabolism in their adipose tissue. Our results validate liver specific GK activation as a strategy for diabetes therapy and provide new insights into the complex GK regulatory network.

Els xenobiòtics són compostos exògens als éssers vius, i encara que no formen part de la seva bioquímica normal, s´incorporen a les seves vies metabòliques. Els xenobiòtics entren a l´organisme per diferents vies: oral (p.o.), intravenosa (i.v.) subcutània (s.c.), intramuscular (i.m.), respiratòria, etc. Els xenobiòtics es poden classificar segons el seu origen i segons els seus efectes. Els organismes, per la seva part, han desenvolupat sistemes de detoxificaxió per eliminar-los. En aquest treball, s´estudien xenobiòtics que tenen o estan desenvolupats per tenir una activitat terapèutica (fàrmacs). Aquests compostos són normalment de naturalesa lipofílica, de forma majoritària amb capacitat de difondre per les membranes cel.lulars, encara que alguns interaccionen amb transportadors específics. Un cop dins de la cèl.lula són normalment difícils d´eliminar. Per aquest motiu, els organismes transformen els xenobiòtics mitjançant processos metabòlics, i així faciliten la seva eliminació. En aquest procés de detoxificació intervenen un conjunt d´enzims poc específics que reconeixen una àmplia gamma de compostos. Les reaccions de metabolisme d´aquests enzims s´anomenen reaccions de biotransformació de fase I, i reaccions de conjugació o de fase II. En el present treball s´estudien les reaccions de fase I, que són processos d´oxidació, reducció i hidròlisi que es donen a temperatura fisiològica. El Citocrom P450, CYP450, és el principal complexe enzimàtic encarregat de les reaccions de fase I, i es caracteritza per la gran varietat de processos que poden catalitzar així com la quantitat de substractes diferents que poden metabolitzar. L´estudi es porta a terme mitjançant experiments in vitro en un sistema d´incubacions en microsomes de fetge de rata. Les isoformes on s´estudien les possibles interaccions en la seva activitat són: CYP1A1/2, CYP2A1/2, CYP2B1/2, CYP2C, CYP2C11, CYP2D1, CYP2E1 i CYP3A1/2. Per assolir aquest objectiu global, es va obtenir i caracteritzar el sistema experimental (microsomes de fetge de rata), es van posar a punt sistemes analítics per avaluar les cinètiques enzimàtiques, i es van desenvolupar models cinètics pel tractament de les dades experimentals.

The TP53-Induced Glycolysis and Apoptosis Regulator (TIGAR) gene was described in 2006 by Dr. Karen Vousden's group in response to the induction of the tumour suppressor gene p53. Since then, numerous studies have focused on clarifying the role of this gene in the metabolism of tumour cells. TIGAR was initially described as an enzyme with bisphosphatase activity on fructose-2,6-bisphosphate, a key metabolite in the positive allosteric regulation of phosphofructokinase-1, which catalyses a key reaction in glycolysis. Through this bisphosphatase activity, TIGAR reduces the levels of fructose-2,6-bisphosphate and, consequently, decreases the glycolytic flux and redirects the metabolites to the pentose phosphate pathway, which is determinant for the antioxidant capacity of cells. Overexpression of TIGAR has been described in multiple tumours, as well as in different cell lines, indicating that this gene confers a growth advantage to these cells. The present doctoral thesis has focused on studying the metabolic function of TIGAR in tumours, as well as the mechanisms that regulate its transcription. The study was carried out on cell lines of cervical cancer and lung cancer in which we have been able to confirm that TIGAR exerts a bisphosphatase function on fructose-2,6-bisphosphate. TIGAR has proven to be key in the response of the cervical cancer cell line HeLa to the blockage of glycolysis, either by inhibiting the expression of the PFKFB3 gene by interfering RNA technology, or by blocking the PFK-2 protein by the drug 3PO. The blockage of glycolysis increases oxidative stress and the phosphorylation of the kinase Akt, which is required for the induction of TIGAR. Furthermore, through metabolomic studies we have been able to describe for the first time the involvement of TIGAR in the entrance of pyruvate to the tricarboxylic acid cycle, in the mitochondria. Finally, and in relation to the mechanisms that regulate the transcription of TIGAR, we have proved that the transcription factor Nrf2, key in the regulation of the antioxidant activity of tumour cells, controls the expression of TIGAR in HeLa cells. In lung cancer cells, where the over activation of Nrf2 is related to chemo resistance and radiotherapy, the relationship between Nrf2 and TIGAR seems to be indirect. With the results presented in this doctoral thesis we have contributed to a better understanding of the role of TIGAR in tumour metabolism and have laid the foundations for future studies aimed at blocking this protein in tumours.<br>El gen TP53-Induced Glycolysis and Apoptosi Regulator (TIGAR) va ser descrit l'any 2006 pel grup de la Dra. Karen Vousden en resposta a l’activació del supressor tumoral p53. Des de llavors, nombrosos estudis s'han centrat en aclarir el paper d'aquest gen en el metabolisme de les cèl·lules tumorals. Inicialment, la funció atribuïda a TIGAR va ser la de bisfosfatasa de la fructosa-2,6-bisfosfat, metabòlit clau en la regulació al·lostèrica positiva de l’enzim fosfofructoquinasa-1, que catalitza la una reacció clau en la glucòlisi. Mitjançant aquesta activitat bisfosfatasa, TIGAR redueix els nivells de fructosa-2,6-bisfosfat i, en conseqüència, frena en flux glicolític i redirigeix els metabòlits a la via de les pentoses fosfat. És per aquest motiu que TIGAR es va descriure com un gen amb capacitat antioxidant. La present tesi doctoral s'ha centrat en estudiar la funció metabòlica de TIGAR en línies tumorals, així com els mecanismes que regulen la seva transcripció. Amb aquests estudis hem pogut demostrar que TIGAR és clar en la resposta de les cèl·lules al bloqueig de la glucòlisi, ja sigui per la inhibició de l'expressió del gen PFKFB3 mitjançant la tecnologia de RNA d'interferència, com pel bloqueig de la proteïna PFK-2 mitjançant el fàrmac 3PO. El bloqueig de la glucòlisi provoca un augment de l'estrès oxidatiu i de la fosforil·lació de la quinasa Akt, necessària per a la inducció de TIGAR.que al seu torn condueix a una inducció de TIGAR. D’altra banda, estudis metabolòmics ens han permès descriure per primera vegada l’acció de TIGAR en nivells inferiors de la glicòlisi, afectant l’entrada del piruvat al cicle de Krebs. Finalment, hem pogut comprovar que el factor de transcripció Nrf2, clau en la regulació de l'activitat antioxidant de les cèl·lules, controla l'expressió de TIGAR en una línia cel·lular de càncer de cèrvix. En cèl·lules de càncer de pulmó, en canvi, la relació entre Nrf2 i TIGAR sembla ser indirecta. Amb els resultats presentats en aquesta tesi doctoral hem contribuït a entendre millor el paper de TIGAR en el metabolisme tumoral i hem establert les bases per a futurs estudis dirigits al bloqueig d'aquesta proteïna als tumors.

Orientador: Carmen Silvia Passos Lima<br>Dissertação (mestrado) - Universidade Estadual de Campinas. Faculdade de Ciências Médicas<br>Made available in DSpace on 2018-08-16T03:51:55Z (GMT). No. of bitstreams: 1 Scherr_MarcelaCustodio_M.pdf: 1746928 bytes, checksum: 1590f1bcf4c880aa0dbd063756c2c777 (MD5) Previous issue date: 2010<br>Resumo: A habilidade de metabolizar xenobióticos é variável em humanos e em cães e está relacionada a genótipos distintos de genes polimórficos. Os portadores do genótipo CYP1A2 1117TT foram classificados como metabolizadores lentos de fármacos. Ainda, os alelos variantes T e C dos polimorfismos CYP1A2 C1117T e GSTA3 I71L foram descritos como menos eficazes na ativação e inativação de carcinógenos do que os selvagens, respectivamente. Assim, humanos e cães podem apresentar diferentes respostas a medicamentos e estar sob riscos distintos de ocorrência de tumores. As frequências dos genótipos dos polimorfismos CYP1A2 C1117T e GSTA3 I71L em cães de diferentes raças e sem raça definida (SRD) não foram ainda descritas. O objetivo deste estudo foi o de identificar as frequências dos genótipos em 105 cães da raça boxer, 84 cães da raça rottweiler, 110 cães da raça pastor alemão e 99 animais SRD, para verificar se ocorre distribuição distinta de genótipos em animais de diferentes grupos. O genótipos foram identificados por amplificação das regiões gênicas de interesse, em amostras de sangue periférico, por meio da reação em cadeia da polimerase seguida por sequenciamento ou digestão enzimática dos fragmentos amplificados. A frequência do genótipo homozigoto selvagem CC do polimorfismo CYP1A2 C1117T foi maior em cães de raça (96,4% versus 87,9%, P= 0,002) e boxers (98,0% versus 87,9%, P= 0,005) do que em cães SRD. Frequências genotípicas similares foram observadas em cães estratificados por aspectos clínicos. Apenas o genótipo variante CC do polimorfismo GSTA3 I71L foi identificado em nossos cães. Nossos resultados sugerem que cães de raça metabilizam fármacos e ativam carcinógenos de forma mais eficaz do que cães SRD e podem obter maior benefício medicamentoso e estar sob risco maior de tumores. Já o metabolismo de xenobióticos pela enzima GSTA3 parece similar em cães de diferentes raças e não influenciar diferentemente a resposta a medicamentos e o risco de tumores nesses animais<br>Abstract: The ability to metabolize xenobiotics is variable among humans and dogs, and is related to the distinct genotypes of polymorphic genes. Carriers of the CYP1A2 1117TT genotype were classified as poor metabolizers of drugs. In addition, the varying T and C alleles of CYP1A2 C1117T e GSTA3 I71L polymorphisms were described as less efficient in the activation and inactivation of carcinogens than the wild alleles, respectively. Thus, humans and dogs seem to obtain distinct benefits under drug treatments and also to be under different risks of tumours. The frequencies of genotypes of the refered genes in dogs of different breeds and mixed breed are yet not described. The objective of this study was to identify the frequencies of the genotypes in 105 boxers, 84 rottweilers, 110 german shepherds and 99 mixed breed dogs with the purpose of to verify if a distinct distribution of genotypes occurs in animals of different groups.The genotypes were evaluated by amplifying the area of interest of the genes, in peripheral blood samples, using a polymerase chain reaction followed by a sequencing or enzymatic digestion of the amplified fragments. The frequency of the homozygous wild CC genotype of the CYP1A2 C1117 polymorphism was higher in dogs of pure breed (96,4% versus 87,9%, P= 0.002) and boxers (98.0% versus 87,9%, P= 0.005) than in mixed breed dogs. Similar frequencies of the genotypes were seem in dogs stratified by clinical features. Just the CC varying genotype of the GSTA3 I71L polymorphism was identified in all dogs. The results of this study suggest that pure breed dogs might be more efficient in metabolizing drugs and in activating carcinogens than mixed breed dogs, and therefore, may obtain larger therapeutic benefits under treatment for diseases and may be under higher risk for tumors. In contrast, the metabolim of xenobiotics by the GSTA3 enzyme seems to be similar among dogs of different breeds and not to influence on a different way the drug metabolism or tumour risk in these animals<br>Mestrado<br>Biologia Estrutural, Celular, Molecular e do Desenvolvimento<br>Mestre em Fisiopatologia Médica

A lipoaspiração é a cirurgia estética mais comumente realizada pelas mulheres no mundo. Uma vez que o tecido adiposo é um órgão metabolicamente ativo, alguns autores sugeriram que a remoção cirúrgica do tecido adiposo subcutâneo (TAS) pudesse exercer efeitos importantes no perfil metabólico. Além disso, estudos experimentais observam ganho de gordura compensatório no tecido adiposo intacto em resposta à lipectomia. O exercício físico regular induz a inúmeras melhoras no perfil metabólico e na composição corporal, promovendo o aumento do gasto energético total e preservação da massa magra. Estudos sobre os efeitos integrados do exercício e da lipoaspiração em humanos são inexistentes na literatura. Desta forma, este estudo visou avaliar o efeito da lipoaspiração acompanhada de treinamento físico sobre o perfil metabólico, a adiposidade e a distribuição de gordura corporal em mulheres adultas e eutróficas (20 a 35 anos; IMC: 23,8 ± 2.2 Kg/m2). Trinta e seis mulheres foram submetidas à cirurgia de lipoaspiração abdominal. Dois meses após a cirurgia, as voluntárias foram divididas em dois grupos: treinado (LT; n=18) e sedentário (LS; n=18). O treinamento físico consistiu de treinamento aeróbio e de força, 3x/semana, por 16 semanas. A metodologia empregada incluiu avaliação do peso corporal, composição corporal (pesagem hidrostática), distribuição da gordura abdominal (tomografia computadorizada), consumo alimentar (diário alimentar), perfil lipídico, concentração de citocinas anti e proinflamatórias, de adiponectina e de leptina antes (PRE), dois meses após a cirurgia (POS-2) e seis meses após a cirurgia (POS-6). A capacidade física (determinada pelo VO2max, e por testes de repetição máxima para membros inferiores e superiores), a taxa metabólica de repouso (TMR) (calorimetria indireta), a determinação do diâmetro médio dos adipócitos e a expressão gênica dos fatores de transcrição para adipogênese, das citocinas anti e proinflamatórias, da leptina e da adiponectina no tecido adiposo subcutâneo abdominal e femoral foram avaliadas nos tempos PRE e POS-6. O consumo alimentar permeceneu inalterado ao longo do estudo. Seis meses após a cirurgia, o grupo LS apresentou tendência ao retorno do peso e da massa gorda total aos valores basais, bem como aumento do tecido adiposo visceral (TAV) e queda da TMR. O grupo LT também apresentou queda da TMR, mas manteve os valores de peso corporal e de massa gorda diminuídos em resposta à cirurgia no POS-6 e apresentou aumento da massa magra e manutenção do TAV. Ambos os grupos apresentaram queda das concentrações de adiponectina e aumento do colesterol total, da LDL-colesterol e da razão LDL/Apolipoproteína B no POS-6. Por meio dos resultados, podemos concluir que a remoção do tecido adiposo abdominal subcutâneo (TAS) em sujeitos eutróficos parece desencadear mecanismos que favorecem a reposição e o crescimento compensatório de tecido adiposo, em especial na cavidade visceral, além de acarretar na diminuição das concentrações de adiponectina, sugerindo possíveis efeitos deletérios no risco cardiovascular em longo prazo. A prática de exercício físico após a cirurgia é de extrema importância para preservar os efeitos da cirurgia na composição corporal, prevenir contra o crescimento compensatório de gordura na cavidade visceral e atenuar seus possíveis efeitos deletérios no metabolismo em longo prazo<br>Liposuction is the most communly cosmetic surgery performed worlwide. Because adipose tissue is metabolicaly active, researchers have suggested that the surgical removal of fat through liposuction could benefically affect the metabolic profile. In addition, in many species, when body fat is removed, it is recovered rapidly due to compensatory fat growth at intact depots. Regular exercise training benefits metabolic profile and body composition by increasing energy expenditure and preserving fat free mass. Studies about the associated effects of liposuction and physical exercise lack in the literature. Thus the aim of this study was to evaluate the effects of liposuction associated with exercise training on metabolic profile, adiposity and body fat distribution in adult normal weight women (20 to 35 years old, BMI: 23,8 ± 2.2 Kg/m2). Thirty-six women underwent a small-volume abdominal liposuction (mean fat aspirate supernantant: 1240.3 ± 363.6 ml). Two months after surgery were randomly divided into two groups: trained (TR; n=18) and sedentary control (SC; n=18). The four-month exercise program consisted of aerobic plus resistance training, thrice a week. Body composition (hydrostatic weighing), body fat distribution (computer tomography), dietary intake (food records), lipid profile, plasmatic concentration of citokyne, adiponectin and leptin were assessed at baseline (PRE), two (POST-2) and six months after surgery (POST-6). Physical capacity (by VO2max, one repetition maximum (1RM) bench and leg press ), resting energy expenditure (TMR - indirect calorimetry), adipocyte size and gene expression of adipogenesis transcription factors, leptin, adiponectin and citokyne were assessed at PRE and POST-6. Dietary intake was unchanged throughout the study. Six months after surgery, LS group showed increased visceral adipose tissue (TAV), decreased TMR and a tendency to return body weight and fat to baseline values. LT group also showed decreased TMR, but sustained liposuction-induced decreased body weight and fat, increased fat free mass and preserved TAV at POST-6. Both groups showed decreased levels of adiponectin and increased levels of total cholesterol, LDL, cholesterol and LDL/ApoB ratio at POST-6. In conclusion, abdominal subcutaneous fat removal in normal weight subjects triggers a compensatory increase of fat, specially towards the visceral cavity, and lowers adiponectin levels, which could enhance long-term cardiovascular risk. Additionally, exercise training plays a very important role in preserving against the compensatory increase of visceral fat and attenuating possible long-term deleterious effects

