Synchrotron X-ray powder diffraction and extended X-ray absorption fine structure spectroscopy studies on nanocrystalline ZrO(2)-CaO solid solutions

The crystal structure and the local atomic order of a series of nanocrystalline ZrO(2)-CaO solid solutions with varying CaO content were studied by synchrotron radiation X-ray powder diffraction and extended X-ray absorption fine structure (EXAFS) spectroscopy. These samples were synthesized by a pH-controlled nitrate-glycine gel-combustion process. For CaO contents up to 8 mol%, the t' form of the tetragonal phase (c/a > 1) was identified, whereas for 10 and 12 mol% CaO, the t '' form (c/a=1; oxygen anions displaced from their ideal positions in the cubic phase) was detected. Finally, the cubic phase was observed for solid solutions with CaO content of 14 mol% CaO or higher. The t'/t '' and t ''/cubic compositional boundaries were determined to be at 9 (1) and 13 (1) mol% CaO, respectively. The EXAFS study demonstrated that this transition is related to a tetragonal-to-cubic symmetry change of the first oxygen coordination shell around the Zr atoms.

Vortex configuration flow cell based on low temperature cofired ceramics as a compact chemiluminescence microsystern

The integration of optical detection methods in continuous flow microsystems can highly extend their range of application, as long as some negative effects derived from their scaling down can be minimized. Downsizing affects to a greater extent the sensitivity of systems based on absorbance measurements than the sensitivity of those based on emission ones. However, a careful design of the instrumental setup is needed to maintain the analytical features in both cases. In this work, we present the construction and evaluation of a simple miniaturized optical system, which integrates a novel flow cell configuration to carry out chemiluminescence (CL) measurements using a simple photodiode. It consists of a micro-mixer based on a vortex structure, which has been constructed by means of the low-temperature cofired ceramics (LTCC) technology. This mixer not only efficiently promotes the CL reaction due to the generated high turbulence but also allows the detection to be carried out in the same area, avoiding intensity signal losses. As a demonstration, a flow injection system has been designed and optimized for the detection of cobalt(H) in water samples. It shows a linear response between 2 and 20 mu M with a correlation of r > 0.993, a limit of detection of 1.1 mu M, a repeatability of RSD = 12.4 %, and an analysis time of 17 s. These results demonstrate the suitability of the proposal to the determination of compounds involved in CL reactions by means of an easily constructed versatile device based on low-cost instrumentation.

Porous silicon optical cavity structure applied to high sensitivity organic solvent sensor

The present work reports the thermal annealing process, the number of layer and electrochemical process effect in the optical response quality of Bragg and microcavity devices that were applied as organic solvent sensors. These devices have been obtained by using porous silicon (PS) technology. The optical characterization of the Bragg reflector, before annealing, showed a broad photonic band-gap structure with blue shifted and narrowed after annealing process. The electrochemical process used to obtain the PS-based device imposes the limit in the number of layers because of the chemical dissolution effect. The interface roughness minimizations in the devices have been achieved by using the double electrochemical cell setup. The microcavity devices showed to have a good sensibility for organic solvent detection. The thermal annealed device showed better sensibility feature and this result was attributed to passivation of the surface devices. (c) 2007 Elsevier Ltd. All rights reserved.

Genome structure of a Saccharomyces cerevisiae strain widely used in bioethanol production

