Differential stress response and susceptibility to oxidative injury in alveolar macrophages from C57BL/6J and C3H/HeJ mice following oxidant exposure

Studies have demonstrated a variable response to ozone among individuals and animal species and strains. For instance, C57BL/6J mice have a greater inflammatory response to ozone exposure than C3H/HeJ mice. In these studies, I utilized these strain differences in an effort to derive a mechanistic explanation to the variable strain sensitivity to ozone exposure. Therefore, alveolar macrophages (AM) from C57BL/6J and C3H/HeJ mice were exposed in vitro to hydrogen peroxide ($\rm H\sb2O\sb2$), heat and acetyl ceramide or in vivo to ozone. Necrosis and DNA fragmentation in macrophages from the two murine strains were determined to assess cytotoxicity following these treatments. In addition, synthesis and expression of the stress proteins, stress protein 72 (SP72) and heme oxygenase (HO-1), were examined following treatments. The in vitro experiments were conducted to eliminate the possibility of in vivo confounders (i.e., differences in breathing rates in the two strains) and thus directly implicate some inherent difference between cells from the two murine strains. $\rm H\sb2O\sb2$ and heat caused greater cytotoxicity in AM from C57BL/6J than C3H/HeJ mice and DNA fragmentation was a particularly sensitive indicator of cell injury. Similarly, AM from C57BL/6J mice were more sensitive to ozone exposure than cells from C3H/HeJ mice. Exposure to either 1 or 0.4 ppm ozone caused greater cytotoxicity in macrophages from C57BL/6J mice compared to macrophages from C3H/HeJ mice. The increased sensitivity of AM to injury was associated with decreased synthesis and expression of stress proteins. AM from C57BL/6J mice synthesized and expressed significantly less stress proteins in response to heat and ozone than AM from C3H/HeJ mice. Heat treatment resulted in greater synthesis and expression of SP72. In addition, macrophages from C57BL/6J mice expressed lower amounts of HO-1 than macrophages from C3H/HeJ mice following 0.4 ppm ozone exposure. Therefore, AM from C57BL/6J mice are more susceptible to oxidative injury than AM from C3H/HeJ mice which might be due to differential expression of stress proteins in these cells. ^

Comportamiento del perfil de ácidos grasos de aceites y materias grasas hidrogenadas sometidos a calentamiento prolongado

La fritura es un proceso altamente utilizado a nivel industrial y casero, el que considera someter a las grasas a temperaturas cercanas a los 200°C, con lo cual se producen complicados cambios físicos y químicos. En Chile, se utilizan para freír mantecas hidrogenadas las cuales han mostrado alto contenido en ácidos grasos trans (AGT). Este trabajo tiene como objetivo determinar los cambios en el perfil en ácidos grasos, con énfasis en AGT, de diferentes tipos de materias grasas hidrogenadas y aceites vegetales sometidos a calentamiento. Para esto se evaluó y se sometió a un ciclo de calentamiento por 50 h 7 diferentes tipos de materias grasas y como control se consideró a aceite de girasol, se evaluaron los tiempos 0, 10, 20, 30, 40 y 50 h. La composición en ácidos grasos se realizó por cromatografía gas - liquido previa preparación de los ésteres metílicos según Norma española, UNE-EN ISO 5509. Los resultados mostraron que la composición en ácidos grasos de todas las materias grasas estudiadas presentaron modificación durante el tratamiento térmico, observándose una disminución de los ácidos grasos poliinsaturados y un aumento o preservación de los ácidos grasos monoinsaturados y saturados. Los ácidos grasos trans mostraron contenidos importantes en mantecas hidrogenadas (20%), los que se mantuvieron constantes durante todo el calentamiento. El aceite de girasol y el de girasol alto oleico resultaron ser los más adecuados para freír, ya que no mostraron presencia de AGT y se mantuvieron aptos para ser utilizados hasta las 40 h de calentamiento.