La 3,4-metilenodioximetanfetamina (MDMA, éxtasis) es una anfetamina sustituida de consumo frecuente y abusivo. La enzima principal que participa en el metabolismo de fase I de la MDMA, la isoforma 2D6 (CYP2D6) del citocromo P450, resulta también inhibida por la MDMA. Además, ésta es a la vez metabolizada por otras isoformas de CYP, por ejemplo la CYP1A2. La contribución de esta enzima y los posibles cambios en su actividad tras una administración de MDMA nunca han sido estudiados in-vivo. En consecuencia, se realizó un ensayo clínico donde los marcadores, dextrometorfano y cafeína se administraron tras una dosis oral de MDMA. En base a la farmacocinética de ambos marcadores se evaluaron posibles cambios en la actividad de las enzimas. En base al ratio metabólico urinario de dextrometorfano i dextrorfano (MR) se calculó la vida-media de degradación de CYP2D6. Tras una dosis de MDMA, el Cmax i AUC del dextrometorfano aumentó aproximadamente 10 veces con la correspondiente disminución en los parámetros farmacocineticos de dextrorfano. Se aumentó el MR casi 100 veces después de una dosis de MDMA, con un 67% de los sujetos superando la antimoda de 0.3 para la asignación del fenotipo de metabolizador lento de CYP2D6. La actividad de CYP2D6 se recuperó después de 10 días con una vida media de degradación de CYP2D6 de 46.6 h. La farmacocinética de la cafeína y sus metabolitos no fue afectada por la MDMA. Se debería avisar los consumidores de MDMA de las consecuencias de tal inhibición. A pesar de que hay muchas evidencias en animales sugiriendo que el MDMA es una neurotoxina serotonergica, todavía hay mucho debate sobre cuál es la causa de estos cambios cerebrales a largo plazo. La investigación apunta a la producción excesiva de especies reactivos de oxigeno (ROS) en el cerebro después de la administración de MDMA. La MDMA induce hipertermia, la liberación excesiva de dopamina cerebral y lleva a la desregulación energética del metabolismo. Conjuntamente, el metabolismo de MDMA produce un catecol reactivo, cuyos productos causan neurotoxicidad serotonergica en ratas. Cualquiera de los factores anteriores podrían ser la causa de la excesiva producción de ROS y los consecuentes cambios serotonergicos. A tenor de estas hipótesis, se investigó si diferentes temperaturas corporales afectarían el metabolismo de MDMA. Se administró MDMA a ratas a tres temperaturas ambientales distintas con el fin de prevenir o exacerbar la hipertermia inducida por MDMA. Se determinaron las concentraciones plasmáticas de MDMA y sus metabolitos principales durante las 6 h posteriores a la administración de la droga. Después de siete días, se sacrificaron los animales y se determinaron las cantidades de índoles cerebral. La administración de MDMA a 15ºC bloqueó la respuesta hipertermica y la disminución a largo plazo de 5-HT encontrada en ratas administradas a 21.5 ºC. A 15ºC, las concentraciones plasmáticas de MDMA aumentaron significativamente mientras que las concentraciones de sus metabolitos disminuyeron en comparación con ratas administradas a 21.5ºC. En contraste, la hipertermia y las deficiencias de indoles fueron exacerbadas en ratas tratadas a 30ºC. Se observó que las concentraciones plasmáticas de metabolitos de MDMA aumentaron significativamente en estos animales. La depleción a largo plazo de 5-HT no estuvo potenciada por la perfusión intrastriatal de MDMA después de una dosis sistémica de MDMA. Además, la interferencia del metabolismo de MDMA con la administración del inhibidor de catecol-o-metiltransferasa, entacapona, potenció la neurotoxicidad de MDMA, indicando que los metabolitos que son sustratos para este enzima podrían contribuir a la neurotoxicidad. Estos resultados tienen implicaciones tanto con el papel de la temperatura en el mecanismo del desarrollo de la neurotoxicidad del MDMA como en el abuso en humanos donde la hipertermia esta asociado con casos de toxicidad aguda. Se ha sugerido también que la causa de la depleción de 5-HT por MDMA es debido a un aumento de niveles de tirosina en el cerebro, cuyo hidroxilación no enzimática conduce a la formación de radicales libres derivados de la dopamina. En consecuencia, se propuso que el metabolismo de MDMA en compuestos pro-oxidantes fuera el paso limitante del proceso. En una serie de experimentos se encontraron niveles más altos de hipertermia aguda, concentraciones plasmáticas de tirosina, MDMA y sus metabolitos después de una dosis toxica de MDMA (15 mg/kg i.p.) versus una dosis no-toxica (7.5 mg/kg i.p.). La administración de una dosis no-toxica de MDMA (7.5 mg/kg i.p.) en conjunto con L-tirosina (0.2 mmol/kg i.p.) produció un aumento similar de niveles de tirosina en el suero con los niveles encontrados tras una dosis toxica de MDMA, sin embargo, los niveles de 5-HT cerebral permanecieron en niveles normales. Una dosis no-toxica de MDMA en combinación con una dosis alta de tirosina (0.5 mmol/kg i.p.) causó depleciones a largo plazo en ratas administradas a 21.5ºC pero no a 15ºC, condiciones conocidas por disminuir el metabolismo de MDMA. Al mismo tiempo, la perfusión estriatal de MDMA en combinación con tirosina (0.5 mmol/kg i.p.) en ratas hipertermicas no causaron depleciones de 5-HT. En contraste, se observaron reducciones significativas en 5-HT cerebral tras la administración de una dosis no-toxica de MDMA en ratas en condiciones de hipertermia en combinación con entacapona o acivisina, compuestos capaces de interferir con el metabolismo de MDMA o aumentar la disponibilidad de sus metabolitos en el cerebro, respectivamente. En conjunto estos datos indican que a pesar de que la tirosina y la hipertermia pueden contribuir a la neurotoxicidad inducida por la MDMA, el metabolismo de la droga parece ser el paso limitante.

Cada cop més evidencies suggereixen que els astròcits participen en les altes funcions cerebrals, controlant des de la transmissió sinàptica fins a les ones cerebrals globals i els processos d’aprenentatge i memòria. Diferents mecanismes han sigut proposats com a responsables d’aquests processos mediats per astròcits, entre ells, l’alliberació de gliotransmissors a partir de les senyals de calci així com la de lactat semblen els principals efectors. L’existència d’aquest control de les funcions cerebrals per part dels astròcits suggereix que aquestes cèl·lules poden regular les funcions cerebrals en resposta a experiència tan com les neurones, constituint el fenomen de plasticitat astrocitària. En neurones s’ha demostrat que el conegut factor de transcripció CREB, coordina les plasticitats sinàptica i intrínseca. El fet que, en astròcits, l’activació de CREB també està regulada per activitat cerebral, situa aquest factor de transcripció com a la diana ideal per promoure canvis dependents d’activitat en astròcits. En aquesta tesi hem analitzat l’efecte de l’activació de la transcripció depenent de CREB en astròcits, centrant-nos en l’excitabilitat del calci i en el metabolisme d’aquestes cèl·lules. Hem demostrat que l’activació de la transcripció depenent de CREB redueix les senyals citosòliques de calci a través del mitocondri a la vegada que augmenta l’alliberació de lactat, dos canvis que poden tenir impacte en la transmissió sinàptica. Una altra contribució important d’aquest estudi es l’anàlisi molecular dels mitocondris dels astròcits, que ha revelat que aquestes cèl·lules poden utilitzar metabòlits que no són glucosa, com ara àcids grassos, per respondre a les necessitats metabòliques energètiques. Els nostres resultats estableixen el CREB en astròcits con un eix de la plasticitat astrocitària i revelen la interacció entre la plasticitat i el metabolisme energètic en astròcits. Aquests descobriments constitueixen un avenç mecanístic i conceptual en el coneixement de la biologia dels astròcits i com aquestes cèl·lules poden controlar l’aprenentatge i la memòria.<br>An increasing body of evidence suggests that astrocytes participate in higher-brain functions, controlling from synaptic transmission to global brain waves and learning and memory processes. Different mechanisms have been proposed to mediate these astrocyte-dependent processes, astrocytic lactate release and calcium-dependent gliotransmission being the main known effectors. The existence of control of brain functions by astrocytes suggests that astrocytes may shape brain functions in response to experience as much as neurons, thus constituting the phenomenon of astrocyte plasticity. In neurons, the transcription factor CREB is the best known coordinator of synaptic and intrinsic plasticity. The fact that, in astrocytes, CREB activation is also activity-dependent, positions CREB as an ideal target to promote plasticity-related changes in astrocytes, too. In this thesis, we have analyzed the effect of the activation of CREB-dependent transcription in astrocytes, specifically regarding calcium signals and metabolism. We have demonstrated that activation of CREB-dependent transcription reduces cytosolic calcium events via mitochondria and increases in lactate release, which may have impact on synaptic transmission. An important contribution of the study is the molecular analysis of astrocytic mitochondria, which has revealed that astrocytes may use fuels other than glucose such as fatty acids to meet basic energy metabolic demands. Taken together, our results establish astrocytic CREB as a hub in astrocyte-plasticity and shed light on the interplay between plasticity and energy metabolism in astrocytes; these findings constitute a conceptual and mechanistic advance in the knowledge of astrocytic biology and how these cells may control learning and memory.

[Truncated abstract] Arachidonic acid is oxidised in vivo by non-enzymatic (free radical) or enzymatic pathways (cyclooxygenase, lipoxygenase, and cytochrome P450) to form a range of biologically active eicosanoids. Specifically, arachidonic acid is metabolised by cytochrome P450 -hydroxylase to produce vasoactive 20-hydroxyeicosatetraenoic acid (20-HETE), and by 5-lipoxygenase to produce proinflammatory leukotriene B4 (LTB4), which can further be metabolised by -hydroxylase to from 20-OH-LTB4 and 20-COOH-LTB4. F2-Isoprostanes (F2-IsoPs) are produced through free radical attack on arachidonic acid and have been recognised as the most reliable markers of lipid peroxidation in vivo. The metabolic syndrome (MetS) is characterised by abdominal obesity, hypertension, insulin resistance, glucose intolerance, and dyslipidemia. It is associated with low-grade inflammation and oxidative stress and an increased risk of developing cardiovascular diseases. Dietary weight loss is strongly recommended for the management of the MetS and can potentially minimise the risk of cardiovascular diseases and diabetes in individuals with the MetS. Little is known regarding the role of these arachidonic acid metabolites in the MetS and the effect of weight loss on their metabolism. Chapter three comprised of three in vitro studies aimed to examine 20-HETE synthesis in human blood cells. 20-HETE acts as a second messenger for vasoconstrictor actions of angiotensin II (Ang II) and endothelin-1 (ET-1) in renal and mesenteric beds. Human neutrophils and platelets are integral to the inflammatory process. ... Production of LTB4 and 20-OH-LTB4 was significantly lower compared with controls (P<0.005) and remained so after adjustment for neutrophil count (P<0.05).The weight loss intervention resulted in a 4.6kg reduction in body weight and a 6.6cm decrease in waist circumference and a significant increase in LTB4 and 20-OH- LTB4 in the weight loss group. Chapter Five continued to investigate the role of other arachidonic acid metabolites, 20-HETE and F2-IsoPs in the MetS and the effect of weight loss. In the case-control study (Human study 1), plasma and urinary 20-HETE and F2-IsoPs were significantly elevated in the MetS group, but no significant difference was found in stimulated-neutrophil 20-HETE. A significant gender x group interaction was observed in that women with the MetS had higher urinary 20-HETE and F2-IsoPs compared to controls (P<0.0001). In a randomised controlled trial (Human study 2), relative to the weight- maintenance group, a 4.6 kg loss in weight resulted in a 2 mmHg fall in blood pressure but did not alter the production of 20-HETE or F2-IsoPs. No significant differences were shown in 20-HETE released from stimulated-neutrophils before and after weight loss. 20-HETE and oxidative stress may be important mediators of cardiovascular disease risk in the MetS. Although a 4% reduction in body weight reduced BP, there were no changes in plasma or urinary 20-HETE or F2-IsoPs. In summary, in vitro studies show that human neutrophils and platelets can produce 20-HETE in response to Ang II and ET-1, and human studies demonstrate that the presence of MetS has a significant impact on arachidonic acid metabolism and effective weight loss can restore leukocyte synthesis of LTB4.