Bioethanol is a biofuel produced mainly from the fermentation of carbohydrates derived from agricultural feedstocks by the yeast Saccharomyces cerevisiae. One of the most widely adopted strains is PE-2, a heterothallic diploid naturally adapted to the sugar cane fermentation process used in Brazil. Here we report the molecular genetic analysis of a PE-2 derived diploid (JAY270), and the complete genome sequence of a haploid derivative (JAY291). The JAY270 genome is highly heterozygous (similar to 2 SNPs/kb) and has several structural polymorphisms between homologous chromosomes. These chromosomal rearrangements are confined to the peripheral regions of the chromosomes, with breakpoints within repetitive DNA sequences. Despite its complex karyotype, this diploid, when sporulated, had a high frequency of viable spores. Hybrid diploids formed by outcrossing with the laboratory strain S288c also displayed good spore viability. Thus, the rearrangements that exist near the ends of chromosomes do not impair meiosis, as they do not span regions that contain essential genes. This observation is consistent with a model in which the peripheral regions of chromosomes represent plastic domains of the genome that are free to recombine ectopically and experiment with alternative structures. We also explored features of the JAY270 and JAY291 genomes that help explain their high adaptation to industrial environments, exhibiting desirable phenotypes such as high ethanol and cell mass production and high temperature and oxidative stress tolerance. The genomic manipulation of such strains could enable the creation of a new generation of industrial organisms, ideally suited for use as delivery vehicles for future bioenergy technologies.

Trypanocidal structure-activity relationship for cis- and trans-methylpluviatolide

The trypanocidal activity of racemic mixtures of cis- and trans-methylpluviatolides was evaluated in vitro against trypomastigote forms of two strains of Trypanosoma cruzi, and in the enzymatic assay of T. cruzi gGAPDH. The cytotoxicity of the compounds was assessed by the MTT method using LLC-MK2 cells. The effect of the compounds on peroxide and NO production were also investigated. The mixture of the trans stereoisomers displayed trypanocidal activity (IC(50) similar to 89.3 mu M). Therefore, it was separated by chiral HPLC, furnishing the and (+) (-)-enantiomers. Only the (-)-enantiomer was active against the parasite (IC(50) similar to 18.7 mu M). Despite being inactive, the (+)-enantiomer acted as an antagonistic competitor. Trans-methylpluviatolide displayed low toxicity for LLC-MK(2) cells, with an IC(50) of 6.53 mM. Furthermore, methylpluviatolide neither inhibited gGAPDH activity nor hindered peroxide and NO production at the evaluated concentrations. (C) 2008 Elsevier Ltd. All rights reserved.

Anti-Group A Streptococcal Vaccine Epitope STRUCTURE, STABILITY, AND ITS ABILITY TO INTERACT WITH HLA CLASS II MOLECULES

Streptococcus pyogenes infections remain a health problem in several countries due to poststreptococcal sequelae. We developed a vaccine epitope (StreptInCor) composed of 55 amino acids residues of the C-terminal portion of the M protein that encompasses both T and B cell protective epitopes. The nuclear magnetic resonance (NMR) structure of the StreptInCor peptide showed that the structure was composed of two microdomains linked by an 18-residue alpha-helix. A chemical stability study of the StreptInCor folding/unfolding process using far-UV circular dichroism showed that the structure was chemically stable with respect to pH and the concentration of urea. The T cell epitope is located in the first microdomain and encompasses 11 out of the 18 alpha-helix residues, whereas the B cell epitope is in the second microdomain and showed no alpha-helical structure. The prediction of StreptInCor epitope binding to different HLA class II molecules was evaluated based on an analysis of the 55 residues and the theoretical possibilities for the processed peptides to fit into the P1, P4, P6, and P9 pockets in the groove of several HLA class II molecules. We observed 7 potential sites along the amino acid sequence of StreptInCor that were capable of recognizing HLA class II molecules (DRB1*, DRB3*, DRB4*, and DRB5*). StreptInCoroverlapping peptides induced cellular and humoral immune responses of individuals bearing different HLA class II molecules and could be considered as a universal vaccine epitope.

