Morphological and thermal analyses of flexible polyurethane foams containing commercial calcium carbonate

One filler often utilized in flexible polyurethane foams is calcium carbonate (CaCO3) because it is non-abrasiveness, non-toxicity and facilitated pigmentation. However, it is observed that the excess of commercial CaCO3 utilized in industry possibly causing permanent deformations and damaging the quality of the final product. The effect of different concentrations of commercial CaCO3, in flexible foams, was studied. Different concentrations of CaCO3 were used for the synthesis of flexible polyurethane foams, which were submitted to morphological and thermal analyses to verify the alterations provoked by the progressive introduction of this filler.

Characterization of novel genes predominantly expressed in the epididymis – from genome analyses to genetically modified mice

In mammals, post-testicular sperm maturation taking place in the epididymis is required for the spermatozoa to acquire the abilities required to fertilize the egg in vivo. The epididymal epithelial cells secrete proteins and other small molecules into the lumen, where they interact with the spermatozoa and enable necessary maturational changes. In this study different in silico, in vitro and in vivo approaches were utilized in order to find novel genes responsible for the function of the epididymis and post-testicular sperm maturation in the mouse. Available online genomic databases were analyzed to identify genes potentially expressed in the epididymis, gene expression profiling was performed by studying their expression in different mouse tissues, and significance of certain genes to fertility was assessed by generating genetically modified mouse models. A recently discovered Pate (prostate and testis expression) gene family was found to be predominantly expressed in the epididymis. It represents one of the largest known gene families expressed in the epididymis, and the members code for proteins potentially involved in defense against microorganisms. Through genetically modified mouse models CRISP4 (cysteine-rich secretory protein 4) was identified to regulate sperm acrosome reaction, and BMYC to inhibit the expression of the Myc proto-oncogene in the developing testis. A mouse line expressing iCre recombinase specifically in the epididymis was also generated. This model can be used to generate conditional, epididymis-specific knock-out models, and will be a valuable tool in fertility studies.

18F-Labeled Presynaptic Dopaminergic Tracers. Synthesis, Radiochemical Analyses, and Preclinical Evaluation of [18F]FDOPA and [18F]CFT

Dysfunction of the dopaminergic system in brain is involved in several pathological conditions such as Parkinson’s disease and depression. 2β-Carbomethoxy-3β-(4-[18F] fluorophenyl)tropane ([18F]CFT) and 6-[18F]fluoro-L-dopa ([18F]FDOPA) are tracers for imaging the dopaminergic function with positron emission tomography (PET). Peripheral uptake of [18F]FDOPA is also used in the localization and diagnosis of neuroendocrine tumors. [18F]FDOPA and [18F]CFT can be synthesized by electrophilic fluorodestannylation. However, the specific radioactivity (SA) in the electrophilic fluorination is low with traditional synthetic methods. In this study, [18F]FDOPA and [18F]CFT were synthesized using post-target-produced [18F]F2 as an electrophilic fluorination agent. With this method, tracers are produced with sufficient SA for neuroreceptor studies. Specific aims in this study were to replace Freon-11 in the production of [18F]FDOPA due to the ozone depleting properties of this solvent, to determine pharmacological specificity and selectivity of [18F]CFT with respect to monoamine transporters, and to compare the ability of these tracers to reflect the degree of nigral neuronal loss in rats in which the dopaminergic system in the brain had been unilaterally destroyed by 6- OHDA. Post-target-produced [18F]F2 was successfully used in the production of [18F]FDOPA and [18F]CFT. The SA achieved was substantially higher than in previous synthetic methods. Deuterated compounds, CD2Cl2, CDCl3 and C3D6O, were found to be suitable solvents for replacing Freon-11. Both [18F]FDOPA and [18F]CFT demonstrated nigrostriatal dopaminergic hypofunction and correlated with the number of nigral dopaminergic neurons in the 6-OHDA lesioned rat. However, the dopamine transporter (DAT) tracer [18F]CFT was more sensitive than the dopamine synthesis tracer [18F]FDOPA in detecting these defects because of the higher non-specific uptake of [18F]FDOPA. [18F]CFT can also be used for imaging the norepinephrine transporter (NET) because of the specific uptake into the locus coeruleus. The observation that [18F]CFT exhibits specific uptake in the pancreas warrants further studies in humans with respect to potential utility in pancreatic imaging