Tetrahydrobiopterin is the cofactor for the hydroxylation of phenylalanine, tyrosine and tryptophan and is therefore essential for the production of monoamine neurotransmitters. Neopterin, a biosynthetic precusor of tetrahydrobiopterin, and biopterin appear in urine. In normal subjects the urinary neopterin to biopterin ratio has been found to be about 1.00. In patients suffering from Alzheimer's disease, Down's syndrome and depression the urinary neopterin to biopterin ratio has been found to be elevated. In some Alzheimer's and depressed patients the increased urinary neopterin to biopterin ratio is proportional to the severity of the disease. Folates were found not to increase tetrahydrobiopterin biosynthesis in the rat as previously thought. Methotrexate was found to reduce liver biopterin levels and increas_ urinary biopterin levels in the rat. Methotrexate also reduced brain pterin levels but had no influence on liver pterin. Urinary isoxanthopterin, found in some patients, was found to be derived from biopterin and neopterin in the rat. Isoxanthopterin is proposed as an indicator of the levels of tetrahydrobiopterin turnover.

La tesis busca contribuir a un mejor entendimiento de los complejos conflictos ecológico-distributivos en las fronteras de extracción, donde el aumento del metabolismo de las sociedades industriales conduce a una mayor destrucción del medio ambiente en países ricos en recursos naturales alrededor del mundo. Se desarrollan tres casos de estudios –Camisea, Conga y Sierra del Divisor, los cuales se explicarán más adelante. Se utiliza la teoría de flujos y fondos (Georgescu-Roegen, 1971) para desarrollar representaciones analíticas de los procesos económicos centrales que intervienen en cada caso de estudio, de forma tal que se facilita la comprensión de la anatomía de estos conflictos socio-ambientales. La tesis presenta una contribución metodológica empírica que combina dos enfoques: las triadas de valoración ambiental para representar el propósito económico (Farrell, 2007) y el análisis integrado multi-escala del metabolismo de la sociedad y los ecosistemas (MuSIASEM, en inglés) para representar el proceso económico (Giampietro y Mayumi, 2000a, 2000b, 2009). En este sentido, el propósito económico de un actor social define los límites –frontera y duración– del proceso económico, y por lo tanto, las identidades de los elementos del proceso en términos de flujos y fondos. Esto puede ser entendido como la etapa pre-analítica de la representación MuSIASEM del proceso económico en cuestión. El caso de estudio de Camisea analiza el complejo de energía-agua-minería y plantea la interrogante sobre cuáles son las implicancias de largo plazo en el sistema energético nacional del crecimiento del sector minero en Perú, favorecido por el gobierno. Esta pregunta se responde mediante el análisis de las interacciones entre el fondo de actividad humana y los flujos de energía exosomática a través de múltiples escalas de la economía peruana para los años 2000 y 2010, con una proyección al 2020. Los resultados empíricos indican: (1) el altísimo ratio metabólico de electricidad (eEMR) del sector minero (61.6 MJ/h en 2010), el cual resultó ser 11 veces el eEMR del sector de construcción y manufactura; y (2) el posible incremento de la proporción de electricidad utilizada por el sector minero, lo cual podría reducir la disponibilidad de gas natural de Camisea –la principal reserva de combustibles fósiles– para el resto de la sociedad peruana. Por lo tanto, se argumenta que el fuerte apoyo del gobierno al crecimiento del sector minero tendría que ser revisado. El caso de estudio de Conga en los Andes explora la anatomía del conflicto ecológico distributivo entre la empresa minera y los campesinos. Se complementa el concepto de tierra Ricardiana –un fondo indestructible– con el concepto de materiales de tierra, el cual es susceptible de cambio cualitativo, y por lo tanto puede ser fondo o flujo del proceso económico. Desde la perspectiva de la empresa minera, los materiales de tierra son considerados flujos del proceso de extracción de minerales, lo cual está en conflicto con la perspectiva de los campesinos, quienes consideran los mismos materiales de tierra como fondos del proceso de producción de leche fresca. En otras palabras, desde el punto de vista de los campesinos “el agua vale más que el oro”, es decir, el oro como valor de cambio en el mercado es menos importante que el agua como valor de soporte de la vida, tanto biofísica como espiritual. El caso de estudio de Sierra del Divisor en la Amazonia aplica una representación simplificada de MuSIASEM con el fin de describir los procesos económicos clave en dos períodos de tiempo, antes y después de la posible construcción del proyecto de ferrocarril transcontinental entre Brasil y Perú, que podría cruzar la Sierra del Divisor y los territorios Isconahuas. Los procesos económicos estudiados incluyen: la producción industrial de soya en Brasil, la minería de oro aluvial en Perú, las pesca de comunidades nativas, la producción de arroz de pequeños agricultores, y las actividades de caza y recolección de pueblos indígenas viviendo en aislamiento voluntario.<br>The thesis aims to contribute toward improved understanding of complex ecological distribution conflicts at the commodity frontiers, where increasing metabolism in industrial societies is leading to increased environmental destruction in resource rich countries throughout the world. The thesis consists of three case studies of social metabolism in Peru (i.e. Camisea, Conga, and Sierra del Divisor). Analytical representations of the central economic processes are developed based on the flow/fund theory (Georgescu-Roegen, 1971) in order to explore the anatomy of these environmental conflicts. The thesis develops an empirical methodological contribution that combines two approaches: the environmental valuation triadics representation of economic purpose (Farrell, 2007) and the multi-scale integrated analysis of societal and ecosystem metabolism (MuSIASEM) representation of economic process (Giampietro and Mayumi, 2000a, 2000b, 2009). That is, the social actor’s economic purpose defines the boundaries of the economic process (i.e. frontier and duration), and therefore, the process elements identities in terms of flows and funds. This can be understood as the pre-analytical step of the MuSIASEM representation of an economic process. The Camisea case study analyzes the energy-water-mining complex, and poses the specific question: What are the long-term national energy system implications of the government-supported growth of the mining sector? This question is addressed by analyzing interactions between funds of human economic activity and flows of exosomatic energy across scales of the Peruvian economy, in 2000 and 2010, with a projection for 2020. The MuSIASEM empirical results indicate: (1) the extremely high electricity metabolic rate (eEMR) of the mining sector (61.6 MJ/h in 2010), which was found to be 11 times the eEMR of the building and manufacturing sector; and (2) the potential increase of the flow share of electricity used by the mining sector, which could reduce the availability of Camisea natural gas –the main fossil fuels reserve– for the rest of society. Based on these implications, it is argued that the Peruvian government strong support for growth of the mining sector may have to be reconsidered. The Conga case study in the Andes explores the anatomy of the ecological distribution conflict between the mining corporation and the campesinos (peasants). By complementing the concept of Ricardian land—an indestructible fund—with the concept of land materials, which is susceptible to qualitative change, and therefore can be either a fund or a flow element of the economic process, we illustrate that the minerals extraction process of multinational companies, which treats this land material as a flow, stands in conflict with the milk production process of campesinos, because that process is using these land materials as a fund, that is, in order to make production possible. In other words, from the perspective of campesinos (and the common sense) “el agua vale más que el oro” –that is, the market exchange value of gold is less important than the life support value of water (i.e. biophysical and spiritual). The Sierra del Divisor case study in the Amazonia applies the MuSIASEM in a simplified way in order to describe key economic processes in two periods of time, before and after the potential construction of the transcontinental Brazil-Peru railway project that would cross the Sierra del Divisor tropical rainforest, representing a major change in the boundary conditions of the observed system. The economic processes studied in this case include: industrial soybean production in Brazil, alluvial gold mining extraction in Peru, fishing by native communities, rice production by small farmers, and hunting and gathering activities of indigenous people living in voluntary isolation.

Deficiências nutricioniais são freqüentemente associadas a aumento das taxas de morbidade e mortalidade no pós-operatório. Descreve-se há mais de 50 anos a relação existente entre perda de peso corporal pré-operatório e o aumento da mortalidade no pós-operatório. Vários fatores, além da desnutrição em si, contribuem para este quadro, entre eles a própria doença de base, a cirurgia, infecções associadas, dentre outros. Sabidamente, na presença de trauma físico e ou metabólico, são observadas alterações no metabolismo das proteínas e dos aminoácidos, além da liberação e aumento de hormônios catabolizantes. O presente trabalho objetiva avaliar se a infusão endovenosa de aminoácidos e glicose, em via periférica, em pacientes submetidos ao estresse cirúrgico, altera esse estresse e/ou resulta em retenção de nitrogênio aminoacídico. Foram estudados 18 pacientes, triados e operados pela mesma equipe médica captados do ambulatório do Serviço de Cirurgia Geral do Hospital Universitário da Universidade Federal de Alagoas. Após serem avaliados e, consentida a realização do protocolo de estudo, foram submetidos à avaliação nutricional e, aleatoriamente, divididos em dois grupos. O grupo I, ou controle, durante o período transoperatório, recebeu solução de ringer com lactato e glicose 5%, como rotineiramente utilizado pelo Serviço de Anestesia e Cirurgia da Universidade Federal de Alagoas. O grupo II, ou teste, além da infusão de soro ringer com lactato e glicose 5%, recebeu aminoácidos 6,6% e glicose 16,6%, equivalente, respectivamente, a 50 gramas de proteína e 250 g de glicose. Observou-se, nos resultados, que não houve diferença estatística entre os grupos estudados, no que se refere à sexo, faixa etária, exames bioquímicos, aminoácidos e concentração urinária de adrenalina no pré e pós-operatório. Observou-se, ainda, que o grupo II, que recebeu solução de aminoácidos e glicose, não excretou mais nitrogênio nos períodos pré, trans e pós-operatório, em comparação ao grupo I, mantendo os valores semelhantes ao grupo I após 24 horas do início da anestesia. No entanto, o grupo II, embora tenha excretado quantidades semelhantes de nitrogênio em comparação ao grupo I, teve um balanço nitrogenado, em média, menos negativo que o grupo I (p<0,05), retendo aproximadamente 40% a mais de nitrogênio aminoacídico. Conclui-se, desta forma, que a infusão parenteral de nitrogênio aminoacídico 6,6% e glicose 16,6% no transoperatório pode ser benéfico aos pacientes sob estresse cirúrgico.<br>It's been described for almost 50 years that weight loss at the preoperative period increases the postoperative mortality. Many factors, besides the malnutrition itself, contribute to this situation, for example, the patient illness, the surgical procedure, associated infections and others. It’s also known that in trauma, organic or not, there are disorders in proteic and aminoacid metabolism and an increase on release of catabolic hormones. This study has the objective to evaluate if the endovenous infusion of a amino acid and glucose solution, in a peripheric vein, in patients submitted to a surgical procedure, modifies the patients response to the trauma and/or the nitrogen retention. We studied 18 patients from the Surgery Division of the Clinical Hospital of the Federal University of Alagoas. The same medical team followed all patients. After evaluation and consent of the realization of the study, they went on an assessment of the nutritional status and were randomized in 2 groups. The group I (control group), during the perioperative period received a solution of lactate ringer and glucose 5%, as is usually done at this hospital. The group II (test group), besides the referred solution, received a mixture of amino acids 6,6% and glucose 16,6%, meaning 50g of protein and 250g of glucose during the transoperatory period. There was no statistic difference between the groups studied in what concerns to age, sex, biochemistry tests, aminoacids and adrenalin urinary concentration at the pre or postoperative periods. It has also been observed that group II, that received a solution with aminoacid and glucose, hasn’t excreted more nitrogen through pre, trans and postoperative periods in comparation to group I, maintaining similar values of group I 24 hours after the anesthesic procedure. However, though group II had excreted similar values of nitrogen in comparation to group I, it had a nitrogened balance less negative than group I (p<0,05) retaining approximately 40% more aminoacidic nitrogen. From that, we conclude that the parenteral infusion of aminoacidic nitrogen 6,6% and glucose 16,6% at the transoperative period may be a benefit to patients under surgical stress.

Les lipins són una família conservada evolutivament de fosfatases de fosfatidat (PAP1) dependents de Mg2+, que generen diacilglicerol per a la síntesi de fosfolípids i triacilglicerol. En mamífers, la família consta de lipina-1, lipina-2 i lipina-3. Mentre en ratolins la mutació del gen Lpin1 causa lipodistròfia, les mutacions deletèries en el gen LPIN1 en humans no afecten la distribució del greix. No obstant, persones amb diabetis tipus 2 mostren nivells reduïts de l'expressió de LPIN1 i de l'activitat PAP1. Aquesta tesi estudia el paper de les lipins en el teixit adipós humà, la adipogènesi i la lipòlisi. Descobrim que la expressió de gens i proteïnes lipin és alterada en el teixit adipós de les persones amb diabetis tipus 2. Silenciant cada membre de la família lipin en la línia cel•lular humana de preadipòcits del síndrome Simpson-Golabi-Behmel (SGBS), mostrem que mentre que els tres membres tenen un paper en el primers estadis de l’adipogènesi, els preadipòcits silenciats de lipin es diferencien i acumulen lípids neutres, la qual cosa condueix a la hipòtesi de l'existència de vies alternatives per a la síntesi de triacilglicerol en adipòcits humans quan es reprimeix l'expressió de les lipin. Les lipin participen també en el reciclatge d'àcids grassos alliberats mitjançant la via lipolítica. Després de la inducció de la lipòlisi, les lipines són defosforilades i es desplacen a la membrana del reticle endoplasmàtic, on exerceixen la seva funció enzimàtica. Aquesta activació és induïda pels àcids grassos alliberats i s'inverteix amb la presència d’albúmina o triacsin C. La inducció d’adipòcits silenciats de cada lipina demostra el seu paper en el metabolisme dels lípids neutres. En resum, les lipin semblen no tenir un paper imprescindible en la adipogènesi humana però sí poden comprometre el reciclatge d'àcids grassos, important per a la homeòstasis lipídica.<br>Las lipinas son una familia de fosfatasas de fosfatidato (PAP1) dependientes de Mg2+ evolutivamente conservadas, que generan diacilglicerol para la síntesis de fosfolípidos y triacilglicerol. En mamíferos, la familia consiste en lipina-1, lipina-2, y lipina-3. Mientras en ratones la mutación del gen Lpin1 causa lipodistrofia, las mutaciones deletéreas en el gen LPIN1 en humanos no afectan a la distribución de grasa. Sin embargo, los individuos con diabetes tipo 2 manifiestan niveles reducidos de expresión de LPIN1 y de actividad PAP1. En esta tesis doctoral se estudia la función de las lipinas en el tejido adiposo humano, la adipogénesis y la lipólisis. Descubrimos que la expresión génica y proteica de las lipinas está alterada en el tejido adiposo de individuos con diabetes tipo 2. La depleción de cada miembro de las lipinas en la línea celular humana de preadipocitos del síndrome Simpson–Golabi–Behmel (SGBS), mostró que, a pesar de que los tres miembros tienen un papel en la adipogénesis temprana, los adipocitos deplecionados de lipinas se diferencian y acumulan lípidos neutros, llevándonos a la hipótesis de la existencia de vías alternativas para la síntesis de triacilglicerol en adipocitos humanos cuando la expresión de las lipinas es reprimida. Las lipinas también intervienen en el reciclaje de los ácidos grasos liberados por la vía lipolítica. Tras la inducción de la lipólisis, las lipinas son defosforiladas y se desplazan a la membrana del retículo endoplásmico, donde ejercen su función. Esta activación es inducida por los ácidos grasos liberados, y revertida con albúmina o triacsin C. La depleción de cada lipina en adipocitos SGBS y posterior inducción de la lipólisis, demuestra su papel en el metabolismo de lípidos neutros. En resumen, las lipinas parecen no tener un papel indispensable en la adipogénesis humana pero sí comprometer el reciclaje de ácidos grasos, importante para la homeostasis lipídica.<br>Lipins are evolutionarily conserved Mg2+-dependent phosphatidate phosphatases (PAP1) that generate diacylglycerol for phospholipid and triacylglycerol synthesis. In mammals the Lipin family consists of lipin-1, lipin-2 and lipin-3. Whereas mutations in the Lpin1 gene cause lipodystrophy in mouse models, LPIN1 deleterious mutations in humans do not affect fat distribution. However, reduced LPIN1 expression and PAP1 activity have been described in participants with type 2 diabetes. In this doctoral thesis we investigate the roles of all lipin family members in human adipose tissue, adipogenesis and lipolysis. We found that adipose tissue gene and protein expression of the lipin family is altered in type 2 diabetes. Depletion of every lipin family member in a human Simpson–Golabi–Behmel syndrome (SGBS) pre-adipocyte cell line showed that even though all members alter early stages of adipogenesis, lipin-silenced cells differentiate and accumulate neutral lipids, pointing to the hypothesis of alternative pathways for triacylglycerol synthesis under repression of lipin expression. Lipins also have a role in the recycling of the fatty acids released by the lipolytic pathway. They become dephosphorylated upon lipolytic induction, and translocate to their active site, the endoplasmic reticulum membrane. This activation is induced by fatty acids and reversed with albumin or triacsin C. Depletion of every lipin member and subsequently stimulation of lipolysis in SGBS adipocytes revealed a role for lipins in neutral lipid metabolism. Overall, our data support that lipins may not have an indispensable role in adipogenesis, but their depletion compromise fatty acid recycling and lipid homeostasis.

