Molecular mobility of n-ethylene glycol dimethacrylate glass formers upon free radical polymerization

Quando um líquido evita a cristalização durante o arrefecimento, diz-se que entra no estado sobrearrefecido. Se a temperatura continuar a diminuir, o consequente aumento da viscosidade reflecte-se na mobilidade molecular de tal maneira que os tempos característicos se tornam da mesma ordem de grandeza que os tempos acessíveis experimentalmente. Se o arrefecimento continuar, o líquido altamente viscoso acaba por vitrificar, i.e. entra no estado vítreo onde apenas os movimentos locais são permitidos. Os monómeros da família n -etileno glicol dimetacrilato ( n -EGDMA, para n = 1 até 4, que constituem o objecto deste estudo, facilmente evitam a cristalização, sendo pois bons candidatos para estudar a mobilidade molecular nos estados sobrearrefecido e vítreo. A Espectroscopia de Relaxação Dieléctrica (DRS) foi a técnica escolhida para obter informação detalhada sobre a sua dinâmica molecular (Capítulos 1 e 2). A primeira parte deste trabalho consistiu na caracterização dieléctrica dos processos de relaxação existentes acima e abaixo da temperatura de transição vítrea (g T ), a qual aumenta com o aumento do peso molecular (w M ), sendo este resultado confirmado por Calorimetria Diferencial de Varrimento (DSC). No que respeita ao processo cooperativo a , associado à transição vítrea, e ao processo secundário b, observa-se uma dependência com w M , enquanto que o outro processo secundário, g , aparenta ser independente deste factor (Capítulo 3). Nos capítulos seguintes, foram levadas a cabo diferentes estratégias com o objectivo de clarificar os mecanismos que estão na origem destas duas relaxações secundárias (b e g ), assim como conhecer a sua respectiva relação com a relaxação principal (a ). Do estudo, em tempo real, da polimerização isotérmica via radicais livres do TrEGDMA por Calorimetria de Varrimento Diferencial com Modulação de Temperatura (TMDSC), levado a cabo a temperaturas abaixo da g T do polímero final, concluem-se entre outros, dois importantes aspectos: i) que a vitrificação do polímero em formação conduz a graus de conversão relativamente baixos, e ii) que o monómero que está por reagir é expulso da rede polimérica que se forma, dando lugar a uma clara separação de fases (Capítulo 4). Com base nesta informação, o passo seguinte foi estudar separadamente a polimerização isotérmica do di-, tri- e tetra-EGDMA, dando especial atenção às alterações de mobilidade do monómero ainda por reagir. Com as restrições impostas pela formação de ligações químicas, as relaxações a e b detectadas no monómero tendem a desaparecer no novo polímero formado, enquanto que a relaxação g se mantém quase inalterada. Os diferentes comportamentos que aparecem durante a polimerização permitiram a atribuição da origem molecular dos processos secundários: o processo g foi associado ao movimento twisting das unidades etileno glicol, enquanto que a rotação dos grupos carboxilo foi relacionada com a relaxação b (Capítulo 5). No que respeita ao próprio polímero, um processo de relaxação adicional foi detectado, pol b , no poly-DEGDMA, poly-TrEGDMA e poly-TeEGDMA, com características similares ao encontrado nos poli(metacrilato de n -alquilo). Este processo foi confirmado e bem caracterizado aquando do estudo da copolimerização do TrEGDMA com acrilato de metilo (MA) para diferentes composições (Capítulo 6). Para finalizar, o EGDMA, o elemento mais pequeno da família de monómeros estudada, além de vitrificar apresenta uma marcada tendência para cristalizar quer a partir do estado líquido ou do estado vítreo. Durante a cristalização, a formação de uma fase rígida afecta principalmente o processo a , cuja intensidade diminui sem no entanto se observarem modificações significativas na dependência do tempo de relaxação característico com a temperatura. Por outro lado, o processo secundário b torna-se melhor definido e mais estreito, o que pode ser interpretado em termos de uma maior homogeneidade dos micro-ambientes associados aos movimentos locais(Capítulo 7).

