Persistent organic pollutants (POPs) and adipose tissue hypoxia have been shown to independently affect adipocyte functions. The goals of this study were to (1) determine the effect of PCB-77, PCB-153, and DDE on the differentiation of human preadipocytes, and (2) investigate the cross-talk between PCB-77 and hypoxia in differentiated human adipocytes. First, human preadipocytes were exposed to PCB-77, PCB-153, or DDE during the entire 14-day differentiation period. We found no effect of low POP levels on lipid accumulation. Second, differentiated human adipocytes were exposed to a combination of PCB-77 and hypoxia. We demonstrated gene-specific cross-talk between PCB-77 and hypoxia, showing an additive effect of PCB-77 on VEGF, MCP-1, and adiponectin, as well as an inhibition of PCB-77-induced expression of CYP1A1 by hypoxia. This work has expanded our understanding of the role of POPs and hypoxia in differentiated human adipocytes.

La modélisation de la migration des hydrocarbures dans les bassins sédimentaires a pour but d'évaluer leur potentiel pétrolier, en localisant et en quantifiant les accumulations d'hydrocarbures au sein des formations géologiques. Dans cette thèse, nous étudions les modèles de migration de type "Darcy" ainsi que des modèles simplifiés de types "ray-tracing" et "invasion percolation"; l'objectif est de mener une analyse critique et de proposer des améliorations tout en fournissant un guide pour une utilisation pertinente sur des cas d'étude.Tout d'abord, nous faisons une revue des mécanismes de la migration depuis l'échelle des pores jusqu'à l'échelle des bassins, puis nous présentons chacun des modèles.Dans le volet suivant, nous proposons deux algorithmes d'invasion percolation : le premier, adapté aux maillages structurés; le second, permettant de mieux prendre en compte les maillages non structurés. Dans un troisième volet, nous nous intéressons à la comparaison entre ces modèles, en nous concentrant sur ceux de types "Darcy" et "invasion percolation". Nous nous focalisons en premier lieu sur les aspects numériques en nous appuyant sur plusieurs cas tests; puis nous effectuons une comparaison formelle en étudiant la limite asymptotique de la solution du modèle de type "Darcy" en temps long. Nous présentons ensuite une série d'applications dont notamment l'étude d'un cas réel 3D en géométrie complexe.Finalement, nous concluons ce travail avec deux articles. Le premier montre une évolution des modèles de type "Darcy" en utilisant la méthode du raffinement local de maillage, avec une illustration sur un cas d'étude du nord du Koweït. Le deuxième synthétise les principaux résultats obtenus concernant les méthodes de "Darcy" et "d'invasion percolation". / Hydrocarbon migration modeling in sedimentary basins aims to localize and to quantify hydrocarbon accumulations in geological formations in order to estimate their petroleum potential. In this thesis, we study “Darcy” migration models and also simplified migration models such as “ray-tracing” and “invasion percolation”; the purpose is to conduct a critical analysis and to offer improvements while providing a guide for a relevant use on case studies.We start by a review of migration mechanisms from the pore scale to the basin scale, then we present each model.In a following part, we propose two invasion percolation algorithms: the first one is suited to structured grids, the second one allows to take better account of unstructured grids.In a third part, we take an interest in the comparison between the different models and particularly between “Darcy” and “invasion percolation” approaches. First we devote our attention to numerical aspects supported by several use cases; then we realize a formal comparison by studying the asymptotic limit of the “Darcy” model large time solution. Afterwards, we present several applications including the study of a 3D real case in complex geometry.Finally, we conclude this work with two articles. The first one shows an evolution of “Darcy” models by using the method of local grid refinement with an illustration on a case study from northern Kuwait. The second one synthesizes the main results on “Darcy” and “invasion percolation” methods.