A relação entre transferência de lipídeos, idade e aterogênese são complexas e ainda não estão claras. É possível que a troca de lipídeos esteja alterada com a avançar da idade e relacionada com a Doença Arterial Coronariana (DAC). O objetivo deste trabalho foi verificar a hipótese se em indivíduos mais jovens a habilidade da HDL de receber lipídeos é diferente de indivíduos mais velhos com e sem a evidência clínica da DAC. Dentro desses aspectos, foram determinados o diâmetro da partícula desta lipoproteína, a atividade da Paraoxonase (PON1) e sua capacidade de receber lipídeos. Para tanto, foram estudados 86 indivíduos divididos em quatro grupos: adulto jovem (25±4), meia-idade (42±8), idosos sem evidência clínica de DAC (75±6) e idosos com DAC (74±5). Uma nanoemulsão artificial rica em colesterol (LDE) marcada com 3H-TG e 14C-CL ou 3H-CE e 14C-FL foi incubada com plasma. Após a precipitação de outras lipoproteínas, o sobrenadante contendo HDL foi separado e em seguida, medida a radioatividade. O diâmetro da HDL foi medido por laser scattering (nm). Foram constatadas diferenças significativas entre as taxas de transferência de 3H-éster de colesterol (CE) entre os grupos: adulto jovem (3.7±1.0%); meia idade (4.1 ±0.7%) e idosos (5.3±1.8%);p= 0.024. Também ocorreu diferença entre as taxas de transferência do 14C-fosfolipídeo (FL): adulto jovem (18.7±4.6%), meia idade (18.3 ±4.0%) e idosos (20.6±5.3); p=0.0368. Com relação ao tamanho das partículas de HDL, foi encontrada diferença entre os grupos: os grupos adulto jovem (8.9± 0.3nm) e meia idade (8.9± 0.3nm) apresentaram menores diâmetros de HDL quando comparados ao do grupo de idosos sem evidência clínica da DAC (9.7± 1.6);p= 0,0444. As transferências de lipídeos foram expressas em % de radioatividade. A idade correlacionou-se positivamente com a taxa de transferência do 3H- éster de colesterol (r=0.3365; p=0.0036), com a concentração de colesterol total (r=0.4965; p=0.0001) e com a concentração de HDL colesterol (r=0.3559; p=0.0023). Também houve correlação positiva com o tamanho de HDL (r=0.3695; p=0.0013). Em princípio, os indivíduos idosos sem evidência clínica da DAC, aparentemente têm alguma proteção contra a mesma. Desse modo, com o intuito de saber se os resultados encontrados no presente trabalho sustentam a afirmação acima, foi realizada a comparação desse grupo com um grupo de idosos que apresentavam a DAC. O grupo com DAC apresentou menor tamanho de partícula de HDL (8,7±0,7). As taxas de transferência de 3H-CE e de 14C-FL também foram menores neste grupo (3H-CE=3,1 ±2,3 e 3H-TG= 5,1 ±1 ,6). Devido ao importante papel antiaterogênico da HDL, esses resultados podem ser relevantes para estabelecer novos mecanismos existentes entre os aspectos qualitativos dessa lipoproteína, o avanço da idade e a presença da DAC.<br>The relationship between transfer of lipids, age and atherogenesis are complex and yet unclear and is possible that the shift of lipids to HDL may be altered by the aging process and related with coronary artery disease (CAD). We tested the hypothesis whether in younger patients the ability of HDL to receive lipids is different from that of elderly patients with or without CAD. Inside of these aspects, the HDL size, the activity of Paraoxonase (PON1) and its capacity to receive lipids was determined. It was studied, 25 younger, 25 middle age, 36 elderly patients with a coronariography and/or a perfusion scintilography on the last 6 months (11 with CAD, 74±5 yo; and 25 patients without proved CAD, 75±6 yo). An artificial cholesterol-rich nanoemulsion labeled with 3H-TG and 14C-FC or H-CE and 14C-PL was incubated, per 1 hour, with plasma. After chemical precipitation of apoB-containing lipoproteins and the nanoemulsion, the supernatant containing HDL was counted for radioactivity. The HDL diameter was measured by laser-light-scattering. Transfer of CE and PL to HDL was smaller in young patients than in the elderly patients without CAD, but the transfer of the other lipids are equal (CE: young= 3.7±1.0%; middle age= 4.1 ±0.7%; elderly without CAD= 5.3±1.8%; p= 0.024 and PL: young= 18.7±4.6%; middle age= 18.3 ±4.0%; elderly without CAD= 20.6±5.3; p=0.0368). The HDL size was greater in elderly group without CAD (9.7± 1.6nm) than in younger (8.9± 0.3nm) and middle age patients (8.9± 0.3nm); p=0,0444. Transfer of lipids to HDL was expressed as % of total incubated radioactivity. The age positively correlated with the transfer of CE (r=0.3365; p=0.0036), with the total cholesterol concentration (r=0.4965; p=0.0001) and with the HDL concentration cholesterol (r=0.3559; p=0.0023). Also had positive correlation with the size of HDL (; p=0.0013). In principle, the aged patients without CAD, have some protection against the same one. In this aspect, with intention to know if the results found in the present work support the affirmation above, was compared this group with a group of aged that presented the CAD. Comparing elderly patients without CAD with elderly patients with CAD, the transfer of CE and FL as well as HDL size was smaller in the CAD group (CE=3.1±2.3 and TG= 5.1±1.6; 8.7±0.7nm). Due to HDL important antiatherogenic roles, this result can be relevant to establish new mechanisms and risk factors in aging and in CAD.

Background: The metabolic syndrome is associated with an increased risk of diabetes and vascular disease. In order to understand the pathophysiological processes underlying such risk, it is necessary to develop a better understanding of normal fat metabolism and abnormalities associated with the syndrome. The hypothesis tested in this thesis is that specific abnormalities in adipose tissue and muscle fat metabolism characterise the metabolic syndrome. Methods: Fasting biochemical parameters were measured in a cohort of overweight men with and without the metabolic syndrome. Stable-isotope labeling and arterio-venous difference measurements were conducted in 18 men to elucidate pathways of exogenous and endogenous fat metabolism under fasting and postprandial conditions in adipose tissue and skeletal muscle. In addition, a pilot study of the effects of heat and electrical stimulation on adipose tissue metabolism was undertaken. Results: Cohort study - The prevalence of the metabolic syndrome depended on the definition used. Total cholesterol and apoB were greater in those with the metabolic syndrome than in those without. There was no difference in fasting NEFAs. Metabolic investigation - There was significant postprandial uptake of NEFA from the circulating NEFA pool by adipose tissue. Chylomicrons were confirmed as the preferred substrate of LPL. There was preferential uptake of FAs derived from chylomicron hydrolysis. There was release of NEFA across muscle. In the metabolic syndrome, adipose tissue NEFA output is lower during fasting and falls less following a meal than in the healthy obese. Clearance of dietary-derived TG is lower across both adipose tissue and muscle in the metabolic syndrome. Pilot study – Heat increased measures of lipolysis whereas electrical stimulation had no effect. Conclusions: Fat metabolism in individuals with the metabolic syndrome is characterised by metabolic inflexibility but not insulin resistance.

Made available in DSpace on 2014-06-11T19:23:01Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-01-22Bitstream added on 2014-06-13T19:28:48Z : No. of bitstreams: 1 cristofalo_r_me_botib.pdf: 456212 bytes, checksum: 2db6199317eddbdd0f0bd4bf0b636d1a (MD5)<br>Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)<br>O aumento da atividade dos radicais livres e da produção de citocinas pró-inflamatórias por monócitos pode estar envolvido na patogênese da pré-eclâmpsia. A silibinina é o componente mais ativo da silimarina (Silybum marianum), um flavonóide polifenólico que possui efeitos anti-oxidante e anti-inflamatório. O objetivo do presente estudo foi avaliar o efeito da silibinina sobre a produção de citocinas pró e anti-inflamatórias, bem como sobre a liberação de ânion superóxido (O2 -) por monócitos de gestantes com pré-eclâmpsia.Foram selecionadas 30 gestantes com préeclâmpsia (PE), 30 gestantes normais (GN) e 30 mulheres normais nãográvidas (NG). Monócitos de sangue periférico foram incubados na presença ou ausência de silibinina nas concentrações de 5 e 50 g/mL por 60min e, estimulados ou não com ester de forbol (PMA) para determinação de O2 -. Estes monócitos também foram estimulados ou não com lipopolissacáride (LPS) por 18h e, no sobrenadante das culturas foram determinadas, pela técnica de ELISA, as seguintes citocinas: fator de necrose tumoral-alfa (TNF- ), Interleucinas (IL-) IL-6, IL-10, IL-12, IL-15, fator estimulador de colônias de granulócitos e monócitos (GM-CSF) e fator de crescimento e transformação (TGF- . A atividade da enzima superóxido dismutase (SOD) foi avaliada em hemolisado de eritrócitos dos três grupos estudados.Níveis significativamente maiores de O2 - foram liberados por monócitos de gestantes com PE quando comparados com GN e NG, confirmando o estado ativado dessas células. A atividade da SOD foi significativamente maior no grupo de gestantes com PE. Os níveis endógenos de TNFforam significativamente mais elevados nas pacientes com PE, quando comparados com os grupos GN e NG, enquanto os níveis de IL-10 foram significativamente menores nas gestantes com PE. A capacidade de produção de citocinas pelos...<br>Increased free radical activity and high proinflammatory cytokine production by monocytes may be implicated in the pathogenesis of preeclampsia. Silibinin is a major active component of silymarin (Silybum marianum), a polyphenolic plant flavonoid that has antioxidant and anti-inflammatory effects. This study investigated the in vitro effect of silibinin on monocyte production of pro- and anti-inflammatory cytokines, as well as on superoxide anion (O2-) release in pregnant women with preeclampsia.Thirty preeclamptic (PE), 30 healthy pregnant (HP) and 30 healthy non-pregnant (NP) women were included. Peripheral blood monocytes were incubated with or without silibinin at 5 and 50ug/mL for 60 min, and stimulated with or without phorbol myristate acetate (PMA) for the assessment of O2-. These cells were also cultured with or without lipopolysaccharide (LPS) for 18h and tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-6, IL-10, IL-12p40, IL-15, granulocyte-macrophage colony-stimulating factor (GM-CSF) and transforming growth factor-beta1 (TGF- 1) in monocyte culture supernatants were determined by ELISA. The activity of the enzyme superoxide dismutase (SOD) was evaluated in erythrocyte lisates of the three groups studied.Monocytes of preeclamptic patients release significantly higher levels of O2 - in comparison to HP and NP women confirming the activation state of these cells. SOD activity in erythrocytes was significantly higher in preeclamptic patients. The endogenous levels of TNFwere significantly higher in PE patients than in HP and NP groups, while IL-10 production was significantly lower in PE women. The levels of IL-6, IL-12 and GM-CSF spontaneously produced by monocytes were higher in HP and PE groups than in NP women. The capacity of cytokine production by LPS-stimulated monocytes was preserved in all the three groups studied... (Complete abstract click electronic access below)

Made available in DSpace on 2014-12-02T11:16:38Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-04-01Bitstream added on 2014-12-02T11:21:05Z : No. of bitstreams: 1 000777381_20151101.pdf: 335580 bytes, checksum: 320f77e5c5865613fa8703a80223014c (MD5) Bitstreams deleted on 2015-11-03T14:39:13Z: 000777381_20151101.pdf,. Added 1 bitstream(s) on 2015-11-03T14:40:15Z : No. of bitstreams: 1 000777381.pdf: 3018222 bytes, checksum: d974504f28696fef1888197540e512b4 (MD5)<br>Cefamicina C é um antibiótico produzido por Streptomyces clavuligerus de grande interesse, em virtude de seu maior grau de resistência a enzimas do tipo β-lactamases comparativamente a outros antibióticos beta-lactâmicos. Cefamcina C é produzida em pequenas concentrações na natureza assim como acontece com todos os metabólitos secundários. Assim, investimentos em programas de melhoria de linhagens e de otimização de meios de cultura e operação de biorreatores são aspectos-chave para o aumento da produção industrial. Porém, a eficácia de tais estratégias pode ser limitada, o que requer o uso de novas abordagens. Neste sentido, a engenharia metabólica é uma área importante que alia ferramentas de quantificação de fluxos metabólicos e de técnicas de biologia molecular para melhoria de linhagens. O estudo dos fluxos metabólicos permite, por meio de análise criteriosa do metabolismo do micro-organismo, identificar gargalos metabólicos na rota biossintética de um produto de interesse para, posteriormente, propor possíveis soluções para o aumento da produção. No presente projeto realizou-se o estudo dos fluxos metabólicos de S. clavuligerus com vistas a encontrar formas de operação do metabolismo que conduzam a aumentos da produção de cefamicina C. Para isto, primeiramente foi desenvolvido um meio de cultivo quimicamente definido contendo maltose como fonte de carbono e lisina como fonte de nitrogênio, para que análises do caldo fermentado não tivessem interferência de componentes desconhecidos. Tal meio possibilitou a obtenção de biomassa em torno de 9 g.L-1 e cefamicina C ao redor de 200 mg.L-1 em processo contínuo. Este modo de operação foi possível somente em biorreator devido ao controle de pH, pois em shaker as variações deste parâmetro inviabilizaram o processo. Paralelamente, foi construído um modelo metabólico com 78 reações e 81 metabólitos, sendo 10 externos e 71 internos, que ...<br>Cephamycin C is an antibiotic produced by Streptomyces clavuligerus of great interest due to its higher degree of resistance to β-lactamases as compared to other beta- lactam antibiotics. Cephamycin C is produced in small concentrations in nature as with all secondary metabolites. Therefore, investments in improvement of strains and optimization, of culture media and operation of bioreactors are key aspects to increase the production. However, the effectiveness of such strategies may be limited, requiring the use of new approaches. In this aspect, the metabolic engineering is an important field that combines quantification of metabolic fluxes and molecular techniques for improving strains. The study of metabolic fluxes enables one to identify metabolic bottlenecks through careful analysis of metabolism of the microorganism, and to suggest ways to increase production. In this project we carried out the study of metabolic fluxes in S. clavuligerus aiming to find ways to increase the production of cephamycin C. For this, first we developed a chemically defined culture medium because this kind of media does not cause interference in the analyses. The developed medium contained maltose as carbon source and lysine as nitrogen source and resulted in 9 g.L-1 of biomass and 200 mg.L-1 of cephamycin C in the continuous process. This mode of operation was only possible in the bioreactor due to its pH control. Due to variations in the pH, the continuous process in shaken-flasks became unviable. The metabolic model was constructed with 78 reactions and 81 metabolites (10 external and 71 internal) which suitably described the metabolism of S. clavuligerus. This model was simulated with the aid of Optflux, a multi-task software developed for this purpose. The profiles of maltose, lysine, biomass, cephamycin C, clavulanic acid, external protein and CO2 evolution were monitored. These data were used for the model simulations. The results allowed...