Futures of automobile industry and challenges on sustainable development and mobility

Portugal had only very few foresight exercises on the automobile sector, and the most recent one was a survey held in a project on work organisation systems in the automobile industry, its recent historical paths and the special strategies of location of companies (the WorTiS project). This involved several teams with different disciplinary backgrounds and from two Portuguese universities. The provisional main results of the first round of a Delphi survey held in Portugal on the automotive sector were already published, but a further analysis was not yet done. This foresight survey was done under the WorTiS project, developed in 2004 by IET – Research Centre on Enterprise and Work Innovation (at FCT-UNL), and financed by the Portuguese Ministry of Science and Technology. Some of this experience on foresight analysis is also been transferred to other projects, namely the WORKS project on work organisation restructuring in the knowledge society that received the support from EC and still is running. The majority of experts considered having an average of less knowledge in almost all the scenario topics presented. This means that information on the automotive industry is not spread enough among academics or experts in related fields (regional scientists, innovation economists, engineers, sociologists). Some have a good knowledge but in very specialised fields. Others have expertise on foresight, or macroeconomics, or management sciences, but feel insecure on issues related with futures of automobile sector. Nevertheless, we considered specially the topics where the experts considered themselves to have some knowledge. There were no “irrelevant” topics considered as such by the expert panel. There are also no topics that are not considered a need for co-operation. The lack of technological infrastructures was not considered as a hindered factor for the accomplishment of any scenario. The experts’ panel considered no other international competence besides US, Japan or Germany in these topics. Special focus will be made in this paper on the topic 2. Public policy and automobile industries, and more specifically on the technological and/or research policies issues, where one can specify the automobile’s role in transport policies with further implications like environment, safety, energy, mobility.

Strategic analysis on urban mobility

A Work Project, presented as part of the requirements for the Award of a Masters Degree in Management from the NOVA – School of Business and Economics

Agenesis of human pancreas due to decreased half-life of insulin promoter factor 1.

Neonatal diabetes mellitus can be transient or permanent. The severe form of permanent neonatal diabetes mellitus can be associated with pancreas agenesis. Normal pancreas development is controlled by a cascade of transcription factors, where insulin promoter factor 1 (IPF1) plays a crucial role. Here, we describe two novel mutations in the IPF1 gene leading to pancreas agenesis. Direct sequence analysis of exons 1 and 2 of the IPF1 gene revealed two point mutations within the homeobox in exon 2. Genetic analysis of the parents showed that each mutation was inherited from one parent. Mutations localized in helices 1 and 2, respectively, of the homeodomain, decreased the protein half-life significantly, leading to intracellular IPF1 levels of 36% and 27% of wild-type levels. Both mutant forms of IPF1 were normally translocated to the nucleus, and their DNA binding activity on different known target promoters was similar to that of the wild-type protein. However, transcriptional activity of both mutant IPF1 proteins, alone or in combination with HNF3 beta/Foxa2, Pbx1, or the heterodimer E47-beta 2 was reduced, findings accounted for by decreased IPF1 steady state levels and not by impaired protein-protein interactions. We conclude that the IPF1 level is critical for human pancreas formation.

Involvement of nuclear factor-kappaB in macrophage migration inhibitory factor gene transcription up-regulation induced by interleukin- 1 beta in ectopic endometrial cells.

OBJECTIVE: To investigate the involvement of the nuclear factor (NF)-kappaB in the interleukin (IL)-1 beta-mediated macrophage migration inhibitory factor (MIF) gene activation. DESIGN: Prospective study. SETTING: Human reproduction research laboratory. PATIENT(S): Nine women with endometriotic lesions. INTERVENTION(S): Endometriotic lesions were obtained during laparoscopic surgery. MAIN OUTCOME MEASURE(S): The MIF protein secretion was analyzed by ELISA, MIF mRNA expression by quantitative real-time polymerase chain reaction (PCR), NF-kappaB translocation into the nucleus by electrophoresis mobility shift assay, I kappaB phosphorylation and degradation by Western blot, and human MIF promoter activity by transient cell transfection. RESULT(S): This study showed a significant dose-dependent increase of MIF protein secretion and mRNA expression, the NF-kappaB translocation into the nucleus, I kappaB phosphorylation, I kappaB degradation, and human MIF promoter activity in endometriotic stromal cells in response to IL-1 beta. Curcumin (NF-kappaB inhibitor) significantly inhibited all these IL-1 beta-mediated effects. Analysis of the activity of deletion constructs of the human MIF promoter and a computer search localized two putative regulatory elements corresponding to NF-kappaB binding sites at positions -2538/-2528 bp and -1389/-1380 bp. CONCLUSION(S): This study suggests the involvement of the nuclear transcription factor NF-kappaB in MIF gene activation in ectopic endometrial cells in response to IL-1 beta and identifies a possible pathway of endometriosis-associated inflammation and ectopic cell growth.