Microstructural and mechanical characteristics of W-2Ti and W-1TiC processed by Hot Isostatic Pressing

It has been demonstrated that mechanical alloying and subsequent consolidation by hot isostatic pressing (HIP) is a successful route to produce dispersion strengthened W alloys with properties satisfying the design requirements of particular plasma facing components in the fusion reactor. However, the presence of the alloying element as a phase filling large interstices between W particles appears to reduce the mechanical properties of these alloys. In order to limit this phase separation induced by the HIP treatment and the detrimental effects on the mechanical properties, the enhancement of the mechanical alloying process, and the effect of a postconsolidation heat treatment in an reducing atmosphere, have been investigated.

Analysis of crystallographic slip and grain boundary sliding in a Ti-45Al-2Nb-2Mn (at%)-0.8 vol%TiB2 alloy by high temperature in situ mechanical testing

This work aims to contribute to a further understanding of the fundamentals of crystallographic slip and grain boundary sliding in the γ-TiAl Ti–45Al–2Nb–2Mn (at%)–0.8 vol%TiB2 intermetallic alloy, by means of in situ high-temperature tensile testing combined with electron backscatter diffraction (EBSD). Several microstructures, containing different fractions and sizes of lamellar colonies and equiaxed γ-grains, were fabricated by either centrifugal casting or powder metallurgy, followed by heat treatment at 1300 °C and furnace cooling. in situ tensile and tensile-creep experiments were performed in a scanning electron microscope (SEM) at temperatures ranging from 580 °C to 700 °C. EBSD was carried out in selected regions before and after straining. Our results suggest that, during constant strain rate tests, true twin γ/γ interfaces are the weakest barriers to dislocations and, thus, that the relevant length scale might be influenced by the distance between non-true twin boundaries. Under creep conditions both grain/colony boundary sliding (G/CBS) and crystallographic slip are observed to contribute to deformation. The incidence of boundary sliding is particularly high in γ grains of duplex microstructures. The slip activity during creep deformation in different microstructures was evaluated by trace analysis. Special emphasis was placed in distinguishing the compliance of different slip events with the Schmid law with respect to the applied stress.

Influencia de la atmósfera de sinterización en las propiedades mecánicas de los aceros P/M AISI 430.LInfluence of sintering atmosphere on the mechanical properties of steel P/M AISI 430L

Se ha estudiado el acero inoxidable pulvimetalúrgico AISI 430L, comparando la sinterización en dos atmósferas diferentes; en vacío, y en una atmósfera que contiene nitrógeno. Se ha desarrollado un tratamiento térmico con objeto de incrementar las propiedades mecánicas, mediante la modificación microestructural de los nitruros complejos de hierro y cromo precipitados durante la etapa de sinterización. Se han evaluado las propiedades físicas y a la vez se ha realizado un análisis microestructural con el fin de relacionar la microestructura con el incremento en las propiedades mecánicas. Influence of sintering atmosphere on the mechanical properties of steel P / M AISI 430L. It has studied the stainless steel powder metallurgy AISI 430L. It has compared the sintering in two different atmospheres; in vacuum, and in an atmosphere containing nitrogen. It has developed a heat treatment with the aim of improving the mechanical properties. This has been done through microstructural modification of complex nitrides of iron and chromium precipitates during the phase of sintering. Physical properties have been evaluated and are been performing a microstructural analysis for microstructure related to the increase in mechanical properties.

