Thesis (Ph. D.)--West Virginia University, 2010. / Title from document title page. Document formatted into pages; contains vii, 105 p. : ill. (some col.). Includes abstract. Includes bibliographical references (p. 92-105).

Thesis (Ph. D.)--Miami University, Dept. of Zoology, 2007. / Title from second page of PDF document. Includes bibliographical references.

Toll-like receptor (TLR)-4 is a pattern recognition receptor (PRR) that recognises the pathogen-associated molecular pattern (PAMP) lipopolysaccharide (LPS) produced by Gram-negative bacteria. LPS binds to Myeloid differentiation 2 (MD-2)/TLR-4 heterodimers, driving their dimerisation and inducing a conformational change of the intracellular TLR-4 toll/interleukin-1 receptor (TIR) domains. The adaptor protein Myeloid differentiation primary response gene 88 (MyD88)-adaptor-like (Mal)/TIR domain-containing adaptor protein (TIRAP) then binds to the TIR domains of TLR-4 and acts as a bridge for MyD88 which goes on to form the myddosome, a large protein complex of six to eight MyD88 molecules and four Interleukin-1 receptor- associated kinase (IRAK) 4 and four IRAK1/2 molecules. This triggers a signalling cascade which results in nuclear factor (NF)-κB transcription factor activation and production of pro-inflammatory effector molecules such as the cytokine Tumour Necrosis Factor (TNF)-α. Upon activation TLR-4 is also endocytosed where it interacts with a second set of adaptor proteins TIR-domain-containing adaptor- inducing interferon (IFN)-β (TRIF)-related adaptor molecule (TRAM) and TRIF to initiate the type I IFN response. How TLR-4 dimerisation results in the formation of the oligomeric myddosome is not fully understood, but it is possible that the stoichiometry of Mal/TIRAP may be important in the formation of this protein complex. The aim of my thesis was to determine the stoichiometry of Mal/TIRAP at the plasma membrane of immortalised bone marrow derived macrophages (iBMDMs) and whether this stoichiometry changes upon stimulation with different TLR-4 ligands. To investigate Mal/TIRAP stoichiometry I first developed a viral transduction experimental cell model to visualise fluorescently labelled Mal/TIRAP. Mal/TIRAP-/- iBMDMs were lentivirally transduced with a Mal/TIRAPHALO construct. The halotag was fluorescently labelled then the cells were stimulated with TLR-4 ligands, such as LPS, fixed at different time points, then imaged. Total internal reflection fluorescence (TIRF) microscopy was used to image the plasma membrane and photobleaching experiments performed to determine Mal/TIRAP stoichiometry. I developed a computer-based analysis pipeline to analyse the resulting photobleaching data. Under resting conditions, Mal/TIRAP is present at the plasma membrane in clusters of approximately ten Mal/TIRAP molecules per cluster. After five minutes of stimulation with 10 ng/ml LPS Mal/TIRAP redistributes into cluster sizes of approximately six, twelve and much larger. After ten and fifteen minutes stimulation with 10 ng/ml LPS the clusters return to the resting size of approximately ten Mal/TIRAP molecules per cluster with a few much larger clusters remaining present. This confirms the rapid time frame within which TLR-4 signalling occurs at the plasma membrane and is consistent with myddosome stoichiometry of six MyD88 molecules or proposed super myddosomes of twelve MyD88 molecules. The computer-based analysis pipeline developed can be used to analyse any protein of interest at the plasma membrane. Protein ligands have also been found to activate TLR-4; for example allergens, such as Fel d 1 and Der p 2, as well as endogenous damage associated molecular patterns (DAMPs), such as extracellular matrix (ECM) proteins, for example fragments of fibronectin and tenascin-C. The mechanism by which these proteins interact with TLR-4 and induce signalling is unclear. Proteins from the ECM (fragments FNIII1c, FNIII13-14, FNIII9-E and FNIII9-E-14 from fibronectin and the fibrinogen-like globe (FBG) domain of tenascin-C) were tested using a transient transfection assay in HEK293 cells and shown to activate TLR-4. In conclusion, I have developed new tools and methodology to investigate how TLR-4 signals in response to LPS and DAMPs in living cells. Whether DAMP- activated TLR-4 forms similar signalling complexes to those induced by LPS will form part of a future study.