Neuropeptide-inducible upregulation of proteasome activity precedes nuclear factor kappa B activation in androgen-independent prostate cancer cells.

ABSTRACT: BACKGROUND: Upregulation of nuclear factor kappa B (NFκB) activity and neuroendocrine differentiation are two mechanisms known to be involved in prostate cancer (PC) progression to castration resistance. We have observed that major components of these pathways, including NFκB, proteasome, neutral endopeptidase (NEP) and endothelin 1 (ET-1), exhibit an inverse and mirror image pattern in androgen-dependent (AD) and -independent (AI) states in vitro. METHODS: We have now investigated for evidence of a direct mechanistic connection between these pathways with the use of immunocytochemistry (ICC), western blot analysis, electrophoretic mobility shift assay (EMSA) and proteasome activity assessment. RESULTS: Neuropeptide (NP) stimulation induced nuclear translocation of NFκB in a dose-dependent manner in AI cells, also evident as reduced total inhibitor κB (IκB) levels and increased DNA binding in EMSA. These effects were preceded by increased 20 S proteasome activity at lower doses and at earlier times and were at least partially reversed under conditions of NP deprivation induced by specific NP receptor inhibitors, as well as NFκB, IκB kinase (IKK) and proteasome inhibitors. AD cells showed no appreciable nuclear translocation upon NP stimulation, with less intense DNA binding signal on EMSA. CONCLUSIONS: Our results support evidence for a direct mechanistic connection between the NPs and NFκB/proteasome signaling pathways, with a distinct NP-induced profile in the more aggressive AI cancer state.

Regulation of gammadelta versus alphabeta T lymphocyte differentiation by the transcription factor SOX13.

alphabeta and gammadelta T cells originate from a common, multipotential precursor population in the thymus, but the molecular mechanisms regulating this lineage-fate decision are unknown. We have identified Sox13 as a gammadelta-specific gene in the immune system. Using Sox13 transgenic mice, we showed that this transcription factor promotes gammadelta T cell development while opposing alphabeta T cell differentiation. Conversely, mice deficient in Sox13 expression exhibited impaired development of gammadelta T cells but not alphabeta T cells. One mechanism of SOX13 function is the inhibition of signaling by the developmentally important Wnt/T cell factor (TCF) pathway. Our data thus reveal a dominant pathway regulating the developmental fate of these two lineages of T lymphocytes.

Association with coregulators is the major determinant governing peroxisome proliferator-activated receptor mobility in living cells.

The nucleus is an extremely dynamic compartment, and protein mobility represents a key factor in transcriptional regulation. We showed in a previous study that the diffusion of peroxisome proliferator-activated receptors (PPARs), a family of nuclear receptors regulating major cellular and metabolic functions, is modulated by ligand binding. In this study, we combine fluorescence correlation spectroscopy, dual color fluorescence cross-correlation microscopy, and fluorescence resonance energy transfer to dissect the molecular mechanisms controlling PPAR mobility and transcriptional activity in living cells. First, we bring new evidence that in vivo a high percentage of PPARs and retinoid X receptors is associated even in the absence of ligand. Second, we demonstrate that coregulator recruitment (and not DNA binding) plays a crucial role in receptor mobility, suggesting that transcriptional complexes are formed prior to promoter binding. In addition, association with coactivators in the absence of a ligand in living cells, both through the N-terminal AB domain and the AF-2 function of the ligand binding domain, provides a molecular basis to explain PPAR constitutive activity.

Transcriptional activation of the GLUT2 gene by the IPF-1/STF-1/IDX-1 homeobox factor.