Orientador:Maria Lucia Gonsales da Costa Araujo<br>Banca: José Roberto Ernandes<br>Banca: Ruy de Souza Junior<br>Banca: Alberto Colli Badino Junior<br>Banca: José Gregório Cabrera Gomez<br>Resumo: Cefamicina C é um antibiótico produzido por Streptomyces clavuligerus de grande interesse, em virtude de seu maior grau de resistência a enzimas do tipo β-lactamases comparativamente a outros antibióticos beta-lactâmicos. Cefamcina C é produzida em pequenas concentrações na natureza assim como acontece com todos os metabólitos secundários. Assim, investimentos em programas de melhoria de linhagens e de otimização de meios de cultura e operação de biorreatores são aspectos-chave para o aumento da produção industrial. Porém, a eficácia de tais estratégias pode ser limitada, o que requer o uso de novas abordagens. Neste sentido, a engenharia metabólica é uma área importante que alia ferramentas de quantificação de fluxos metabólicos e de técnicas de biologia molecular para melhoria de linhagens. O estudo dos fluxos metabólicos permite, por meio de análise criteriosa do metabolismo do micro-organismo, identificar gargalos metabólicos na rota biossintética de um produto de interesse para, posteriormente, propor possíveis soluções para o aumento da produção. No presente projeto realizou-se o estudo dos fluxos metabólicos de S. clavuligerus com vistas a encontrar formas de operação do metabolismo que conduzam a aumentos da produção de cefamicina C. Para isto, primeiramente foi desenvolvido um meio de cultivo quimicamente definido contendo maltose como fonte de carbono e lisina como fonte de nitrogênio, para que análises do caldo fermentado não tivessem interferência de componentes desconhecidos. Tal meio possibilitou a obtenção de biomassa em torno de 9 g.L-1 e cefamicina C ao redor de 200 mg.L-1 em processo contínuo. Este modo de operação foi possível somente em biorreator devido ao controle de pH, pois em shaker as variações deste parâmetro inviabilizaram o processo. Paralelamente, foi construído um modelo metabólico com 78 reações e 81 metabólitos, sendo 10 externos e 71 internos, que ...<br>Abstract: Cephamycin C is an antibiotic produced by Streptomyces clavuligerus of great interest due to its higher degree of resistance to β-lactamases as compared to other beta- lactam antibiotics. Cephamycin C is produced in small concentrations in nature as with all secondary metabolites. Therefore, investments in improvement of strains and optimization, of culture media and operation of bioreactors are key aspects to increase the production. However, the effectiveness of such strategies may be limited, requiring the use of new approaches. In this aspect, the metabolic engineering is an important field that combines quantification of metabolic fluxes and molecular techniques for improving strains. The study of metabolic fluxes enables one to identify metabolic bottlenecks through careful analysis of metabolism of the microorganism, and to suggest ways to increase production. In this project we carried out the study of metabolic fluxes in S. clavuligerus aiming to find ways to increase the production of cephamycin C. For this, first we developed a chemically defined culture medium because this kind of media does not cause interference in the analyses. The developed medium contained maltose as carbon source and lysine as nitrogen source and resulted in 9 g.L-1 of biomass and 200 mg.L-1 of cephamycin C in the continuous process. This mode of operation was only possible in the bioreactor due to its pH control. Due to variations in the pH, the continuous process in shaken-flasks became unviable. The metabolic model was constructed with 78 reactions and 81 metabolites (10 external and 71 internal) which suitably described the metabolism of S. clavuligerus. This model was simulated with the aid of Optflux, a multi-task software developed for this purpose. The profiles of maltose, lysine, biomass, cephamycin C, clavulanic acid, external protein and CO2 evolution were monitored. These data were used for the model simulations. The results allowed...<br>Doutor

Thesis (Ph. D.)--University of North Carolina at Chapel Hill, 2006.<br>Title from electronic title page (viewed Oct. 10, 2007). "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Environmental Sciences and Engineering, School of Public Health." Discipline: Environmental Sciences and Engineering; Department/School: Public Health.

Submitted by Erika Demachki (erikademachki@gmail.com) on 2014-12-29T15:35:49Z No. of bitstreams: 2 license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Dissertação - Letícia de Almeida Leandro - 2013.pdf: 2111763 bytes, checksum: c992b02afb4a59e318265d06577e6454 (MD5)<br>Approved for entry into archive by Erika Demachki (erikademachki@gmail.com) on 2014-12-29T15:40:58Z (GMT) No. of bitstreams: 2 license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Dissertação - Letícia de Almeida Leandro - 2013.pdf: 2111763 bytes, checksum: c992b02afb4a59e318265d06577e6454 (MD5)<br>Made available in DSpace on 2014-12-29T15:40:58Z (GMT). No. of bitstreams: 2 license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Dissertação - Letícia de Almeida Leandro - 2013.pdf: 2111763 bytes, checksum: c992b02afb4a59e318265d06577e6454 (MD5) Previous issue date: 2013-02-28<br>The analysis of the energetic and respiratory metabolism of Taenia crassiceps cysticerci innoculated into the encefalum of female BALB/c mice was performed. After 30 days of infection the mice were treated with low dosages (3.0 and 6.0 mg/kg) of albendazole and praziquantel aiming the observation of the alterations caused by a hostile environment on the parasite's metabolism. The aim of this study was to detect the influence of low dosages of anti-helminthic drugs on the production of organic acids related to the intermediary metabolism (oxaloacetate, malate, fumarate, succinate, citrate and α-ketoglutarate), carbohydrates metabolism (pyruvate, lactate, propionate, oxaloacetate and malate) and fatty acids and proteins catabolism (β-hydroxibutyrate, acetoacetate, acetate, α-ketoglutarate and oxalate) by T. crassiceps cysticerci innoculated into the CNS of BALB/c mice. Regarding the glycolisis, it was possible to detect lactate in all samples which shows that this parasite performs a lactic fermentation. As to the intermediary metabolism it was possible to detect oxaloacetate, citrate, malate, succinate, fumarate and α-ketoglutarate which indicates that the cysticerci may have used the tricarboxilic acid cycle (TCA) to produce energy. However facing the results found the evidences show that the cysticerci used a metabolic pathway denominated reversion of the TCA for energy production which presents as main end products propionate and acetate. There were no significant differences between the groups treated with the anti-helminthic and the control one except that the group treated with 6.0 mg/kg of albendazole presented acetate as main end product of the TCA reversion while the other groups presented propionate. In the groups treated with praziquantel, both concentrations, there was no production of acetate and in the 6.0 mg/kg of praziquntel there was no production of β-hydroxibutyrate wheter in the groups treated with both concentrations of albendazol there was an increase in the production of this organic acid which indicates the maximization of the energetic pathway of fatty acids oxidation and/or of the proteic catabolism. It was possible to conclude that T. crassiceps cysticerci when exposed to the conditions of this study preferred an anaerobic energy production pathway and probably performed the catabolism of carbohydrates, proteins and fatty acids.<br>Realizou-se a análise do metabolismo energético e respiratório de cisticercos de Taenia crassiceps inoculados no encéfalo de camundongos BALB/c fêmeas. Após 30 dias de infecção, esses camundongos foram tratados com baixas doses (3,0 e 6,0 mg/kg) de albendazol e praziquantel visando observação de alterações provocadas no metabolismo do parasito. O objetivo deste trabalho foi detectar a influência de baixas doses desses anti-helmínticos na produção dos ácidos orgânicos do metabolismo intermediário (citrato, α-cetoglutarato, succinato, fumarato, malato, oxaloacetato), de carboidratos (piruvato, lactato, oxaloacetato, malato, acetato e propionato), de ácidos graxos e proteínas (β-hidroxibutirato, acetoacetato, acetato, α-cetoglutarato e oxalato) realizados por cisticercos de T. crassiceps implantados no SNC de camundongos BALB/c fêmeas. Com relação a glicólise, detectou-se lactato em todas as amostras mostrando que esse parasito fez fermentação lática; em relação ao metabolismo intermediário foi possível detectar os ácidos orgânicos citrato, α-cetoglutarato, succinato, fumarato, malato e oxaloacetato, indicando que os cisticercos apresentaram elementos do ciclo do ácido cítrico para produzir energia. Porém diante dos resultados encontrados a principal evidência é de que esses cisticercos utilizaram uma via denominada reversão do ciclo do ácido cítrico para produzirem energia, obtendo como produtos finais propionato e acetato. Não se observou diferenças significativas entre os grupos que sofreram tratamentos e os grupos controle exceto pelo fato de que, o grupo tratado com albendazol 6,0 mg/kg de peso do camundongo teve como principal produto final da reversão do ciclo do ácido cítrico, o acetato, ao contrário dos demais grupos que tiveram o propionato; nos grupos tratados com praziquantel não houveram a produção de acetato como nos demais grupos e no grupo tratado com praziquantel 6,0 mg/kg não houve a produção de β-hidroxibutirato; nos grupos tratados com albendazol observou-se um aumento na produção de β-hidroxibutirato indicando que esse grupo maximizou a via de produção energética através da β-oxidação de ácidos graxos e/ou através do catabolismo proteico.Concluímos que os cisticercos de T. crassiceps, nas condições do presente trabalho, deram preferência para uma via de produção energética anaeróbia e provavelmente catabolizaram, além de carboidratos, proteínas e ácidos graxos para produzirem ATP.

La actividad sináptica regula una larga lista de procesos esenciales para el desarrollo y la supervivencia neuronal mediante cambios en los niveles de Ca2+ citoplasmático. Modula desde la morfología del árbol dendrítico neuronal hasta la plasticidad neuronal o los mecanismos de protección contra el daño oxidativo. La entrada de Ca2+ a través de los receptores sinápticos promueve diferentes cambios a nivel celular que van desde la remodelación del citoesqueleto a la estimulación de diferentes cascadas de señalización que finalmente activan diferentes factores de transcripción implicados en la regulación de toda una serie de programas de transcripción. Durante las primeras semanas tras el nacimiento tiene lugar el período, enmarcado dentro del desarrollo del sistema nervioso, en que se da una mayor tasa de crecimiento de dendritas y axones. El crecimiento neuronal debe ir acompañado de la síntesis de nuevos lípidos, necesarios para la generación de nuevas membranas. A pesar de que los mecanismos implicados en la regulación del crecimiento de dendritas y axones han sido ampliamente estudiados, se conoce muy poco con respecto a los cambios a nivel metabólico destinados a la síntesis de las biomoléculas necesarias para abastecer la generación de nuevas membranas. El principal objetivo de este trabajo ha sido estudiar el mecanismo por el que la actividad sináptica es capaz de regular el metabolismo neuronal para promover la síntesis de diferentes metabolitos precursores para la síntesis de los lípidos necesarios para el crecimiento neurítico. El principal modelo utilizado en este estudio ha sido los cultivos primarios de neuronas corticales de embriones de rata. La actividad sináptica se ha estimulado mediante un protocolo basado en la desinhibición de la red neuronal, que consiste en el tratamiento combinado con Bicuculina y 4-aminopiridina. Los resultados de esta tesis demuestran que la actividad sináptica estimula la captación y el metabolismo de la glucosa incrementando la transcripción del principal transportador de glucosa en neuronas (Glut3) y de diferentes enzimas implicadas en la glucólisis (HK2, PKM1/2 y PFKFB3). Todo el proceso está regulado por un mecanismo caracterizado por la activación de dos factores de transcripción: CREB y HIF-1α. La activación de CREB, uno de los principales factores de transcripción regulados por la actividad sináptica, promueve la expresión de Glut3 y de la enzima ubiquitina ligasa, Siah2. La actividad de esta última es necesaria para promover la estabilización y, en consecuencia, la activación de HIF-1α, que finalmente promueve la expresión de las enzimas implicadas en la glucólisis. La inhibición de la glucólisis o el bloqueo de la actividad de HIF-1α son suficientes para inhibir la estimulación del crecimiento neuronal por la actividad sináptica. De acuerdo con estos resultados en cultivos celulares, la inhibición de esta vía o la eliminación de este factor de transcripción durante las primeras semanas de desarrollo postnatal en roedores tiene como consecuencia una reducción de la complejidad del árbol dendrítico. Tal como se muestra en el presente trabajo, la actividad sináptica también podría estar implicada en la regulación de diferentes orgánulos como son los peroxisomas y las mitocondrias. En el primer caso se ha observado que la actividad sináptica promueve la síntesis de diferentes implicados tanto el desarrollo y el mantenimiento de los peroxisomas (Pex5, Pex11b y Pex13) como en el metabolismo lipídico peroxisomal (ABCD2 y ACOT8). Cuanto a las mitocondrias se ha comprobado que las neuronas estimuladas sinápticamente activas presentan un incremento en el transporte anterógrado mitocondrial a lo largo del axón que depende de la actividad de HIF-1α.<br>During the first weeks after birth occurs the period of the nervous system development in which takes place the highest dendritic and axonal growth ratio. Neuronal growth must be accompanied by the synthesis of new lipids, which are necessary for the formation of new membranes. Although the mechanisms involved in the regulation of dendrites and axons growth have been widely studied, there is very little known about the changes at the metabolic level involved in the synthesis of the biomolecules necessary to supply the formation of new membranes. The main goal of this doctoral thesis has been studying the mechanism by which synaptic activity, one of the most important inducers of neurite growth, regulates neuronal metabolism to promote the synthesis of different precursor metabolites involved in the synthesis of lipids required for neuritic growth. The results of this thesis show that synaptic activity stimulates glucose uptake and metabolism by increasing the transcription of the main glucose transporter in neurons, Glut3 and different enzymes involved in glycolysis. The whole process is regulated by a mechanism characterized by the activation of two transcription factors: CREB and HIF-1α. The activation of CREB, one of the main transcription factors regulated by synaptic activity, promotes the expression of Glut3 and the ubiquitin-protein ligase, Siah2. The activity of the latter is necessary to promote the stabilization and, consequently, the activation of HIF-1α, which finally promotes the expression of enzymes involved in glycolysis. Glycolysis inhibition or blocking of HIF-1α activity are sufficient to inhibit stimulation of neuronal growth by synaptic activity. A shown in this thesis, synaptic activity may also be involved in the regulation of peroxisomes and mitochondria. In the first case, it has been observed that synaptic activity promotes the synthesis of different agents involved in the development and maintenance of peroxisomes and in peroxisomal lipid metabolism. In regard to mitochondria, it has been proven that synaptically active neurons exhibit an increase in mitochondrial anterograde transport along the axon which requires HIF-1α activity.