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate: Tereza Morávková Supervisor: Assoc. Prof. Přemysl Mladěnka, Pharm.D., Ph.D. Title of Thesis: Assessment of the complex stoichiometry of the active dexrazoxane metabolite ADR-925 with iron and copper by the standard Job'smethod Iron and copper are important trace elements which participate on many physiological processes in humans. Their kinetics in the organism is tightly regulated since both lack or excess of these elements are associated with pathological states. Free ions of iron and copper can catalyse reactive oxygen species (ROS) production and hence cause damage to proteins and DNA. Imbalance in these metals is linked with diabetes, cardiotoxicity, cirrhosis of the liver and neurodegenerative diseases as Alzheimer's disease or Parkinson's disease. Dexrazoxane is a drug with documented protecting effect against cardiotoxicity of anthracyclines. A former theory associated its protective effects against these cytotoxic drugs with the iron-chelating properties of its active metabolite ADR-925. The goal of this diploma thesis was to explore if ADR-925 is able to chelate Fe2+ , Fe3+ , Cu+ a Cu2+ ions at physiologically and pathophysiologically relevant pH values (4,5; 5,5; 6,8 and 7,5) and...

La contamination simultanée des écosystèmes aquatiques par des polluants et des nutriments est un problème majeur dont les effets sur les macrophytes sont encore méconnus. L'objectif de cette thèse était d'étudier les effets de deux types de polluants : herbicides et éléments trace métalliques (ETM), en combinaison avec des ressources variables (azote, phosphore, ou carbone) sur Myriophyllum spicatum, en utilisant des paramètres provenant de l'écologie chimique et de l'écotoxicologie. Les tests ont montré des effets contrastés des herbicides sur la plante, mais pas d'effet des ETM. Dans tous les cas, les modifications de ressources (nitrate, ratio N:P ou saccharose) ont induit des effets importants sur la physiologie et la stœchiométrie de la plante, pouvant altérer ses réponses à un stress supplémentaire comme les polluants. Ces résultats suggèrent qu'un déséquilibre des ressources en présence de polluants pourrait modifier leurs effets sur les macrophytes, et soulignent également la nécessité d'ajouter des paramètres plus informatifs dans les tests afin de répondre aux enjeux du stress multiple et améliorer l'évaluation du risque environnemental / Simultaneous contamination of aquatic ecosystems by pollutants and nutrients is a major problem whose effects on macrophytes are still unknown. The objective of this thesis was to study the effects of different pollutants, herbicides and trace metal elements (TME), in combination with varying resource availability (nitrogen, phosphorus, or carbon) on Myriophyllum spicatum, using parameters from chemical ecology and ecotoxicology. Tests showed contrasting effects of herbicides, but no effects of the TME. In all cases, resource modifications (nitrate, N:P ratio or sucrose) induced significant effects on the physiology and stoichiometry of the plant, which might alter its response to additional stress like pollutants. These results suggest that an imbalance of resources in the presence of pollutants may lead to unforeseen changes in the combined effects on macrophytes, and also highlight the need to add more informative parameters in tests to meet the challenges of multiple stress and improve environmental risk assessment.