Structure activity relationship, acute toxicity and cytotoxicity of antimycobacterial neolignan analogues

Objectives The study`s aims were to evaluate the antimycobacterial activity of 13 synthetic neolignan analogues and to perform structure activity relationship analysis (SAR). The cytotoxicity of the compound 2-phenoxy-1-phenylethanone (LS-2, 1) in mammalian cells, such as the acute toxicity in mice, was also evaluated. Methods The extra and intracellular antimycobacterial activity was evaluated on Mycobacterium tuberculosis H37Rv. Cytotoxicity studies were performed using V79 cells, J774 macrophages and rat hepatocytes. Additionally, the in-vivo acute toxicity was tested in mice. The SAR analysis was performed by Principal Component Analysis (PCA). Key findings Among the 13 analogues tested, LS-2 (1) was the most effective, showing promising antimycobacterial activity and very low cytotoxicity in V79 cells and in J774 macrophages, while no toxicity was observed in rat hepatocytes. The selectivity index (SI) of LS-2 (1) was 91 and the calculated LD50 was 1870 mg/kg, highlighting the very low toxicity in mice. SAR analysis showed that the highest electrophilicity and the lowest molar volume are physical-chemical characteristics important for the antimycobacterial activity of the LS-2 (1). Conclusions LS-2 (1) showed promising antimycobacterial activity and very weak cytotoxicity in cell culture, as well as an absence of toxicity in primary culture of hepatocytes. In the acute toxicity study there was an indication of absence of toxicity on murine models, in vivo.

Human salivary gland morphogenesis: myoepithelial cell maturation assessed by immunohistochemical markers

Aims: Myoepithelial cells are important components of salivary gland structure, aiding the expulsion of saliva from acinar lobules. The aim was to evaluate the expression of smooth muscle actin (SMA), calponin, caldesmon, CD10, CD29, S100 protein, glial fibrillary acidic protein (GFAP) and p63 in myoepithelial cells during salivary gland morphogenesis to understand the maturation process of these cells and their possible use in the diagnosis of salivary gland lesions. Methods and results: Major and minor human salivary glands at various stages of development, derived from fetuses at 8-26 weeks of gestation, were studied immunohistochemically. Fully developed salivary glands were used as controls. The protein p63 was present in all stages of salivary gland morphogenesis from initial bud to terminal bud stage. CD29, S100 and calponin were detected increasingly as salivary gland structure matured and in fully developed salivary gland. Proteins GFAP, CD10 and caldesmon were not observed in myoepithelial cells of salivary glands. Conclusions: The proteins SMA, calponin, CD29, S100 and p63, which are present from the earliest stages of salivary gland maturation, are valuable myoepithelial markers but, although very specific, are not exclusive markers for this cell type.

Human alveolar bone cell proliferation, expression of osteoblastic phenotype, and matrix mineralization on porous titanium produced by powder metallurgy

Objective: This study aimed at investigating the influence of the porous titanium (Ti) structure on the osteogenic cell behaviour. Materials and methods: Porous Ti discs were fabricated by the powder metallurgy process with the pore size typically between 50 and 400 mm and a porosity of 60%. Osteogenic cells obtained from human alveolar bone were cultured until subconfluence and subcultured on dense Ti (control) and porous Ti for periods of up to 17 days. Results: Cultures grown on porous Ti exhibited increased cell proliferation and total protein content, and lower levels of alkaline phosphatase (ALP) activity than on dense Ti. In general, gene expression of osteoblastic markers-runt-related transcription factor 2, collagen type I, alkaline phosphatase, bone morphogenetic protein-7, and osteocalcin was lower at day 7 and higher at day 17 in cultures grown on porous Ti compared with dense Ti, a finding consistent with the enhanced growth rate for such cultures. The amount of mineralized matrix was greater on porous Ti compared with the dense one. Conclusion: These results indicate that the porous Ti is an appropriate substrate for osteogenic cell adhesion, proliferation, and production of a mineralized matrix. Because of the three-dimensional environment it provides, porous Ti should be considered an advantageous substrate for promoting desirable implant surface-bone interactions.