Madera termo-tratada de frondosas para uso estructural

La madera termotratada es madera modificada mediante un proceso térmico a elevadas temperaturas que le proporciona mayor estabilidad dimensional y durabilidad sin incorporar productos químicos perjudiciales para el medio ambiente. Hasta el momento se ha aplicado fundamentalmente a madera de coniferas por motivos económicos, siendo su uso más habitual en ambientes exteriores o de elevada humedad, como elementos de revestimiento no estructurales, carpinterías, mobiliario de jardín, etc. En la presente tesis se estudia la viabilidad de la madera termotratada de frondosas para uso estructural, en particular fresno (Fraxinus excelsior L) y haya (Fagus sylvatica L). Con este fin, y considerando que el termotratamiento modifica la estructura interna de la madera resultando en un nuevo material, se realizan estudios experimentales y numéricos para su caracterización. Estos trabajos se desarrollan bajo el enfoque de la Mecánica de Fractura debido a la pérdida de resistencia y aumento de fragilidad que presenta el material, especialmente a tracción perpendicular a las fibras. Así mismo, se lleva a cabo una recopilación de las bases, fundamentos y metodologías de esta teoría aplicados a madera sin tratar y otros materiales debido a la inexistencia de este tipo de estudios en madera termotratada. De igual manera se realiza un programa de caracterización mecánica del material para determinar sus propiedades elásticas considerando un modelo ortótropo, necesarios en la investigación del comportamiento a fractura. El trabajo derivó en el desarrollo de un nuevo método de ensayo para la determinación multiparamétrica a partir de un sólo espécimen, proporcionando resultados mucho más robustos que los obtenidos con la metodología convencional de ensayos. En base a los trabajos realizados, considerando las limitaciones de resistencia y fragilidad, así como la dudosa aplicabilidad de las normativas existentes en madera sin tratar, se aconseja no utilizar tratamientos térmicos intensos en elementos estructurales primarios. Se propone su aplicación en elementos secundarios, de manera que un posible colapso no implique una pérdida de fiabilidad global de la estructura. Se estudia la viabilidad de un panel sandwich innovador y ecológico para fachadas expuesto a cargas de viento, compuesto de madera termotratada en las caras y panel aislante de fibras de madera con función estructural en el alma. Esta investigación se desarrolló dentro del proyecto de investigación Europeo "Holiwood", Holistic implementation of European thermal treated hardwood (TMT) in the sector of construction industry and noise protection by sustainable, knowledge-based and value added products, perteneciente al sexto Programa Marco. ABSTRACT Hcat-trcatcd wood is modified wood by a thermal process at high temperatures which provides greater dimensional stability and durability without adding harmful chemicals to the environment. It has been mainly applied to softwoods due mainly to economical reasons, being its most common use outdoors or in high humidity environments, as non-structural elements, furniture, etc. The present Thesis studies the feasibility of heat-treated hardwoods for structural uses, particularly ash (Fraxinus excelsior L) and beech (Fagus sylvatica L). To this end, and considering that heat treatment modifies the internal structure of the wood resulting in a new material, experimental and numerical studies are performed for its characterization. This investigation is developed under the approach of Fracture Mechanics due to the loss of strength and the increase in brittlcncss of the material, especially in tension perpendicular to the grain. Likewise, it holds a collection of the bases, foundations and methodologies of this theory applied to untreated wood and other materials due to the lack of such studies in heat-treated wood. In addition, studies for the mechanical characterization of the material are performed in order to determine the elastic properties considering an orthotropic model. This work is necessary in the investigation of the fracture behavior. It led to the development of a new test method for multiparameter determination by using just a single specimen, providing much more robust results than those obtained with conventional test methodology. Based on this investigation, and considering the limitations of strength and brittleness, and the questionable applicability of existing standards for untreated wood, it is advised not to use intense heat treatments in primary structural elements. It is proposed the application to secondary elements, so that a possible collapse does not involve a loss of overall reliability of the structure. It is studied the feasibility of an innovative and ecological sandwich panel for facades exposed to wind loads, composed by heat-treated wood faces and insulating wood fiberboard with structural function in the core. This investigación was developed within the European research project "Holiwood", Holistic implementation of European thermal treated hardwood (TMT) in the sector of construction industry and noise protection by sustainable, knowledge-based and value added products, of the Sixth Framework Program.

Determination of a refractive index and an extinction coefficient of standard production of CVD-graphene

The refractive index and extinction coefficient of chemical vapour deposition grown graphene are determined by ellipsometry analysis. Graphene films were grown on copper substrates and transferred as both monolayers and bilayers onto SiO2/Si substrates by using standard manufacturing procedures. The chemical nature and thickness of residual debris formed after the transfer process were elucidated using photoelectron spectroscopy. The real layered structure so deduced has been used instead of the nominal one as the input in the ellipsometry analysis of monolayer and bilayer graphene, transferred onto both native and thermal silicon oxide. The effect of these contamination layers on the optical properties of the stacked structure is noticeable both in the visible and the ultraviolet spectral regions, thus masking the graphene optical response. Finally, the use of heat treatment under a nitrogen atmosphere of the graphene-based stacked structures, as a method to reduce the water content of the sample, and its effect on the optical response of both graphene and the residual debris layer are presented. The Lorentz-Drude model proposed for the optical response of graphene fits fairly well the experimental ellipsometric data for all the analysed graphene-based stacked structures.