É sabido que o ensino de ciências no Brasil está entre os piores do mundo. Atualmente, a sociedade discute com bastante frequência este quadro: pensam-se as razões que levam a educação a estar tão defasada, e o que leva os estudantes a não gostarem da disciplina de Química. Uma dessas razões pode ser a forma como os conteúdos são passados para os discentes, a qual muitas vezes é descontextualizada e nada atrativa. As histórias em quadrinhos têm sido utilizadas como instrumento didático de aprendizagem em diferentes áreas no ensino médio, no entanto na área da química, ainda são pouco usadas. Baseado nisso o presente trabalho tem como objetivo desenvolver um estudo de caso elaborando uma revista em quadrinhos para explicar o conteúdo de estequiometria a alunos do segundo ano do ensino médio. A hipótese é de que a revista em quadrinhos, por ter um caráter lúdico, possa facilitar a aprendizagem do conteúdo. Primeiramente foi criada a revista, depois foi realizado um estudo de caso em uma escola estadual em Gravataí/RS, onde participaram da atividade utilizando a revista, 60 alunos de três turmas do segundo ano do ensino médio. A avaliação foi realizada por meio de prétestes e pós-testes e com o uso de questionários com perguntas abertas e fechadas, sendo que as respostas foram categorizadas sob um enfoque quantitativo. Os resultados encontrados mostraram que os estudantes ficaram motivados com a atividade e tiveram um maior entendimento do conteúdo quando utilizaram a revista em quadrinhos, sendo que 26% dos estudantes melhoraram suas respostas em frente ao conteúdo abordado. Além disso, observou-se que cerca de 25% compreenderam melhor os conceitos e os cálculos estequiométricos. Sendo assim, acredita-se que a revista em quadrinhos possa ser uma boa estratégia de ensinoaprendizagem para o ensino de Química. / It is a known fact that science education in Brazil is among the worst in the world. Nowadays, society often discusses this situation and considers the reasons why education is so outdated and why students do not like the subject of chemistry. One reason may be the method used to teach the subject matter, which is often decontextualized and unattractive. The hypothesis was that the comic book, due to its ludic character, might facilitate the understanding of this subject. First the comic book was created; then a case study was done at a state school in Gravataí/RS, where 60 students from three groups in the second year of high school participated in the activity. The evaluation was done through pretests and posttests and using questionnaires with open- and closed-ended questions, classifying the answers with a quantitative focus. The results showed that the students were motivated by the activity and had a better understanding of the subject when they used the comic book, with 26% of the students improving their answers about the subject matter. It was also observed that 25% of the students better understood the concepts and the stoichiometric calculus. Thus, we believe that comic books may be a good teachinglearning strategy for the study of chemistry.

abstract: Primary production in aquatic ecosystems is often limited by the availability of nitrogen (N) and/or phosphorus (P). Animals can substantially alter the relative availability of these nutrients by storing and recycling them in differential ratios. Variation in these stoichiometric traits, i.e., the elemental phenotype, within a species can link organismal evolution to ecosystem function. I examined the drivers of intraspecific variation in the elemental phenotype of aquatic consumers to test for the generality of these effects. Over a thermal gradient in Panamá, I found that average specific growth grate and body P content of the mayfly Thraulodes increased with environmental temperature, but that these patterns were due to site-specific differences rather than the direct effects of warmer temperature. In a meta-analysis of published studies, I found that in fishes intraspecific variation in dietary N:P ratio had a significant effect on excretion N:P ratio, but only when accounting for consumption. I tested for the effects of variation in consumption on excretion N:P ratio among populations of the fish Gambusia marshi in the Cuatro Ciénegas basin in Coahuila, Mexico. G. marshi inhabits warm groundwater-fed springs where it often co-occurs with predatory fishes and cool runoff-dominated wetlands which lack predators. Using stoichiometric models, I generated predictions for how variation in environmental temperature and predation pressure would affect the N:P ratio recycled by fishes. Adult female G. marshi excretion N:P ratio was higher in runoff-dominated sites, which was consistent with predators driving increased consumption rates by G. marshi. This result was supported by a diet ration manipulation experiment in which G. marshi raised on an ad libitum diet excreted N:P at a lower ratio than fish raised on a restricted diet ration. To further support the impacts of predation on phenotypic diversification in G. marshi, I examined how body morphology varied among habitats and among closely related species. Both among and within species, predation had stronger effects on morphology than the physical environment. Overall, these results suggest that predation, not temperature, has strong effects on these phenotypic traits of aquatic consumers which can alter their role in ecosystem nutrient cycling through variation in consumption rates. / Dissertation/Thesis / Doctoral Dissertation Biology 2017