Storage proteins and cell wall mobilisation in seeds of Sesbania virgata (Cav.) Pers. (Leguminosae)

The endosperm of seeds of Sesbania virgata (Cav.) Pers. accumulates galactomannan as a cell wall storage polysaccharide. It is hydrolysed by three enzymes, one of them being alpha-galactosidase. A great amount of protein bodies is found in the cytoplasm of endospermic cells, which are thought to play the major role as a nitrogen reserve in this seed. The present work aimed at understanding how the production of enzymes that degrade storage compounds is controlled. We performed experiments with addition of inhibitors of transcription (actinomycin-d and alpha-amanitin) and translation (cycloheximide) during and after germination. In order to follow the performance of storage mobilisation, we measured fresh mass, protein contents and alpha-galactosidase activity. All the inhibitors tested had little effect on seed germination and seedling development. Actinomycin-d and cycloheximide provoked a slight inhibition of the storage protein degradation and concomitantly lead to an elevation of the alpha-galactosidase activity. Although alpha-amanitin showed some effect on seedling development at latter stages, it presented the former effect and did not change galactomannan degradation performance. Our data suggest that some of the proteases may be synthesised de novo, whereas alpha-galactosidase seems to be present in the endosperm cells probably as an inactive polypeptide in the protein bodies, being probably activated by proteolysis when the latter organelle is disassembled. These evidences suggest the existence of a connection between storage proteins and carbohydrates mobilisation in seeds of S. virgata, which would play a role by assuring a balanced afflux of the carbon and nitrogen to the seedling development.

Cell wall polysaccharides from cell suspension cultures of the Atlantic Forest tree Rudgea jasminoides (Rubiaceae)

Rudgea jasminoides (Rubiaceae) is a tropical tree species native of the Atlantic Forest in the south of Brazil. Previous studies with leaf cell walls of R. jasminoides showed a different proportion of cross-linked glycans compared to what is usually reported for eudicots. However, due to the difficulties of working with whole plant organs, cell suspensions of R. jasminoides, consisting of predominantly undifferentiated cells with mainly primary cell walls, were used to examine cell walls and extracellular soluble polysaccharides (EP) released into the culture medium. Sugar composition and linkage analysis showed homogalacturonans, xylogalacturonans and arabinogalactans to be the predominant EP. In the cell wall, homogalacturonans and arabinogalactans are the major pectins, and xyloglucans and xylans are the major cross-linking glycans. The presence of xylogalacturonans in the R. jasminoides cell cultures seems to be related to the occurrence of a homogeneous cell suspension with loosely attached cells. Although all alkali extractions from the cell walls yielded amounts of xyloglucan that exceed those of the xylans, the latter was found in a proportion that is higher than what has been usually reported for primary cell walls of most eudicots. The xyloglucan from cell walls of cell suspension cultures of R. jasminoides has low fucosylation levels and high proportion of galactosyl residues, a branching pattern commonly found in storage cell-wall xyloglucans.

Cell death and lumen formation in spheroids of MCF-7 cells

3D (three-dimensional) cell culture permits a more integrated analysis of the relationship between cells, inserting them into a structure more closely resembling the cellular microenvironment in vivo. The development of in vitro parameters to approximate in vivo 3D cellular environments makes a less reductionist interpretation of cell biology possible. For breast cells, in vitro 3D culture has proven to be an important tool for the analysis of luminal morphogenesis. A greater understanding of this process is necessary because alterations in the lumen arrangement are associated with carcinogenesis. Following lumen formation in 3D cell culture using laser scanning confocal microscopy, we observed alterations in the arrangement of cytoskeletal components (F-actin and microtubules) and increasing levels of cell death associated with lumen formation. The formation of a polarized monolayer facing the lumen was characterized through 3D reconstructions and the use of TEM (transmission electron microscopy), and this process was found to occur through the gradual clearing of cells from the medullary region of the spheroids. This process was associated with different types of cell death, such as apoptosis, autophagy and entosis. The present study showed that changes in the extracellular matrix associated with long periods of time in 3D cell culture lead to the formation of a lumen in MCF-7 cell spheroids and that features of differentiation such as lumen and budding formation occur after long periods in 3D culture, even in the absence of exogenous extracellular compounds.