Nesta tese de doutorado foram investigados parâmetros imunológicos de 3 espécies de anuros ao longo de diferentes estágios de história de vida, bem como a relação dos níveis plasmáticos de hormônios esteroides com as variáveis imunológicas (Capítulo 1). O capítulo 2 aborda os ajustes sazonais de reguladores metabólicos em diferentes músculos relacionados a reprodução e locomoção no período reprodutivo e de seca em três espécies de anuros. O capítulo 3 aborda a relação causal entre o aumento dos níveis plasmáticos de hormônios esteroides e a imunomodulação da resposta inflamatória e da resposta mediada por proteínas do sistema complemento na espécie R. jimi. Adicionalmente, foram investigados possíveis ajustes anuais do custo energético da resposta inflamatória, da taxa metabólica padrão, e a relação dessas variáveis com os níveis plasmáticos de hormônios esteroides na espécie R. jimi (Capítulo 4). O conjunto de dados dos capítulos 1 e 4 apontam na direção contrária da hipótese de imunossupressão durante a temporada reprodutiva, altos níveis de hormônios esteroides (testosterona e corticosterona) aumentam os paramêtros imunitários, bem como a resposta imune. Os resultados apresentados no capítulo 3 corroboram os efeitos imunomodulatórios do tratamento agudo de testosterona e corticosterona em anfíbios anuros. Por fim, o capítulo 2 mostra a variação sazonal de expressão de proteínas que agem como reguladores metabólicos. Essas reguladores, mediam a manutenção de células e tecidos durante a seca, fazendo com que os músculos associados a reprodução e locomoção não degradem durante este período. Adicionamente, a espécie estivadora ativa vias que diminuem o consumo de ATP, fazendo que com que haja economia de reservas energéticas durante a estivação<br>We investigated the correlation between imunological parameters, steroid plasma levels in 3 anuran species during different life history stages (Chapter1). Chapter 2 explore the seasonal adjusments of metabolic regulators for 3 anuran species. In the 3th chapter, we studied the casual relationship between increase of steroid plasma levels and immunomodulation in males of Rhinella jimi. Additionally, w e investigated the metabolic cost of the immune response, standard metabolic rate, and steroid plasma levels of R. jimi (Chapter 4). Our results point to an opposite direction of the immunocompetence handicap hypothesis (Chapter 1 and 4), high steroid plasma levels are associated to higher immune response. The results found in the chapter 3 corroborates the immunomodulatory effects of testosterone and corticosterone. Chapter 2 shows the seasonal variation of metabolic regulators, which guarantee muscle and cellular maintenance during the dry period for all three species. The aestivating species activates pathways that shut down ATP consumption saving energy during the drought

Coronary heart disease (CHD) is the leading cause of morbidity and mortality in the developed world. Attempts to prevent CHD using LDL lowering medications have been only partly successful and new approaches are under investigation. Significant efforts are being made to develop therapeutics that raise plasma HDL levels to aid in the prevention of CHD. HDL levels are believed to be inversely associated with the risk of developing CHD. Naturally occurring phospholipids such as phosphatidylinositol (PI), have been shown to increase plasma apoA-I levels and HDL levels in animal models and human subjects; but the mechanism remains to be elucidated. Since in humans, HDL is primarily synthesized in the liver, the objective of the present study was to evaluate the underlying molecular mechanism of PI-induced apoA-I and HDL secretion from liver cells. We show that PI doubles apoA-I/HDL secretion at 24h in a model hepatocyte, HepG2, cell culture system. PI-induced apoA-I secretion is unaffected by PI-3-kinase inhibitors but is sensitive to various MAP kinase inhibitors. While the p38MAPK inhibitor SB203580 has no effect on PI-induced apoA-I secretion, the MEK1/2 inhibitor U0126 blocks PI-induced apoA-I secretion. Inhibition of the JNK MAPK pathway by SP600125 also blocks PI mediated apoA-I secretion. Real-time PCR shows no changes in cellular apoA-I mRNA and suggests that PI is not impacting the transcription of the apoA-I gene. However, the degradation of apoA-I is decreased in PI treated HepG2 cells. Collectively, the data from these investigations suggest that PI acts through mitogen and stress-activated protein kinase pathways to increase plasma apoA-I levels by decreasing the degradation of apoA-I.

Made available in DSpace on 2014-06-11T19:28:01Z (GMT). No. of bitstreams: 0 Previous issue date: 2001Bitstream added on 2014-06-13T19:57:13Z : No. of bitstreams: 1 barreiros_rc_me_botfm.pdf: 2089159 bytes, checksum: 5458c167603553aa52e8877b42426d13 (MD5)<br>O metabolismo da frutose, bem como o seu nível sangüíneo em recém-nascidos não está bem esclarecido. A frutose é uma hexose encontrada normalmente no organismo humano e tem seu metabolismo associado à glicose e ao sorbitol. As principais fontes de frutose são os vegetais e o mel. O leite materno não contém frutose. O metabolismo desse açúcar é independente da insulina o que o torna uma boa opção para utilização em pacientes com deficiência desse hormônio. Além da independência da insulina, o metabolismo da frutose é caracterizado por uma rápida fosforilação e rápida conversão em glicogênio e glicose ou conversão em glicerol, para posterior utilização no metabolismo lipídico. A frutose pode ser produzida endogenamente, a partir da glicose, através da via do sorbitol. Apesar de ser utilizado há tempos clinicamente como uma alternativa à glicose, existem poucos trabalhos na literatura determinando os níveis normais em humanos. Isso ocorreu em grande parte devidas dificuldades na dosagem desse carboidrato: em virtude de ser difícil a sua diferenciação da glicose, outra hexose, além da frutose ser encontrada em pouca quantidade em fluídos orgânicos...<br>The metabolism of fructose as well the blood-levels of fructose in newborn infants are not yet well established. Fructose is an hexose found naturally in fruits, vegetables and honey and its metabolism is associated with glucose and sorbitol. The human milk does not contain fructose. The metabolism of fructose is insulin independent, which makes it an alternative to glucose. Besides its independence of insulin, fructose is rapidly metabolized primarily in the liver, where occurs phosphorylation and conversion to glycogen and glucose or to glycerol, to further utilization in lipid metabolism. Fructose, also, can be produced by the human organism, originating from glucose by the sorbitol pathway. Although fructose is utilized since 1893 for medical purposes, there are few studies in the literature about normal levels of fructose in humans. This lack of studies is due in part to difficulties to determine this sugar in human fluids: glucose interferes in the final results and levels of fructose in biological fluids are very low. Our main goal was to establish the normal levels of fructose in newborn infants at 48 hours of life, with adequate weight for gestational age, breast-fed exclusively and to correlate the level of fructose with the levels of glucose and sorbitol. For this purpose we used the High performance liquid chromatography was used. Our study group was selected among breast-fed term newborn... (Complete abstract click electronic access below)

Orientador: Eliana Cotta de Faria<br>Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas<br>Made available in DSpace on 2018-08-09T21:29:39Z (GMT). No. of bitstreams: 1 Bassora_FernandaDutraSantiago_M.pdf: 2612060 bytes, checksum: 0ddcd23de6e1ce1f879247d7a9c9b2d7 (MD5) Previous issue date: 2007<br>Resumo: Está bem estabelecida na literatura especializada a associação inversa entre as concentrações plasmáticas de colesterol das lipoproteínas de alta densidade (HDL-C) e a incidência de doença arterial coronariana (DAC). Além de propriedades anti-oxidante, anti-inflamatória e anti-trombótica, a HDL participa do transporte reverso de colesterol, via pela qual o colesterol é captado das lipoproteínas e das membranas células periféricas e transportado ao fígado para sua excreção na forma livre ou de ácidos biliares. A lipase hepática (LH) possui função crucial no transporte reverso do colesterol, por sua atividade lipolítica e pela função de ligante à lipoproteínas facilitando sua captação tissular. A proteína de transferência de ésteres de colesterol (CETP), e mesma importância metabólica, promove a troca de ésteres de colesterol por triglicérides entre a HDL e as lipoproteínas ricas em triglicérides. Mutações nos genes que codificam estas proteínas têm sido muito estudadas para se compreender a função destas no metabolismo lipídico. O modelo experimental da hiperalfalipoproteinemia tem sido utilizado no decorrer dos últimos anos com o intuito de elucidar os mecanismos de ação da HDL e das proteínas reguladoras do seu metabolismo. A hiperalfalipoproteinemia é caracterizada pelo aumento das concentrações de HDL-C e é causada principalmente por deficiências genética de CETP e/ou LH. Os objetivos desta dissertação foram o de se estabelecer à modulação da hiperalfalipoproteinemia sobre os parâmetros antropométricos, bioquímicos, moleculares (¿514C/T do gene da LH e I405V do gene da CETP) e radiológicos (espessura da camada íntima média de carótidas) em uma amostra populacional brasileira. O estudo foi conduzido em 291 voluntários de ambos os sexos, classificados como hiperalfalipoproteinemicos (Hiper-A), HDL-C =68mg/dL, ou controles, HDL-C<68 e =32 mg/dL, de acordo com o valor do percentil 90, obtido em um estudo prévio do Laboratório de Lípides a partir população normolipidêmica. Os polimorfismos LH-514C/T e CETP I405V foram identificados através de técnicas de reação em cadeia polimerase (PCR) e a espessura da camada íntima-média de carótidas (EIM) pela ultra-sonografia de alta resolução. Em um primeiro trabalho observou-se em um sub-grupo de 169 indivíduos, com a medida da EIM, que somente a idade foi correlacionada com a EIM na hiperalfalipoproteinemia, enquanto que em controles houve modulação positiva pela idade, sexo masculino, pressão arterial sistólica, e controversamente com relatos da literatura, com HDL-C. Apesar de Hiper-A possuir um perfil com maior número de fatores de risco cardiovasculares, a semelhança encontrada na EIM de carótidas, assim como, da freqüência de EIM maior que 1 mm poderia, em parte, ser explicada pela grande diferença de modulação entre os grupos apontando para um traço protetor contra a aterosclerose carotídea em hiperalfalipoproteinemia. A ateroproteção reduzida em controles, tanto em homens quanto em mulheres, está de acordo com a observada associação negativa neste grupo entre EIM e a CETP com possível presença de HDL com a composição química alterada (ricas em TG e pobres em ésteres de colesterol), e ocorreu possìvelmente no sub-grupo masculino, com perfil pró-aterogênico evidente. Em um segundo trabalho, no sub-grupo de 169 indivíduos, com a medida da EIM, foi avaliado o efeito do polimorfismo LH-514C/T sobre a espessura da camada íntima-média de carótidas na hiperalfalipoproteinemia. Não se observou nenhuma variação de EIM em ambos os grupos em função deste polimorfismo. Quando comparados os grupos, o genótipo CC do polimorfismo LH-514C/T mostrou apenas tendência a maior EIM de carótidas em hiperalfalipoproteinemia (p<0,09), mas a freqüência de EIM maior que 1 mm foi igual. Em um terceiro trabalho, em 282-291 indivíduos foram avaliadas as semelhanças de freqüências entre os polimorfismos LH-514C/T e CETP I405V na hiperalfalipoproteinemia e normolipidemia. Ambos apresentaram altas freqüências, similares entre grupos e entre o polimorfismo LH-514C/T, CC 39%, CT+TT 61%; e o polimorfismo CETP I405V: II 26%, IV+VV 74% e CTL: CC 40%, CT+TT 60%, II 43% e IV+VV 57%. Descrevemos o polimorfismo LH-514C/T na hiperalfalipoproteinemia os TT vs CC apresentaram cintura menor, concentrações mais baixas de colesterol plasmático (C), fosfolípides (FL), LDL-C, estimativa do tamanho da LDL (LDL-C/ApoB). O polimorfismo CETP I405V na hiperalfalipoproteinemia em VV vs II, mostrou alta pressão arterial sangüínea e menores concentrações plasmáticas de HDL2TG e HDL3TG. O genótipo IV teve maiores concentrações plasmáticas de ApoAI e pressão arterial diastólica quando comparado com o genótipo II. Em resumo esta dissertação aponta para efeitos ateroprotetores ou neutros da hiperalfalipoproteinemia em uma amostra de população brasileira sobre a aterosclerose carotídea, inclusive no polimorfismo LH -514C/T. Os polimorfismos LH-514C/T e CETP I405V foram muito semelhantes com relação aos lípides e lipoproteínas séricos, mas não às proteínas reguladoras, oferecendo modulação protetora na hiperalfalipoproteinemia<br>Abstract: There is an inverse relationship between plasma concentration of high-density lipoprotein (HDL-C) and the risk of coronary arterial disease (CAD). Beyond anti-oxidant, anti-inflammatory and anti-thrombotic properties, HDL plays a role on the reverse cholesterol transport, where cholesterol is taken from lipoproteins and peripheral cells to the liver for excretion. Hepatic lipase (HL) plays a key role in this process, by its lipolitic activities and ligand functions. Cholesterol ester transfer protein (CETP), of equal metabolic importance, facilitates the exchange of cholesterol ester and triglycerides between HDL and triglyceride rich-lipoproteins. Mutations and polymorphisms of these enzymes have being studied in order to evaluate its activity and metabolic consequences. Hyperalphalipoproteinemia (Hyper-A) has being used in the latest years with the purpose of evaluating the anti and pro-atherogenic mechanisms of HDL and of regulating proteins. The aim of this work was to establish the modulation of hyperalphalipoproteinemia in relation to controls on the anthropometric, biochemical, radiological and molecular manifestations. This study was conducted on 291 volunteers, classified as Hyper-A, HDL-C=68mg/dL and controls, HDL-C <68 e 32 mg/dL according to the percentile 90th, obtained from a local normolipidemic population study. We determined clinic data, lipid, lipoproteins and radiological parameters of volunteers. The HL-514C/T and CETP I405V polymorphism were determined by polymerase chain reaction methods. The carotid intima-media thickness measurements were performed high performance ultrasound. We showed in the first manuscript that although possessing a higher risk coronary vascular disease profile the similarity found in carotid could in part be explained by the striking differences in its modulation between the two groups, indicating a protective trait against carotid atherosclerosis in hyperalphalipoproteinemia. In the Hyper-A population, was only correlated with age, while in controls had a positive correlation with age, male sex, systolic blood pressure, and surprisingly with HDL-C. This dissociation between IMT and HDL-C could be accounted for by a small HDL particle number in CTL. In the manuscript 2, the ¿514C/T polymorphism did not contribute to variations in the carotid IMT and Hyper-A did not modulate the IMT variations, contrary to Rundek et al., (2002) who investigated the ¿514C/T polymorphism on variations in the carotid IMT in 87 stroke-free subjects suggested that CC genotypes had increase of carotid IMT, FMT and HALP. The HL-514C/T e CETP I405V polymorphisms, were no associate, were highly prevalent in the two groups but were not associated with HDL-C. In Hyper-A, LH-514C/T induced lower plasma cholesterol (C), phospholipids (PL), LDL-C and LDL size (LDL-C/ApoB). In Hyper-A CETP I405V decreased blood pressure, reduced TG in HDL subfractions 2 and 3 of (HDL2TG and HDL3TG) and increase ApoAI. The HL -514C/T polymorphism in Hyper-A the TT vs CC had lower waist hip-circumference, cholesterol (C) concentrations, phospholipids (PL), LDL-C and estimated size particle by LDL-C/ApoB. The genotype TT was different between 2 groups: in Hyper-A with relation the CTL, had lower HL, estimated size particle by TG/HDL-C and higher HDL2C, HDL3C, HDL3TG, ApoAI and C concentrations and had higher C, estimated size particle by LDL-C/ApoB, ApoAI, HDL2C, HDL3C and estimated size particle by TG/HDL-C. The CETP I405V polymorphism in Hyper-A, the VV vs II had higher Systolic Blood Pressure and lower HDL2TG e HDL3TG concentrations. The IV genotype had higher ApoAI concentration and Diastolic Blood Pressure. In Hyper A, the VV genotype had higher HDL2C, HDL3C, ApoAI, e TG concentrations and reduced concentration of VLDL- and estimated size particle of LDL by TG/HDL-C. In summary, this work indicates an athero-protector and neutral effect on the carotid atherosclerosis in Hyper-A between HL-514C/T and CETP I405V polymorphisms both modulated for plasma lipids more atheroprotective<br>Mestrado<br>Ciencias Basicas<br>Mestre em Clinica Medica