Analyses of the development and glycoproteins present in the ovarian follicles of Poecilia vivipara (Cyprinodontiformes, Poeciliidae)

The morphofunctional aspects of oogenesis of Poecilia vivipara were studied aiming to understand the reproductive biology and development of species with internal fertilization, particularly those belonging to the family Poeciliidae. The stages of gonadal maturation and follicular development were characterized using mesoscopic, histological, histochemical, and lectin cytochemical analyses. Through mesoscopic evaluation the ovarian development was classified in six phases of development: immature, in maturation I, in maturation II, mature I, mature II, and post-spawn. Based on microscopic examination of the ovaries, we identified the presence of oocytes types I and II during the previtellogenic phase and types III, IV, and V during the vitellogenic phase. As oogenesis proceeded the oocyte cytosol increased in volume and presented increased cytoplasmic granule accumulation, characterizing vitellogenesis. The zona radiata (ZR) increased in thickness and complexity, and the follicular epithelium, which was initially thin and consisting of pavimentous cells, in type III oocytes exhibited cubic simple cells. The histochemical and cytochemical analyses revealed alterations in the composition of the molecular structures that form the ovarian follicle throughout the gonadal development. Our study demonstrated differences in the female reproductive system among fish species with internal and external fertilization and we suggest P. vivipara can be used as experimental model to test environmental toxicity.

In silico phylogenetic and virulence gene profile analyses of avian pathogenic Escherichia coli genome sequences

Avian pathogenic Escherichia coli (APEC) infections are responsible for significant losses in the poultry industry worldwide. A zoonotic risk has been attributed to APEC strains because they present similarities to extraintestinal pathogenic E. coli (ExPEC) associated with illness in humans, mainly urinary tract infections and neonatal meningitis. Here, we present in silico analyses with pathogenic E. coli genome sequences, including recently available APEC genomes. The phylogenetic tree, based on multi-locus sequence typing (MLST) of seven housekeeping genes, revealed high diversity in the allelic composition. Nevertheless, despite this diversity, the phylogenetic tree was able to cluster the different pathotypes together. An in silico virulence gene profile was also determined for each of these strains, through the presence or absence of 83 well-known virulence genes/traits described in pathogenic E. coli strains. The MLST phylogeny and the virulence gene profiles demonstrated a certain genetic similarity between Brazilian APEC strains, APEC isolated in the United States, UPEC (uropathogenic E. coli) and diarrheagenic strains isolated from humans. This correlation corroborates and reinforces the zoonotic potential hypothesis proposed to APEC.

Kinematic gait analyses in healthy Golden Retrievers

Kinematic analysis relates to the relative movement between rigid bodies and finds application in gait analysis and other body movements, interpretation of their data when there is change, determines the choice of treatment to be instituted. The objective of this study was to standardize the march of Dog Golden Retriever Healthy to assist in the diagnosis and treatment of musculoskeletal disorders. We used a kinematic analysis system to analyse the gait of seven dogs Golden Retriever, female, aged between 2 and 4 years, weighing 21.5 to 28 kg, clinically normal. Flexion and extension were described for shoulder, elbow, carpal, hip, femorotibialis and tarsal joints. The gait was characterized lateral and had accepted hypothesis of normality for all variables, except for the stance of hip and elbow, considering a confidence level of 95%, significance level α = 0.05. Variations have been attributed to displacement of the stripes during movement and the duplicated number of reviews. The kinematic analysis proved to be a consistent method of evaluation of the movement during canine gait and the data can be used in the diagnosis and evaluation of canine gait in comparison to other studies and treatment of dogs with musculoskeletal disorders.