The homeodomain protein PDX-1, referred as IPF-1/STF-1/IDX-1, is a transcriptional factor that plays a critical role in the control of several genes expressed in the pancreatic islet. PDX-1 gene expression has been previously shown to be reduced in cultured beta-cell lines chronically exposed to high glucose concentrations. As the glucose transporter type 2 (GLUT2) gene expression is selectively decreased in the beta-pancreatic cells of experimental models of diabetes, we postulated that the loss of GLUT2 gene expression in the pancreatic islets of diabetic animals may be due to the loss of PDX-1 transacting function on the GLUT2 gene. We, therefore, investigated the potential role of PDX-1 in the transcriptional control of GLUT2. We have identified a repeat of a TAAT motif (5'-TAATA-ATAACA-3') conserved in the sequence of the human and murine GLUT2 promoters. Recombinant PDX-1 binds to this GLUT2TAAT motif in electrophoretic mobility shift experiments. PDX-1 antiserum detects the formation of the complex of PDX-1 with the GLUT2TAAT motif in nuclear extracts from the pancreatic insulin-secreting cell line, beta TC3. The GLUT2TAAT motif was mutated in the murine GLUT2 promoter (-1308/+49 bp) linked to a luciferase reporter gene and transfected into beta TC3 cells. Compared with the transcriptional activity of the wild type promoter, that of the mutated promoter decreases by 41%. Multiple copies of the GLUT2TAAT motif were ligated 5' to a heterologous promoter and transfected into a PDX-1-expressing cell line (beta TC3) and into cell lines lacking the homeobox factor (InR1-G9 and JEG-3). The GLUT2TAAT motif mediates the activation of the heterologous promoter in the PDX-1-expressing cell line but not in InR1-G9 or JEG-3 cell lines. Furthermore, cotransfection in a PDX-1-deficient cell line with the expression vector encoding PDX-1 transactivates specifically the heterologous promoter containing the multimerized GLUT2TAAT motif. These data demonstrate that the murine GLUT2 promoter is controlled by the PDX-1 homeobox factor through the identified GLUT2TAAT motif.

High expression of the HMG box factor sox-13 in arterial walls during embryonic development.

Members of the Sox gene family of transcription factors are defined by the presence of an 80 amino acid homology domain, the High Mobility Group (HMG) box. Here we report the cloning and initial analysis of murine Sox-13 . The 984 amino acids Sox-13 protein contains a single HMG box, a leucine zipper motif and a glutamine-rich stretch. These characteristics are shared with another member of the Sox gene family, Sox-6. High level embryonic expression of Sox-13 occurs uniquely in the arterial walls of 13.5 days post coitum (dpc) mice and later. Low level expression was observed in the inner ear of 13.5 dpc mice and in a limited number of cells in the thymus of 16.5 dpc mice, from which Sox-13 was originally cloned. At 18.5 dpc, Sox-13 is expressed in the tracheal epithelium below the vocal cord and in the hair follicles. The Sox-13 protein binds to the consensus HMG box motif, AACAAAG, but does not transactivate transcription through a concatamer of this motif. Sox-13, like other members of the Sox family likely plays an important role in development.

Identification of the minimal promoter region of the mouse NKX5-3, a transcription factor implicated in eye development.

Early ocular development is controlled by a complex network of transcription factors, cell cycle regulators, and diffusible signalling molecules. Together, these molecules regulate cell proliferation and apoptosis, and specify retinal fate. NKX5-3 is a homeobox transcription factor implicated in eye development. The analysis of the 5'-flanking region of the mouse Nkx5-3 gene revealed a predicted TATA-less promoter sequence between -416 and -166 of the translation start site. To functionally characterise Nkx5-3 promoter activity, serial deletions of the promoter sequence were introduced in pGL-3 basic vector and promoter activity of these 5'- and 3'-deleted constructions was tested in HeLa and CHO cells. Transactivation assays identified a region between -350 and -296 exhibiting promoter-like activity. Combined analysis by deletions and point mutations showed that this sequence, containing multiple Sp1 binding sites was necessary to promote transcriptional activity. Binding of Sp1 to this region was confirmed by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation, using an antibody specific for Sp1. Altogether, these results demonstrated that the immediate upstream region of Nkx5-3 gene possessed a strong intrinsic promoter activity in vitro, suggesting a potential role in Nkx5-3 transcription in vivo.