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Thesis (DTech (Chemical Engineering))--Cape Peninsula University of Technology, 2017. / The high rate of industrialisation in most developing countries has brought about challenges of wastewater management especially in the mineral processing industry. Cyanide has been used in base metal extraction processes due to its lixiviant properties thus, its presence in wastewater generated is inevitable. Furthermore, partial and/or the use of unsuitable treatment methods for such wastewater is a potential hazard to both human and the environment. There are several reports on biotechnological treatments of cyanide containing wastewater but few mineral processing industries have adopted this approach. Hence, the thermodynamic study of biodegradation of cyanide containing wastewater was undertaken. The primary aim of this study was to explore the application of bioenergetic models and biological stoichiometry to determine the functionality and thermodynamic requirements for cyanide degrading isolate (Fusarium oxysporum EKT01/02), grown exclusively on Beta vulgaris, for a system designed for the bioremediation of cyanidation wastewater. Chapter 2 reviews some of the applicable thermodynamic parameters such as enthalpy, entropy, heat of combustion, heat capacity, Gibbs energy, including stoichiometry models in relation to their applicability for microbial proliferation in cyanidation wastewater. The chapter places emphasis on the application of agro-industrial waste as a suitable replacement for refined carbon sources for microbial proliferation in bioremediation systems because such systems are environmentally benign. The choice of using agro-industrial waste is due to organic waste properties, i.e. agro-industrial waste is rich in nutrients and is generated in large quantities. Chapter 3 presents the materials and various standardised methods used to address the research gaps identified in chapter 2. For an organism to degrade free cyanide in wastewater, it must be able to survive and perform its primary function in the presence of such a toxicant. Chapter 4 exemplifies both molecular and biochemical characteristics of Fusarium oxysporum EKT01/02 isolated from the rhizosphere of Zea mays contaminated with a cyanide based pesticide. The molecular analyses confirmed the fungal isolate to be Fusarium oxysporum EKT01/02 and the nucleotide sequence of the isolates were deposited with National Centre for Biotechnology Information (NCBI) with accession numbers KU985430 and KU985431. The biochemical analyses revealed a wide substrate utilisation mechanism of the isolate dominated by aminopeptidase including nitrate assimilation capabilities. A preliminary investigation showed free cyanide degradation efficiency of 77.6% (100 mg CN-/L) after 5 days by the isolate. The excess production of extracellular polymeric substance (EPS) was attributed to the isolates’ strive to protect itself from cyanide toxicity.

É sabido que o ensino de ciências no Brasil está entre os piores do mundo. Atualmente, a sociedade discute com bastante frequência este quadro: pensam-se as razões que levam a educação a estar tão defasada, e o que leva os estudantes a não gostarem da disciplina de Química. Uma dessas razões pode ser a forma como os conteúdos são passados para os discentes, a qual muitas vezes é descontextualizada e nada atrativa. As histórias em quadrinhos têm sido utilizadas como instrumento didático de aprendizagem em diferentes áreas no ensino médio, no entanto na área da química, ainda são pouco usadas. Baseado nisso o presente trabalho tem como objetivo desenvolver um estudo de caso elaborando uma revista em quadrinhos para explicar o conteúdo de estequiometria a alunos do segundo ano do ensino médio. A hipótese é de que a revista em quadrinhos, por ter um caráter lúdico, possa facilitar a aprendizagem do conteúdo. Primeiramente foi criada a revista, depois foi realizado um estudo de caso em uma escola estadual em Gravataí/RS, onde participaram da atividade utilizando a revista, 60 alunos de três turmas do segundo ano do ensino médio. A avaliação foi realizada por meio de prétestes e pós-testes e com o uso de questionários com perguntas abertas e fechadas, sendo que as respostas foram categorizadas sob um enfoque quantitativo. Os resultados encontrados mostraram que os estudantes ficaram motivados com a atividade e tiveram um maior entendimento do conteúdo quando utilizaram a revista em quadrinhos, sendo que 26% dos estudantes melhoraram suas respostas em frente ao conteúdo abordado. Além disso, observou-se que cerca de 25% compreenderam melhor os conceitos e os cálculos estequiométricos. Sendo assim, acredita-se que a revista em quadrinhos possa ser uma boa estratégia de ensinoaprendizagem para o ensino de Química. / It is a known fact that science education in Brazil is among the worst in the world. Nowadays, society often discusses this situation and considers the reasons why education is so outdated and why students do not like the subject of chemistry. One reason may be the method used to teach the subject matter, which is often decontextualized and unattractive. The hypothesis was that the comic book, due to its ludic character, might facilitate the understanding of this subject. First the comic book was created; then a case study was done at a state school in Gravataí/RS, where 60 students from three groups in the second year of high school participated in the activity. The evaluation was done through pretests and posttests and using questionnaires with open- and closed-ended questions, classifying the answers with a quantitative focus. The results showed that the students were motivated by the activity and had a better understanding of the subject when they used the comic book, with 26% of the students improving their answers about the subject matter. It was also observed that 25% of the students better understood the concepts and the stoichiometric calculus. Thus, we believe that comic books may be a good teachinglearning strategy for the study of chemistry.