The crystal structure of the leptospiral hypothetical protein LIC12922 reveals homology with the periplasmic chaperone SurA

Leptospirosis is a world spread zoonosis caused by members of the genus Leptospira. Although leptospires were identified as the causal agent of leptospirosis almost 100 years ago, little is known about their biology, which hinders the development of new treatment and prevention strategies. One of the several aspects of the leptospiral biology not yet elucidated is the process by which outer membrane proteins (OMPs) traverse the periplasm and are inserted into the outer membrane. The crystal structure determination of the conserved hypothetical protein LIC12922 from Leptospira interrogans revealed a two domain protein homologous to the Escherichia coli periplasmic chaperone SurA. The LIC12922 NC-domain is structurally related to the chaperone modules of E. coli SurA and trigger factor, whereas the parvulin domain is devoid of peptidyl prolyl cis-trans isomerase activity. Phylogenetic analyses suggest a relationship between LIC12922 and the chaperones PrsA, PpiD and SurA. Based on our structural and evolutionary analyses, we postulate that LIC12922 is a periplasmic chaperone involved in OMPs biogenesis in Leptospira spp. Since LIC12922 homologs were identified in all spirochetal genomes sequenced to date, this assumption may have implications for the OMPs biogenesis studies not only in leptospires but in the entire Phylum Spirochaetes. (C) 2010 Elsevier Inc. All rights reserved.

Percolation in a network with long-range connections: Implications for cytoskeletal structure and function

Cell shape, signaling, and integrity depend on cytoskeletal organization. In this study we describe the cytoskeleton as a simple network of filamentary proteins (links) anchored by complex protein structures (nodes). The structure of this network is regulated by a distance-dependent probability of link formation as P = p/d(s), where p regulates the network density and s controls how fast the probability for link formation decays with node distance (d). It was previously shown that the regulation of the link lengths is crucial for the mechanical behavior of the cells. Here we examined the ability of the two-dimensional network to percolate (i.e. to have end-to-end connectivity), and found that the percolation threshold depends strongly on s. The system undergoes a transition around s = 2. The percolation threshold of networks with s < 2 decreases with increasing system size L, while the percolation threshold for networks with s > 2 converges to a finite value. We speculate that s < 2 may represent a condition in which cells can accommodate deformation while still preserving their mechanical integrity. Additionally, we measured the length distribution of F-actin filaments from publicly available images of a variety of cell types. In agreement with model predictions, cells originating from more deformable tissues show longer F-actin cytoskeletal filaments. (C) 2008 Elsevier B.V. All rights reserved.

Synthesis, characterization, crystal structure, and biological studies of vanadium complexes with glycolic acid

Synthesis, characterization, crystal structure, and biological studies of two complexes with glycolic acid are described. The solid complexes were formulated as K2[VO(C2H2O3)(C2H3O3)2] H2O (1) and K2[{VO2(C2H2O3)}2] (2) and characterized by X-ray studies, Fourier transform infrared spectroscopy (FTIR), Electron paramagnetic resonance (EPR), and magnetic susceptibility. Conversion of 1 to 2 was studied in aqueous solution by UV-Vis spectroscopy and in the solid state by diffuse reflectance spectroscopy. Complex 2 contains dinuclear [{VO2(C2H2O3)}2]2- anions in which glycolate(2-) is a five-membered chelating ring formed by carboxylate and -hydroxy groups. The geometry around the vanadium in 2 was interpreted as intermediate between a trigonal bipyramid and a square pyramid. Vanadium(IV) is pentacoordinate in 1 as a distorted square pyramid. Complex 1 contains a vanadyl group (V=O) surrounded by two oxygens from deprotonated carboxylate and hydroxy groups forming a five-membered ring. Two oxygens from different glycolates(1-) are bonded to the (V=O) also. Biological analysis for potential cytotoxic effects of 1 was performed using Human Cervix Adenocarcinoma (HeLa) cells, a human cervix adenocarcinoma-derived cell line. After incubation for 48 h, 1 causes 90 and 95% of HeLa cells death at 20 and 200 mol L-1, respectively.