Labor Mobility and Productivity Growth

[cat] Els models de creixement amb aprenentatge suposen que el coneixement après en producció es transmet de forma lliure i instantània a tota l'economia. En con- seqüència, l'economia presenta economies d'escala creixents i el creixement de la productivitat (TFP) és endògena. No obstant, el supòsit de difusió instantània del coneixement és poc realista. La difusió del coneixement necessita temps i algun canal de transmissió. En aquest article suposem que el coneixement es transmet amb la contractació de treballadors nous (learning-by-hiring). En el nostre model la difusió instantània i lliure de coneixement pot ocórrer només dins d'un sector. La difusió de coneixement entre sectors pot ocórrer només a través de la mobilitat de treballadors, i per tant, el mercat de treball determina el nivell i la taxa de creixement de productivitat (TFP). Estudiem com els costos de mobilitat laboral modifiquen l'equilibri sota dos escenaris: creixement endogen i exogen. A més, demostrem que d'altres ineficiències del mercat laboral, com són les taxes o els costos de cerca, poden reduir la mobilitat laboral, i per tant, modificar la TFP.

Labor Mobility and Productivity Growth

[cat] Els models de creixement amb aprenentatge suposen que el coneixement après en producció es transmet de forma lliure i instantània a tota l'economia. En con- seqüència, l'economia presenta economies d'escala creixents i el creixement de la productivitat (TFP) és endògena. No obstant, el supòsit de difusió instantània del coneixement és poc realista. La difusió del coneixement necessita temps i algun canal de transmissió. En aquest article suposem que el coneixement es transmet amb la contractació de treballadors nous (learning-by-hiring). En el nostre model la difusió instantània i lliure de coneixement pot ocórrer només dins d'un sector. La difusió de coneixement entre sectors pot ocórrer només a través de la mobilitat de treballadors, i per tant, el mercat de treball determina el nivell i la taxa de creixement de productivitat (TFP). Estudiem com els costos de mobilitat laboral modifiquen l'equilibri sota dos escenaris: creixement endogen i exogen. A més, demostrem que d'altres ineficiències del mercat laboral, com són les taxes o els costos de cerca, poden reduir la mobilitat laboral, i per tant, modificar la TFP.

Hepatocyte nuclear factor 4alpha transactivates the mitochondrial alanine aminotransferase gene in the kidney of Sparus aurata

Alanine aminotransferase (ALT) plays an important role in amino acid metabolism and gluconeogenesis. The preference of carnivorous fish for protein amino acids instead of carbohydrates as a source of energy lead us to study the transcriptional regulation of the mitochondrial ALT (mALT) gene and to characterize the enzyme kinetics and modulation of mALT expression in the kidney of gilthead sea bream (Sparus aurata) under different nutritional and hormonal conditions. 5′-Deletion analysis of mALT promoter in transiently transfected HEK293 cells, site-directed mutagenesis and electrophoretic mobility shift assays allowed us to identify HNF4α as a new factor involved in the transcriptional regulation of mALT expression. Quantitative RT-PCR assays showed that starvation and the administration of streptozotocin (STZ) decreased HNF4α levels in the kidney of S. aurata, leading to the downregulation of mALT transcription. Analysis of the tissue distribution showed that kidney, liver, and intestine were the tissues with higher mALT and HNF4α expression. Kinetic analysis indicates that mALT enzyme is more efficient in catalyzing the conversion of L-alanine to pyruvate than the reverse reaction. From these results, we conclude that HNF4α transactivates the mALT promoter and that the low levels of mALT expression found in the kidney of starved and STZ-treated fish result from a decreased expression of HNF4α. Our findings suggest that the mALT isoenzyme plays a major role in oxidazing dietary amino acids, and points to ALT as a target for a biotechnological action to spare protein and optimize the use of dietary nutrients for fish culture.

The attenuation of concentrations model: a new method for assessing mercury mobility in sediments

In this work we propose a new approach for the determination of the mobility of mercury in sediments based on spatial distribution of concentrations. We chose the Tainheiros Cove, located in the Todos os Santos Bay, Brazil, as the study area, for it has a history of mercury contamination due to a chloro-alkali plant that was active during 12 years. Twenty-six surface sediment samples were collected from the area and mercury concentrations were measured by cold vapour atomic absorption spectrophotometry. A contour map was constructed from the results, indicating that mercury accumulated in a "hot spot" where concentrations reach more than 1 µg g-1. The model is able to estimate mobility of mercury in the sediments based on the distances between iso-concentration contours that determines an attenuation of concentrations factor. Values of attenuation ranged between 0.0729 (East of the hot spot, indicating higher mobility) to 0.7727 (North of the hot spot, indicating lower mobility).