Las celdas de combustible de óxido sólido (cuyo acrónimo en inglés es SOFC) son dispositivos energéticos capaces de convertir la energía química de un combustible directamente en energía eléctrica. Esto las dota de unas eficiencias eléctricas muy elevadas, que pueden llegar a ser del 80% si se aprovecha su calor residual de alta calidad mediante turbinas. Además, son capaces de funcionar con una gran variedad de combustibles: hidrógeno, gas natural, gas de síntesis, etanol, metanol, etc. Sin embargo, para su inserción en la cadena de producción energética, su temperatura de funcionamiento debería disminuir al rango de 500-700 ºC sin que se redujeran las densidades de potencias eléctricas generadas. Las SOFC convencionales se basan en la conducción de iones oxígeno de su electrolito, que separa la reacción de combustión del combustible en sus semi-reacciones electroquímicas, generando de este modo la energía eléctrica directamente. Al disminuir la temperatura de operación en este tipo de SOFC, con electrolitos (o membranas) delgados e hidrógeno como combustible, la principal limitación de funcionamiento se centra en la activación y reducción del oxígeno que tiene lugar en el electrodo denominado cátodo. Por otro lado, el empleo de otros combustibles basados en carbono no es compatible con los materiales de ánodos actualmente utilizados. Por tanto, es necesario el desarrollo de nuevos cátodos con mejoradas propiedades electrocatalíticas para la reducción de oxígeno a menores temperaturas, cuyas propiedades termo-mecánicas sean compatibles con las del resto de componentes de la celda, y la obtención de ánodos capaces de funcionar con combustibles basados en carbono. La combinación conjunta de varios lantánidos y bario en la estructura perovskita (LalPrpSmsBab)0.58Sr0.4Fe0.8Co0.2O3 ha permitido obtener compuestos con resistencias de polarización de electrodo significantemente menores que las mostradas por el cátodo del estado de la técnica La0.6Sr0.4Fe0.8Co0.2O3 en el rango de temperaturas 450-650 ºC. / Vert Belenguer, VB. (2011). ELECTRODOS AVANZADOS PARA PILAS DE COMBUSTIBLE DE ÓXIDO SÓLIDO (SOFCs) [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/14669 / Palancia

The aryl hydrocarbon receptor (AHR) is a ligand-activated signaling molecule which controls tumor growth and metastasis, T cell differentiation, and liver development. Expression levels of this receptor protein are sensitive to the cellular p23 protein levels in immortalized cancer cell lines. As little as 30% reduction of the p23 cellular content can suppress the AHR function. Here we reported that down-regulation of the p23 protein content in normal, untransformed human bronchial/tracheal epithelial cells to 48% of its content also suppresses the AHR protein levels to 54% of its content. This p23-mediated suppression of AHR is responsible for the repression of (1) the ligand-dependent induction of the cyp1a1 gene transcription; (2) the benzo[a]pyrene- or cigarette smoke condensate-induced CYP1A1 enzyme activity, and (3) the benzo[a]pyrene and cigarette smoke condensate-mediated production of reactive oxygen species. Reduction of the p23 content does not alter expression of oxidative stress genes or production of PGE2. Down-regulation of p23 suppresses the AHR protein levels in two other untransformed cell types, namely human breast MCF-10A and mouse immune regulatory Tr1 cells. Collectively, down-regulation of p23 suppresses the AHR protein levels in normal and untransformed cells and can in principle protect our lung epithelial cells from AHR-dependent oxidative damage caused by exposure to agents from environment and cigarette smoking. The AHR is expressed in triple-negative and non-triple-negative breast cancer cells. It affects breast cancer growth and crosstalk with the estrogen receptor signaling. Normally the AHR is degraded shortly after ligand activation via the action of 26S proteasome. Here we report that the piperazinylpyrimidine compound Q18 triggers AHR protein degradation which is mediated through chaperone-mediated autophagy in triple-negative breast cancer cells (MDA-MB-468 and MDA-MB-231). This lysosomal degradation of AHR exhibits the following characteristics: (1) not observed in non-triple-negative breast cancer cells (MCF-7, T47D, and MDA-MB-361); (2) inhibited by progesterone receptor B but not estrogen receptor alpha; (3) reversed by chloroquine but not MG132; (4) required LAMP2A; (5) triggered by 6 amino-nicotinamide and starvation and (6) involved AHR-LAMP2A interaction mediated by 6 amino-nicotinamide and starvation. The NEKFF sequence localized at amino acid 558 of human AHR is a KFERQ-like motif of chaperone-mediated autophagy, essential for the LAMP2A-mediated AHR protein degradation.