Foi estudado o comportamento bioquímico energético de populações domiciliadas de Panstrongylus megistus de quatro regiões geográficas brasileiras (Região Tropical Atlântica, Região Floresta de Inclusão, Região do Agreste e Região da Caatinga), através dos seguintes parâmetros do metabolismo energético: Proteinas, Glicose, Deidrogenase láctica, Creatino-quinase e respectivas isoenzimas. Os espécimens foram mantidos em jejum de O a 90 dias. Foram observadas diferenças de metabolismo energético entre populações de duas regiões: Tropical Atlântica (Grupo I) Floresta de Inclusão (Grupo II)<br>The population\'s behaviour of Panstrongylus megistus, which is domiciliated in four different geographycal brazilian regions (\"Tropical Altântica\" system and \"Inclusão\" Forest, and \"Agreste\" and \"Caatinga\" regions, was studied by different bioenergetic metabolism parameters: Proteins, Glucose, Lactate dehydrogenase, Creatine Kinase and their respective isoenzymes. These specimens were kept from 0 to 90 days fasting. During this time, it was observed many differences in the energetic metabolism of the vectors of Chagas\' disease in the \"Tropical Atlântica\" system (Grupo I) \"Inclusão\" Forest (Grupo 11)

Os viperídeos estão entre as serpentes de menor custo energético de manutenção, o que é, geralmente, relacionado a seus baixos níveis de deslocamento e a tática de forrageio por emboscada. Dois importantes componentes do alocamento energético destes animais são: 1) o metabolismo de repouso padrão, que está relacionado ao custo de manutenção visceral durante o repouso e 2) o aumento metabólico pós-prandial (AMPP), que está relacionado ao custo do processo digestório. O presente trabalho teve como objetivos estudar as variações da TMRP (taxa metabólica de repouso padrão) e do AMPP em 5 espécies pertencentes ao gênero Bothrops levando em conta aspectos filogenéticos, ontogenéticos e testando hipóteses acerca de possíveis variações destes componentes em função de características ecológicas. Além disso, o presente trabalho testou, através de ferramentas de ponderação filogenética, a hipótese atualmente aceita que prediz que as TMRPs em serpentes estão essencialmente ligadas a estratégia alimentar e não a filogenia. O trabalho está dividido em quatro capítulos, sendo o primeiro devotado a uma introdução geral acerca dos assuntos que serão abordados nos demais. Os resultados e discussões específicas estão divididos em dois capítulos redigidos em forma de artigo, primeiro referente aos estudos das taxas metabólicas de repouso (capítulo 2) e o segundo sobre o AMPP (capítulo 3). Por fim, a conclusão final acerca dos achados e as perspectivas para pesquisas futuras estão presente no capítulo 4.<br>The viperids are among the snakes with the lowest energetic maintenance costs which are generally related to the low mobility and ambush foraging mode. Two important components of energy allocation in these animals are 1) standard metabolic rates (SMR), related to the cost of keeping visceral components during resting and 2) specific dynamic action (SDA) , which is related to the cost of digestion. The present work aimed to study the variations of SMR and SDA in 5 species of the genus Bothrops taking into account aspects of phylogeny and ontogeny plus testing hypotheses about possible variations in these metabolic rates related to ecological characteristics. Furthermore, the present study used phylogenetic weighting tools to test the currently accepted hypothesis that predicts that SMR in snakes is related to the feeding strategy and not to phylogeny. The work is divided into four different chapters. Chapter 1 is devoted to a general introduction about the issues that will be addressed in the others chapters. The results and discussions are divided into two chapters presented as articles, the first referring to studies of SMR ( chapter 2 ) and the second to the SDA (chapter 3 ). Finally, in chapter 4 we include the final conclusion and prospects for future research

A regulação da expressão gênica do GLUT4 induzida pelo jejum e insulina em músculo sóleo de ratos e a participação do fator transcricional Nuclear Factor -kappa B (NF-<font face=\"symbol\">kB). Investigou-se a regulação induzida pelo jejum e insulina na expressão do GLUT4 no músculo sóleo de rato Wistar e a participação do fator transcricional NF-<font face=\"symbol\">kB. Analisou-se sóleo de rato alimentado e jejuado com músculo incubado ou não em KHB com ou sem insulina. Avaliou-se a expressão do GLUT4 por Northern e Western blotting e a atividade de ligação do NF-<font face=\"symbol\">kB ao DNA por Gel Shift. Sóleos de ratos jejuados diminuíram em 24% o mRNA de GLUT4, aumentaram 51% a atividade de ligação do NF-<font face=\"symbol\">kB e diminuíram 30% o conteúdo protéico total comparados aos alimentados. Sóleos de ratos jejuados aumentaram 38% o mRNA do GLUT4, diminuíram 30% a atividade de ligação do NF-<font face=\"symbol\">kB e sem alterar a proteína GLUT4 comparados aos incubados. A incubação com insulina por 70, 120 e 180 minutos do sóleo de ratos jejuados aumentou o mRNA de GLUT4 em 37%, 30% e 25%, diminuiu a atividade de ligação do NF-<font face=\"symbol\">kB em 30%, 40% e 50% e aumentou a proteína GLUT4 em 33%, 40% e 50% em relação a incubação com KHB. Os resultados indicam que o jejum e insulina regulam a expressão de GLUT4 com participação do NF-<font face=\"symbol\">kB.<br>The regulation of GLUT4 gene expression induced by fasting and insulin in soleus muscle of rats and the participation of transcriptional factor Nuclear Factor -kappa B (NF-<font face=\"symbol\">kB). It was investigated the regulation induced by fasting and insulin in the GLUT4 expression in soleus muscle of Wistar rat and the particiation of transcriptional factor NF-<font face=\"symbol\">kB. It was analysed soleus of fed-rats and fasted-rats with muscle incubated ou not in KHB with or without insulin. It was analysed the GLUT4 expression for Northern and Western blotting and the binding activity of NF-<font face=\"symbol\">kB to DNA for Gel Shift. Soleus of fasted-rats decreased 24% the GLUT4 mRNA, increased 51% the binding activity of NF-<font face=\"symbol\">kB and decreased 30% the total protein content compared to fed-rats. Soleus of fasted-rats increased 38% the GLUT4 mRNA, decreased 30% the binding activity of NF-<font face=\"symbol\">kB, without altering the GLUT4 protein compared to the incubated muscles. The incubation with insulin for 70, 120 e 180 minutes of the soleus of fasted-rats increased the GLUT4 mRNA in 37%, 30% e 25%, decreased the binding activity of NF-<font face=\"symbol\">kB in 30%, 40% e 50% and increased GLUT4 protein in 33%, 40% e 50% in relation to incubated muscles with KHB. The results indicate that fasting and insulin regulate the GLUT4 expression with participation of NF-<font face=\"symbol\">kB.

Orientador: Maria Terezinha Serrão Peraçoli<br>Resumo: O aumento da atividade dos radicais livres e da produção de citocinas pró-inflamatórias por monócitos pode estar envolvido na patogênese da pré-eclâmpsia. A silibinina é o componente mais ativo da silimarina (Silybum marianum), um flavonóide polifenólico que possui efeitos anti-oxidante e anti-inflamatório. O objetivo do presente estudo foi avaliar o efeito da silibinina sobre a produção de citocinas pró e anti-inflamatórias, bem como sobre a liberação de ânion superóxido (O2 -) por monócitos de gestantes com pré-eclâmpsia.Foram selecionadas 30 gestantes com préeclâmpsia (PE), 30 gestantes normais (GN) e 30 mulheres normais nãográvidas (NG). Monócitos de sangue periférico foram incubados na presença ou ausência de silibinina nas concentrações de 5 e 50 g/mL por 60min e, estimulados ou não com ester de forbol (PMA) para determinação de O2 -. Estes monócitos também foram estimulados ou não com lipopolissacáride (LPS) por 18h e, no sobrenadante das culturas foram determinadas, pela técnica de ELISA, as seguintes citocinas: fator de necrose tumoral-alfa (TNF- ), Interleucinas (IL-) IL-6, IL-10, IL-12, IL-15, fator estimulador de colônias de granulócitos e monócitos (GM-CSF) e fator de crescimento e transformação (TGF- . A atividade da enzima superóxido dismutase (SOD) foi avaliada em hemolisado de eritrócitos dos três grupos estudados.Níveis significativamente maiores de O2 - foram liberados por monócitos de gestantes com PE quando comparados com GN e NG, confirmando o estado ativado dessas células. A atividade da SOD foi significativamente maior no grupo de gestantes com PE. Os níveis endógenos de TNFforam significativamente mais elevados nas pacientes com PE, quando comparados com os grupos GN e NG, enquanto os níveis de IL-10 foram significativamente menores nas gestantes com PE. A capacidade de produção de citocinas pelos... (Resumo completo, clicar acesso eletrônico abaixo)<br>Abstract: Increased free radical activity and high proinflammatory cytokine production by monocytes may be implicated in the pathogenesis of preeclampsia. Silibinin is a major active component of silymarin (Silybum marianum), a polyphenolic plant flavonoid that has antioxidant and anti-inflammatory effects. This study investigated the in vitro effect of silibinin on monocyte production of pro- and anti-inflammatory cytokines, as well as on superoxide anion (O2-) release in pregnant women with preeclampsia.Thirty preeclamptic (PE), 30 healthy pregnant (HP) and 30 healthy non-pregnant (NP) women were included. Peripheral blood monocytes were incubated with or without silibinin at 5 and 50ug/mL for 60 min, and stimulated with or without phorbol myristate acetate (PMA) for the assessment of O2-. These cells were also cultured with or without lipopolysaccharide (LPS) for 18h and tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-6, IL-10, IL-12p40, IL-15, granulocyte-macrophage colony-stimulating factor (GM-CSF) and transforming growth factor-beta1 (TGF- 1) in monocyte culture supernatants were determined by ELISA. The activity of the enzyme superoxide dismutase (SOD) was evaluated in erythrocyte lisates of the three groups studied.Monocytes of preeclamptic patients release significantly higher levels of O2 - in comparison to HP and NP women confirming the activation state of these cells. SOD activity in erythrocytes was significantly higher in preeclamptic patients. The endogenous levels of TNFwere significantly higher in PE patients than in HP and NP groups, while IL-10 production was significantly lower in PE women. The levels of IL-6, IL-12 and GM-CSF spontaneously produced by monocytes were higher in HP and PE groups than in NP women. The capacity of cytokine production by LPS-stimulated monocytes was preserved in all the three groups studied... (Complete abstract click electronic access below)<br>Mestre