Efecto del tratamiento de harina de girasol con ácido mólico y calor sobre la producción de metano y fermetnación in vitro

Incubations were carried out with batch cultures of ruminal micro-organisms to study the effects of the treatment of sunflower meal (SFM) with malic acid at 150 ºC for 1 (SFM1) or 3 (SFM3) hours on in vitro fermentation. There were no differences (P>0.05) between SFM and SFM1 in the amount of gas and volatile fatty acids (VFA) produced and the disappearance of organic matter (OMD), but CH4 and NH3-N concentrations were reduced (P<0.05) by 11.3 and 14.5% with the malic treatment at 150 ºC for 1 hour, respectively. In contrast, SFM3 treatment reduced when compared to SFM gas and VFA production and OMD by 27.4, 32.5 and 49.6 (P<0.05), respectively, indicating decreased fermentability of SFM. The results indicate that combining malic acid and heat treatment (150ºC) for 1 h could be an effective means to reduce both protein degradability and CH4 production, but increasing the length of the treatment to 3 h resulted in reductions of SFM degradability and VFA production.

Molecular Cloning and Characterization of a Radial Spoke Head Protein of Sea Urchin Sperm Axonemes: Involvement of the Protein in the Regulation of Sperm Motility

Monoclonal antibodies raised against axonemal proteins of sea urchin spermatozoa have been used to study regulatory mechanisms involved in flagellar motility. Here, we report that one of these antibodies, monoclonal antibody D-316, has an unusual perturbating effect on the motility of sea urchin sperm models; it does not affect the beat frequency, the amplitude of beating or the percentage of motile sperm models, but instead promotes a marked transformation of the flagellar beating pattern which changes from a two-dimensional to a three-dimensional type of movement. On immunoblots of axonemal proteins separated by SDS-PAGE, D-316 recognized a single polypeptide of 90 kDa. This protein was purified following its extraction by exposure of axonemes to a brief heat treatment at 40°C. The protein copurified and coimmunoprecipitated with proteins of 43 and 34 kDa, suggesting that it exists as a complex in its native form. Using D-316 as a probe, a full-length cDNA clone encoding the 90-kDa protein was obtained from a sea urchin cDNA library. The sequence predicts a highly acidic (pI = 4.0) protein of 552 amino acids with a mass of 62,720 Da (p63). Comparison with protein sequences in databases indicated that the protein is related to radial spoke proteins 4 and 6 (RSP4 and RSP6) of Chlamydomonas reinhardtii, which share 37% and 25% similarity, respectively, with p63. However, the sea urchin protein possesses structural features distinct from RSP4 and RSP6, such as the presence of three major acidic stretches which contains 25, 17, and 12 aspartate and glutamate residues of 34-, 22-, and 14-amino acid long stretches, respectively, that are predicted to form α-helical coiled-coil secondary structures. These results suggest a major role for p63 in the maintenance of a planar form of sperm flagellar beating and provide new tools to study the function of radial spoke heads in more evolved species.

A family of genes required for maintenance of cell wall integrity and for the stress response in Saccharomyces cerevisiae

The PKC1–MPK1 pathway in yeast functions in the maintenance of cell wall integrity and in the stress response. We have identified a family of genes that are putative regulators of this pathway. WSC1, WSC2, and WSC3 encode predicted integral membrane proteins with a conserved cysteine motif and a WSC1–green fluorescence protein fusion protein localizes to the plasma membrane. Deletion of WSC results in phenotypes similar to mutants in the PKC1–MPK1 pathway and an increase in the activity of MPK1 upon a mild heat treatment is impaired in a wscΔ mutant. Genetic analysis places the function of WSC upstream of PKC1, suggesting that they play a role in its activation. We also find a genetic interaction between WSC and the RAS–cAMP pathway. The RAS–cAMP pathway is required for cell cycle progression and for the heat shock response. Overexpression of WSC suppresses the heat shock sensitivity of a strain in which RAS is hyperactivated and the heat shock sensitivity of a wscΔ strain is rescued by deletion of RAS2. The functional characteristics and cellular localization of WSC suggest that they may mediate intracellular responses to environmental stress in yeast.