The aryl hydrocarbon receptor (AHR) is a transcription factor which heterodimerizes with the aryl hydrocarbon receptor nuclear translocator (Arnt) to regulate downstream gene transcription. For the purpose of studying the crystal structure of human aryl hydrocarbon receptor (hAHR), it is essential to obtain abundant amount of pure recombinant protein.Basing on the benefits of using P. pastoris system to produce recombinant protein, including appropriate folding, secretion of interest proteins to the external environment of the cell, and easier purification process of protein due to the its limited production of endogenous secretory proteins [1], our lab chose P. pastoris yeast as the host to overexpress human AHR. My lab has successfully used the protease-deficient P. pastoris (ySMD1163) strain to express AHR [2], but unfortunately the yield is modest, presumably due to low copy number. My work addressed whether increasing the copy number of hAHR in the yeast genome would increase the expression level of hAHR in Pichia pastoris. Results from my experiments showed that although the copy number correlated with the expression levels of hAHR, the increased expression of the hAHR largely in the pellet, suggesting that the soluble expression of hAHR can’t be enhanced merely by increasing its production.

A new free radical chain process for the allylation of hydrocarbons and some other substrates utilizing substituted allyl bromides (R-H + C=C-C-Br -> R-C-C=C + HBr) has been developed. Good to excellent yields were observed in all cases. Kinetic chain measurements and competition experiments were performed in order to elucidate the mechanism of the reaction. Overall, the results are consistent with a free radical chain process with bromine atom as the chain carrier. Substitution effects on the reactivity of the allyl bromides (CH2=C(Z)CH2Br) and their influence on the overall reaction rate were studied by conducting several competition experiments. The relative rate constants for addition of benzyl radical to CH2=C(Z)CH2Br are: Z=CN(180), COOEt(110), Ph(65), H(1.0). The trend of electronegativity/reactivity of these reactions was very similar to that reported for addition of benzyl radical to substituted alkenes. Other than alkyl aromatics (PhCH3, PhCH(CH3)2), other substrates (i.e., 2- propanol, phenyl cyclopropane) were also tested for this allylation reaction. The magnitude and scope of these reactions, and their synthetic utility is discussed. / Ph. D.

The aryl hydrocarbon receptor nuclear translocator (ARNT) is a promiscuous protein serving as a required dimerization partner for the aryl hydrocarbon receptor (AhR) and hypoxia inducible factor-1α (HIF-1α) transcription factors. Additionally it serves as a potent co-activator for estrogen receptor (ER) signaling. We sought to take advantage of these cross-talk mechanisms by designing an AhR construct that can influence the regulation of these pathways by sequestering ARNT. CΔ553 is a truncated form of the AhR lacking the C-terminal 553 amino acids which harbors the complete transactivation (TAD) and significant portions of the ligand binding (LBD) domains. Altering the LBD allows CΔ553 to become constitutively active and has been shown to associate with ARNT and bind DNA. Without the TAD, CΔ553 cannot recruit co-activators to the promoter so that no activation of gene transcription may occur. Transient transfection studies using a corresponding luciferase reporter plasmid in MCF-7 cells showed that CΔ553 effectively suppressed the AhR, HIF-1α, and ER signaling pathways. RT/real-time QPCR data showed that CΔ553 blocked the up-regulation of the target genes controlled by AhR ( CYP1A1 ), HIF-1α ( VEGF, aldolase C , and LDH-A ), and ER ( GREB1 ) in breast cancer cells. Since both HIF-1α and ER are highly active in ER-positive breast cancers, CΔ553 has the potential to be developed as a protein drug to treat breast cancer by blocking these two signaling pathways. Seeking to determine if complete suppression of genes is possible with CΔ553, a tetracycline regulated retroviral expression system is investigated along with the possibilities for cellular administration via the HIV-1 Tat protein transduction domain. Since ARNT dimerizes with both AhR and HIF-1α, we present further studies looking into the role protein factors play in the activation of each system. Previously within our lab p23 and Cyp40, two components of the hsp90 chaperon complex, were found to facilitate the formation of the AhR•ARNT•DNA binding complex. Analysis of these proteins within the hypoxia signaling pathway found that only p23 was capable of generating the HIF-1α•ARNT•DNA binding complex.