La línea de investigación presentada en esta tesis representa un primer acercamiento entre los estudios sobre Hidrología y Metabolismo Social. La línea nace de la observación de que el agua es evitada en los estudios que tratan el metabolismo y de que la ciencia del agua –si bien reconoce la necesidad de evolucionar hacia la interdisciplinariedad- todavía no ha conseguido conectar los análisis enfocados en la sociedad y en los ecosistemas. La contribución que se hace en este trabajo es precisamente la definición de un marco analítico –el Metabolismo Hídrico de los Socio-ecosistemas- donde se puede establecer esta conexión y que está formado por una propuesta conceptual y un set de herramientas metodológicas. El documento se divide en tres partes donde se discuten las novedades epistemológicas, metodológicas y formales del marco. La Parte I cubre las reflexiones epistemológicas relacionadas con el marco analítico. Éstas comienzan en el Capítulo 1 con la explicación de los restos a los que la ciencia del agua se enfrenta y que están relacionados con la necesidad de encontrar marcos analíticos que puedan proporcionar inputs relevantes para la gestión integrada de los recursos hídricos (GIRH). Al igual que para el caso de otros recursos, la GIRH requiere el establecimiento de una conexión analítica de las dinámicas sociales y de los ecosistemas. La analogía del metabolismo de la sociedad, como una de las piezas claves de la Ciencia de la Sostenibilidad, es una buena opción para establecer esta conexión. Sin embargo, el concepto de metabolismo necesita ser examinado de cerca antes de su uso combinado con otras concepciones de las relaciones entre el ser humanos y la naturaleza. Tras subrayas que el metabolismo de las sociedades y los ecosistemas son dos procesos distintos per conectados, en el Capítulo 2 se propone un esquema para la descripción de las relaciones metabólicas entre ellos. En el capítulo 3, este esquema es adaptado a las condiciones específicas del agua, usando algunos de los conceptos más relevantes en socio- y eco-hidrología. De esta forma, el metabolismo hídrico del socio-ecosistema es definido como el metabolismo del sistema agua-ser humano. La Parte II describe el marco metodológico. Como un marco ampliamente establecido que es capaz de tratar los problemas de escala y de integrar narrativas, el Capítulo 4 presenta el Análisis Integrado Multi-Escala del Metabolismo Social y de los Ecosistemas (MuSIASEM). MuSIASEM se ha seleccionado como raíz y ha sido adaptado al análisis de sistemas complejos agua-ser humano. Dado que el agua presenta importantes diferencias con respecto a los análisis previos en energía, esta adaptación requiere la inclusión de nuevas escalas de análisis –el ‘problemshed’ y el ‘watershed’- y nuevas definiciones del agua como metabolito –como flujo y fondo. En el capítulo 5 se señalan las diferencias y sinergias ente MuSIASEM el análisis de la huella hídrica –como una de las herramientas de la GIRH. En la Parte III se presentan cuatro casos de estudio con dos objetivos. En primer lugar, el Capítulo 6 analiza a sostenibilidad de los patrones metabólicos en el uso del agua en Punjab y Mauricio para testear la aplicación de MuSIASEM a los estudios de agua y para mostrar cómo este tipo de análisis de formaliza. En segundo lugar, el Capítulo 7 muestra como los métodos de contabilidad del agua del análisis de la huella hídrica complementan el análisis de flujos de agua en MuSIASEM, encontrando además una referencia para su contextualización.<br>The research line presented in this dissertation is a first attempt to provide a bridge for the communication between Hydrological studies and Social Metabolism. It was born from the observation that water is neglected in Social Metabolism and that current water science, while certain about the need of evolving towards a more interdisciplinary field, still faces challenges in the connection of social and ecosystem analyses. The contribution made here is the definition of an analytical framework –the Water Metabolism of Socioecosystems- where this connection can be established and which is formed by a conceptual proposal and a methodological toolkit. The document is divided in three parts where the epistemological, the methodological and the formal novelties of the framework are discussed. Part I covers the epistemological reflections related to the analytical framework. It begins in Chapter 1 with the explanation of the challenges faced by current water science and that relate to the need of finding analytical frameworks that contribute useful inputs to integrated management of the water resources (IWRM). As with the case of other resources, IWRM requires the analytical connection of the social and ecosystem dynamics. As a key piece within Sustainability Science the analogy of the metabolism of societies can be used to establish this connection. However, the metabolism concept needs a close examination before its joint use with other conceptions of the relations between humans and nature. After highlighting the need of considering the societal and ecosystem metabolism of socio-ecosystems as two separate but connected processes, a conceptual scheme is proposed in Chapter 2 to describe the metabolic relations between them. In Chapter 3, this scheme is adapted to the specifics of water using some of the most relevant concepts in socio- and eco-hydrology. In this way the water metabolism of socio-ecosystems is defined as the metabolism of the coupled water-human systems. Part II describes the methodological framework. In Chapter 4 the Multi-Scale Assessment of the Societal and Ecosystem Metabolism (MuSIASEM) is presented as an established framework able to deal with the scale issues and the integration of narratives. MuSIASEM is selected as a root and adapted to the analyses of coupled water-human systems. Since water presents some differences with the previous energy-focus analyses, its adaptation requires the inclusion of new scales of analysis –problemshed and watershed- and new definitions of water as a metabolite –as flow and fund. In Chapter 5 the differences and synergies between MuSIASEM and the water footprint analysis –as one of the tools of the IWRM- are highlighted. In part III four case studies are presented with two objectives. First, Chapter 6 assesses the sustainability of the metabolic patterns I Punjab and Mauritius in order to test the adaptation of MuSIASEM to water and to show how this type of analyses is made functional. Second, Chapter 7 shows how the water footprint accounting methods can complement the analysis of the water flows in MuSIASEM and how MuSIASEM, in turn an provide a space for their contextualization. Keywords: Agriculture, Complex Systems, Integrated Water Resources Management, Flow/Fund Model, Grammar, Multilevel Matrixes, MuSIASEM, Scale Issues, Socio-Ecological System, Social Metabolism, Virtual Water, Water, Water Footprint, New Water Culture.

Pharmaceuticals which are used worldwide are designed to facilitate the life for the human society and have an important role in treatment and prevention of disease for both humans and animals. They are ubiquitous in the aquatic environment and are mainly derived from municipal wastewater treatment plants (WWTPs) due to their low removal rate. Therefore, their presence in the environment is directly linked to the human impact. Various biological and abiotic processes in the environment can transform them to transformation products (TPs). In many cases, transformation is already initiated in the human body by a variety of drug-metabolizing enzymes. The metabolites formed through human metabolism present some modifications in their chemical structures that can differ in physicochemical properties to their parent compound. Once they are excreted from the human body, both the unmetabolised parent drug and their metabolites enter WWTPs by means of the sewer system. Since the WWTPs are not designed to remove completely pharmaceutical residues, the fraction not removed after the treatment will eventually end up in the receiving water bodies. Consequently, due to pharmaceutical transformations in the human body, biotransformations in WWTPs and phototransformations in surface water, they can potentially produce a high number of TPs in real world samples which makes their identification a challenge. In this thesis, two different approaches (TPs profiling and suspect screening) based on high resolution mass spectrometry (HRMS) for the detection and identification of TPs of pharmaceuticals were investigated. TPs profiling approach was applied for the identification of phototransformation products of an antiviral zanamivir (ZAN) in batch reactors filled with surface water. On the other hand, suspect screening approach was applied for evaluation of transformation, prioritization and identification of photoTPs of six iodinated contrast media in surface water. Finally, a combination of suspect and TPs profiling approach was applied for the detection of TPs of an anticonvulsant lamotrigine and its main human metabolite lamotrigine N2-glucuronide which were formed as the result of their degradation in both activated sludge and pH dependent hydrolysis. The TPs profiling approach for evaluation of these transformations is illustrated in the example of photodegradation of an antiviral ZAN with identification of its TPs in surface water (Chapter 3.). Here a set of lab- scale experiments was performed in order to determine the susceptibility of ZAN towards photodegradation under simulated and natural sunlight. The identification of the TPs was performed using hydrophilic interaction liquid chromatography coupled to high resolution mass spectrometry (HILIC-HRMS) where four photoTPs were tentatively identified and their proposed structures were rationalized by photolysis mechanisms. Kinetic experiments showed that photodegradation kinetics of ZAN in surface waters would proceed with slow kinetics since upon exposure of aqueous solutions of surface water (20 μg L-1) to simulated sunlight, ZAN was degraded with t1/2 of 3.6 h. Under natural sunlight irradiating surface water, about 30 % of the initial concentration of the antiviral disappeared within 18 days. However, when ZAN and its TPs were retrospectively screened from surface water extracts, neither the parent nor the TPs were detected. The results of this TP profiling used for the identification of TPs of ZAN, although straightforward, suggests that it is not suitable when dealing with a considerably elevated number of TPs formed in batch experiments. However, time and effort needed to be optimised for the structure elucidation of 108 photoTPs of six iodinated contrast media (ICM) (Chapter 3.). Again, the photodegradation study was performed in surface water spiked with the ICMs using a sunlight lab-scale simulator. 108 TPs were generated and each photoTP was characterised by its unique exact mass of the molecular ion and retention time and added to a suspect list. Once the suspect list was generated, the photoTPs were searched in thirteen surface water samples which were extracted using a generic solid- phase extraction method (four cartridges of different chemistries in order to retain ample number of compounds with different chemical properties). Based on their detection frequency (those TPs with the frequency higher than 50 % were deemed important), eleven TPs were prioritized and their structures elucidated by HRMS and NMR (when possible). Out of the eleven prioritised TPs, ten were formed as the result of deiodination (either by deiodination, oxidative deiodination or intramolecular elimination). In the real surface water samples, median concentration of parent compounds was 110 ngL-1 reaching up to 6 µgL-1 for iomeprol while TPs were found at median concentration of 8 ngL-1, reaching up to 0.4 µgL-1 for iomeprol TP651-B. Here detection-based prioritization served as a crucial step to reduce the number of TPs to be identified and thereby reducing costs and time for the subsequent target analysis. This time-effective approach not only guaranties that the degradation products elucidated would be found, but also that they are environmentally relevant. In summary, the proposed screening approach facilitates the evaluation of the degradation of polar compounds at a real scale with a fast detection of TPs without prior availability of the standards. Approach used for detection and identification of TPs of ICM in Chapter 3 was an example of suspect screening where the suspect list of TPs was generated at lab-scale, In Chapter 4, the work started with a suspect screening of lamotrigine (LMG) and related compounds (its human metabolites, synthetic impurities and photoTPs) which were listed from the literature and searched in wastewater and surface water samples. As the result of suspect screening, LMG, three human metabolites and a LMG synthetic impurity (OXO-LMG) were detected in the screened samples. Preliminary results showed significantly higher concentrations of OXO-LMG in wastewater effluent, suggesting its formation in the WWTPs. However, biodegradation reactors amended with mixed liquor at neutral pH showed that LMG is resistant to biodegradation with only about 5 % elimination after 6 days. Since LMG is extensively and predominantly metabolised by phase II metabolism to its N2-glucuronide, this metabolite (LMG-N2-G) was degraded following the same experimental setup. Results showed that this metabolite was the actual source of the TP detected. Additionally, in batch experiments, LMG-N2-G was transformed, following pseudo-first kinetics, to three TPs as a result of i) deconjugation (to LMG), ii) oxidation of the glucuronic acid (to LMG-N2-G-TP430) and iii) amidine hydrolysis in combination with deconjugation (to OXO-LMG). In order to further rationalize the formation of the TP OXO-LMG, the stability of LMG-N2-G and related compounds was studied as a function of pH in the range of 4 – 9. Same as during biodegradation, LMG was stable across the entire pH range tested. However, LMG-N2-G was transformed to three TPs at neutral – basic pH. They were identified as TPs formed after hydrolysis of amidine and guanidine moieties. The third TP detected was an intermediated in the guanidine hydrolysis reaction. Kinetic experiments in wastewater samples at different concentration (20 and 200 nM) and pH (pH 6.5, 7, 8, 8.5 and 9) demonstrated that while the degradation constants were concentration independent, at higher pH, LMG-N2-G degraded at higher rate. The pH-dependent stability experiments of related compounds with different nitrogen N2-substituents on the 1,2,4-triazine ring showed that reaction of amidine and guanidine hydrolysis depends on imine tautomer equilibrium whose formation depends directly on the N2-supsitutent. LMG- N2-G major abiotic TP (amidine hydrolysis TP) was detected in hospital effluent and WWTP influent samples. Having in mind the concentrations of both biotic and abiotic TPs detected, a total mass balance at two- concentration levels batch reactors was closed at 86% and 102%, respectively. In three WWTPs total mass balance of LMG-N2-G ranged from 71-102%. Finally, LMG-N2-G and its TPs were detected in surface water samples with median concentration ranges of 23–186 ngL-1. The work presented in this chapter gives a new insight into the behaviour of glucuronides of pharmaceuticals, suggesting that they might also be sources of yet undiscovered, but environmentally relevant TPs.<br>Els productes farmacèutics, l'ús dels quals s'estén a nivell mundial, estan dissenyats per millorar la qualitat de vida de la societat i juguen un paper clau en el tractament i la prevenció de malalties, tant en homes com en animals. Aquests compostos químics es troben de forma ubíqua en el medi ambient. Això es deu principalment a les estacions depuradores d'aigua residual (EDARs), les quals no són capaces d'eliminar de manera eficient aquest tipus de compostos, ja que no estan dissenyades amb aquesta finalitat. Per tant, la presència de fàrmacs en el medi ambient està directament relacionada amb l'activitat humana. Un cop al medi ambient, l'estructura d'aquests compostos pot ser modificada per diferents processos biològics i abiòtics, generant-se així els que es coneixen com a productes de transformació (PTs). De fet, la transformació dels fàrmacs pot iniciar-se en alguns casos en el cos humà, després de la seva administració a causa de l'activitat metabòlica dels diferents enzims que posseeix l'home. Els metabòlits formats en aquests processos presenten algunes modificacions en les seves estructures químiques pel que fa al compost original, i, en conseqüència, unes propietats fisicoquímiques diferents. Un cop excretats, tant el fàrmac original no metabolitzat com els seus metabòlits arriben a les EDARs mitjançant la xarxa de sanejament municipal d'aigües residuals. La fracció d'aquests compostos que no s'elimina en els diferents tractaments realitzats en l'EDAR, es descarrega juntament amb l'efluent de la planta als aigües receptores. El gran nombre de transformacions que poden experimentar els fàrmacs en el seu cicle de vida a causa del seu metabolisme en el cos humà, la seva biotransformació per microorganismes i la seva fototransformació per llum solar, pot generar un nombre molt elevat de PTs en el medi ambient, i, per tant, la identificació dels mateixos, necessària per avaluar el destí dels fàrmacs en el medi ambient, és un desafiament. En el desenvolupament d'aquesta tesi es van aplicar dues aproximacions analítiques diferents: a)avaluació de perfils de PTs generats en experiments a escala de laboratori i b) anàlisi qualitativa dirigida suspect screening en mostres reals, tots dues basades en espectrometria de masses d'alta resolució (HRMS) per a la detecció i identificació de PTs de productes farmacèutics. L'aproximació d'avaluació de perfils de PTs en reactors a escala de laboratori es va aplicar per identificar productes de fototransformació (fotoPTs) de l'antiviral zanamivir (ZAN) en aigua superficial. L'aproximació de suspect screening es va aplicar per prioritzar i identificar fotoPTs de sis mitjans de contrast radiològics iodats (ICM) en aigua superficial. Finalment, una combinació de les dues aproximacions es va aplicar per detectar PTs de l’anticonvulsiu lamotrigina (LMG) i del seu principal metabòlit humà, el lamotrigina-N2-glucurònid (LMG- N2-G), resultants de la seva degradació tant en fangs activats com a reaccions d'hidròlisi a diferents valors de pH.