Herpes simplex virus DNA packaging sequences adopt novel structures that are specifically recognized by a component of the cleavage and packaging machinery

The product of the herpes simplex virus type 1 UL28 gene is essential for cleavage of concatemeric viral DNA into genome-length units and packaging of this DNA into viral procapsids. To address the role of UL28 in this process, purified UL28 protein was assayed for the ability to recognize conserved herpesvirus DNA packaging sequences. We report that DNA fragments containing the pac1 DNA packaging motif can be induced by heat treatment to adopt novel DNA conformations that migrate faster than the corresponding duplex in nondenaturing gels. Surprisingly, these novel DNA structures are high-affinity substrates for UL28 protein binding, whereas double-stranded DNA of identical sequence composition is not recognized by UL28 protein. We demonstrate that only one strand of the pac1 motif is responsible for the formation of novel DNA structures that are bound tightly and specifically by UL28 protein. To determine the relevance of the observed UL28 protein–pac1 interaction to the cleavage and packaging process, we have analyzed the binding affinity of UL28 protein for pac1 mutants previously shown to be deficient in cleavage and packaging in vivo. Each of the pac1 mutants exhibited a decrease in DNA binding by UL28 protein that correlated directly with the reported reduction in cleavage and packaging efficiency, thereby supporting a role for the UL28 protein–pac1 interaction in vivo. These data therefore suggest that the formation of novel DNA structures by the pac1 motif confers added specificity on recognition of DNA packaging sequences by the UL28-encoded component of the herpesvirus cleavage and packaging machinery.

A Selaginella lepidophylla Trehalose-6-Phosphate Synthase Complements Growth and Stress-Tolerance Defects in a Yeast tps1 Mutant1

The accumulation of the disaccharide trehalose in anhydrobiotic organisms allows them to survive severe environmental stress. A plant cDNA, SlTPS1, encoding a 109-kD protein, was isolated from the resurrection plant Selaginella lepidophylla, which accumulates high levels of trehalose. Protein-sequence comparison showed that SlTPS1 shares high similarity to trehalose-6-phosphate synthase genes from prokaryotes and eukaryotes. SlTPS1 mRNA was constitutively expressed in S. lepidophylla. DNA gel-blot analysis indicated that SlTPS1 is present as a single-copy gene. Transformation of a Saccharomyces cerevisiae tps1Δ mutant disrupted in the ScTPS1 gene with S. lepidophylla SlTPS1 restored growth on fermentable sugars and the synthesis of trehalose at high levels. Moreover, the SlTPS1 gene introduced into the tps1Δ mutant was able to complement both deficiencies: sensitivity to sublethal heat treatment at 39°C and induced thermotolerance at 50°C. The osmosensitive phenotype of the yeast tps1Δ mutant grown in NaCl and sorbitol was also restored by the SlTPS1 gene. Thus, SlTPS1 protein is a functional plant homolog capable of sustaining trehalose biosynthesis and could play a major role in stress tolerance in S. lepidophylla.

Electric Signaling and Pin2 Gene Expression on Different Abiotic Stimuli Depend on a Distinct Threshold Level of Endogenous Abscisic Acid in Several Abscisic Acid-Deficient Tomato Mutants1