No description available.

A new material for use as a proton exchange membrane in fuel cells has been developed: a blend of a perfluorocyclobutane-based block ionomer (S-PFCB) and Poly (vinylidene-co-hexafluoropropylene) (Kynar Flex, KF). This thesis details the work done thus far to characterize the morphology of this material, using small angle x-ray scattering, differential scanning calorimetry, atomic force micrscopy, and some other techniques to a lesser extent. Small angle x-ray scattering (SAXS) of pure S-PFCB showed a strong block copolymer- associated phase separation, on the order of 25 nm. Differential scanning Calorimetry (DSC) confirmed this finding. SAXS also revealed the presence of a peak representing individual ionic aggregates on the order of 3 nm. Finally, it was shown with DSC that no crystallinity develops in the S-PFCB block copolymer, while one of the blocks, known as 6F, crystallizes extensively. SAXS of incremental blend compositions of KF and S-PFCB revealed a steady increase in size of the block copolymer phase separation peak in SAXS, demonstrative of the miscibility of KF and the non-sulfonated 6F block of S-PFCB. Furthermore, this incremental study determined the scattering vector range relevant for comparing amounts of KF crystallinity. DSC of incremental blend compositions revealed two phases of KF crystallinity develops upon cooling a membrane, independent of cooling rate. Atomic force microscopy (AFM), small angle x-ray scattering (SAXS), and differential scanning calorimetry (DSC) corroborate to suggest a nonuniform morphology through the thickness of solution cast membranes. Also, the effect of different casting temperatures and after-casting anneals on morphology was assessed. Future work on this project involves morphological studies at various relative humidities and temperatures, as well as following up on discoveries already made. Finally, transmission electron micrscopy (TEM) should be performed to provide a visual analog, which will greatly help in developing an accurate morphological model. / Master of Science

Dioxins and several halogenated polycyclic aromatic hydrocarbons belong to a class of toxic environmental pollutants that give rise to a myriad of teratogenic and carcinogenic responses and are of major concern from a human health perspective due to their widespread distribution. Apart from an array of toxic endpoints, they affect the expression of a variety of xenobiotic metabolizing enzymes including CYP1A1 and 1A2. Data generated by rodent studies have shown that most, if not all, of their biological and toxic effects are mediated through binding to the aryl hydrocarbon receptor (AhR). Upon ligand binding, AhR translocates into the nucleus and heterodimerizes with AhR-nuclear translocator (Arnt); the heterodimer binds to the dioxin response element (DRE) located upstream to the promoter region of target genes, leading to their transcription. The AhR/Arnt/DRE complex has been well characterized and can be observed readily by the gel shift assay. However, the mechanism for this AhR complex formation is unclear. Baculovirus expressed, metal resin-purified human AhR and Arnt are unable to bind the DRE in a ligand-dependent manner unless crude extracts, such as the rabbit reticulocyte lysate (RRL), are reconstituted with these proteins. Proteins in the RRL are responsible for this restoration of the gel shift complex because the activity is sensitive to both heat and proteolytic treatments. Fractionation of the RRL using centricon devices gave the enriched activity in the C10 retentate fraction (C10R). Screening gel shift assays and immunodepletion studies showed that p23 and CyP40, but not hsp90 and hsp70, could be the protein factors. Purified bacterial expressed p23 restored the gel shift complex; and the mechanism is mediated at the heterodimerization step and is hsp90-dependent. However, p23 is not the major factor since the same amount of C10R as that of purified p23 produced a much more pronounced gel shift activity and was insensitive to geldanamycin and apyrase. CyP40 is unable to restore the complex formation directly; however, our data suggested that some of the CyP40-interacting proteins restore the AhR/Arnt/DRE complex formation.