Experiments were performed on three abscisic acid (ABA)-deficient tomato (Lycopersicon esculentum Mill.) mutants, notabilis, flacca, and sitiens, to investigate the role of ABA and jasmonic acid (JA) in the generation of electrical signals and Pin2 (proteinase inhibitor II) gene expression. We selected these mutants because they contain different levels of endogenous ABA. ABA levels in the mutant sitiens were reduced to 8% of the wild type, in notabilis they were reduced to 47%, and in flacca they were reduced to 21%. In wild-type and notabilis tomato plants the induction of Pin2 gene expression could be elicited by heat treatment, current application, or mechanical wounding. In flacca and sitiens only heat stimulation induced Pin2 gene expression. JA levels in flacca and sitiens plants also accumulated strongly upon heat stimulation but not upon mechanical wounding or current application. Characteristic electrical signals evolved in the wild type and in the notabilis and flacca mutants consisting of a fast action potential and a slow variation potential. However, in sitiens only heat evoked electrical signals; mechanical wounding and current application did not change the membrane potential. In addition, exogenous application of ABA to wild-type tomato plants induced transient changes in membrane potentials, indicating the involvement of ABA in the generation of electrical signals. Our data strongly suggest the presence of a minimum threshold value of ABA within the plant that is essential for the early events in electrical signaling and mediation of Pin2 gene expression upon wounding. In contrast, heat-induced Pin2 gene expression and membrane potential changes were not dependent on the ABA level but, rather, on the accumulation of JA.

Signals involved in wound-induced proteinase inhibitor II gene expression in tomato and potato plants.

Chemical and physical signals have been reported to mediate wound-induced proteinase inhibitor II (Pin2) gene expression in tomato and potato plants. Among the chemical signals, phytohormones such as abscisic acid (ABA) and jasmonic acid (JA) and the peptide systemin represent the best characterized systems. Furthermore, electrical and hydraulic mechanisms have also been postulated as putative Pin2-inducing systemic signals. Most of the chemical agents are able to induce Pin2 gene expression without any mechanical wounding. Thus, ABA, JA, and systemin initiate Pin2 mRNA accumulation in the directly treated leaves and in the nontreated leaves (systemic) that are located distal to the treated ones. ABA-deficient tomato and potato plants do not respond to wounding by accumulation of Pin2 mRNA, therefore providing a suitable model system for analysis of the signal transduction pathway involved in wound-induced gene activation. It was demonstrated that the site of action of JA is located downstream to the site of action of ABA. Moreover, systemin represents one of the initial steps in the signal transduction pathway regulating the wound response. Recently, it was reported that heat treatment and mechanical injury generate electrical signals, which propagate throughout the plant. These signals are capable of inducing Pin2 gene expression in the nontreated leaves of wounded plants. Furthermore, electrical current application to tomato leaves leads to an accumulation of Pin2 mRNA in local and systemic tissues. Examination of photosynthetic parameters (assimilation and transpiration rate) on several types of stimuli suggests that heat-induced Pin2 gene expression is regulated by an alternative pathway from that mediating the electrical current and mechanical wound response.

Determinação da tenacidade à fratura do aço aeronáutico 300M, submetido a diferentes parâmetros de tratamentos térmicos e criogenia, utilizando técnica de corpo cilíndrico com entalhe

Foi realizado estudo sobre a influência da variação de alguns parâmetros do tratamento térmico de têmpera e revenimento, bem como aplicação de criogenia, na tenacidade a fratura do aço aeronáutico 300M, com a utilização da técnica de corpo cilíndrico com entalhe. Primeiramente, realizou-se a construção da curva de transição dúctil frágil, através de ensaio Charpy, para amostras submetidas a três tratamentos térmicos, 1 - Têmpera seguida de Revenimento, 2 - Têmpera seguida de criogenia e revenimento. Não foi observada influência significativa da aplicação da criogenia (tratamento de numero 2) na curva de transição dúctil-frágil em comparação com o tratamento térmico número 1. Em um segundo momento, com o auxílio da técnica de ensaio de corpo cilíndrico com entalhe, realizou-se a medição da tenacidade à fratura das amostras do aço 300M submetido a quatro diferentes tratamento térmicos, Q, V, C e D. Os tratamentos variavam entre si principalmente pela temperatura de austenitização. Os valores de dureza para as amostras submetidas a criogenia (V e D) foram maiores que as dos tratamentos sem criogenia (Q e C), e também apresentaram tenacidade à fratura menor. Com relação à temperatura de austenitização, observou-se que os tratamentos C e D, austenitizados a 980°C, propiciaram a completa dissolução de precipitados primários da matriz, elevando a tenacidade à fratura com relação aos tratamentos realizados na faixa de 870°C, Q e V, superando também os efeitos negativos para a tenacidade à fratura devido ao aumento no tamanho de grão.