Analyses of the usefulness of Software Defined Storage Solutions for Web-based Digital Preservation Applications

Presentation at Open Repositories 2014, Helsinki, Finland, June 9-13, 2014

Network connectance and autonomy analyses of the photosynthetic apparatus in tropical tree species from different successional groups under contrasting irradiance conditions

Biological systems are complex dynamical systems whose relationships with environment have strong implications on their regulation and survival. From the interactions between plant and environment can emerge a quite complex network of plant responses rarely observed through classical analytical approaches. The objective of this current study was to test the hypothesis that photosynthetic responses of different tree species to increasing irradiance are related to changes in network connectances of gas exchange and photochemical apparatus, and alterations in plant autonomy in relation to the environment. The heat dissipative capacity through daily changes in leaf temperature was also evaluated. It indicated that the early successional species (Citharexylum myrianthum Cham. and Rhamnidium elaeocarpum Reiss.) were more efficient as dissipative structures than the late successional one (Cariniana legalis (Mart.) Kuntze), suggesting that the parameter deltaT (T ºCair - T ºCleaf) could be a simple tool in order to help the classification of successional classes of tropical trees. Our results indicated a pattern of network responses and autonomy changes under high irradiance. Considering the maintenance of daily CO2 assimilation, the tolerant species (C. myrianthum and R. elaeocarpum) to high irradiance trended to maintain stable the level of gas exchange network connectance and to increase the autonomy in relation to the environment. On the other hand, the late successional species (C. legalis) trended to lose autonomy, decreasing the network connectance of gas exchange. All species showed lower autonomy and higher network connectance of the photochemical apparatus under high irradiance.

Reproductive phenology, pollination and seed dispersal syndromes on sandstone outcrop vegetation in the "Chapada Diamantina", northeastern Brazil: population and community analyses

Vegetation on rock outcrops in the "Chapada Diamantina" (soil islands) is often aggregated and surrounded by nude rock surfaces, thus creating natural units with well defined limits. The flowering and fruiting cycles of plants on 58 soil islands at altitudes between 1,100 and 1,140 meters above sea leavel were analyzed at Mãe Inácia Peak (12°27' S and 41°28' W) in the "Chapada Diamantina", Bahia, Brazil. The presence/absence of flowering and fruiting species on each soil island, and their respective cover areas were analyzed at both the population and community levels, and the phenophases of flowering and fruiting were observed during 24 successive months. The analyses of pollination and seed dispersal syndromes indicated that animals are more important in pollination than in seed dispersal (which is predominantly by anemochory and autochory). The flowering and fruiting of plants with animal pollination syndromes were correlated with rainfall and temperature. The flowering season varied during the year according to the pollination syndrome involved: entomophily was predominant from summer through autumn, ornithophily was predominant during winter, and anemophily in the spring. The staggered timing of flowering and fruiting among different species provides a nearly continuous supply of resources for the local fauna.

Phylogenetic analyses of Cladophora vagabunda (L.) C. Hoek (Cladophorales, Chlorophyta) from Brazil based on SSU rDNA sequences

The complete SSU rDNA was sequenced for 10 individuals of Cladophora vagabunda collected along the coast of Brazil. For C. rupestris (L.) Kütz. a partial SSU rDNA sequence (1634 bp) was obtained. Phylogenetic trees indicate that Cladophora is paraphyletic, but the section Glomeratae sensu lato including C. vagabunda from Brazil, Japan and France, C. albida (Nees) Kütz., C. sericea (Hudson) Kütz., and C. glomerata (L.) Kütz. is monophyletic. Within this group C. vagabunda is paraphyletic. The sequence identity for the SSU rDNA varied from 98.9% to 100% for the Brazilian C. vagabunda, and from 98.3% to 99.7% comparing the Brazilian individuals to the ones from France and Japan. Sequence identity of the Brazilian C. vagabunda to C. albida and C. sericea vary from 98.0% to 98.6%. The SSU rDNA phylogeny support partially the morphological characteristics presented by Brazilian populations of C. vagabunda. On the other hand, C. rupestris from Brazil does not group with C. rupestris from France, both sequences presenting only 96.9% of identity. The inclusion of sequences of individuals from Brazil reinforces the need of taxonomical revision for the genus Cladophora and for the complex C. vagabunda.

A method for multiple sequential analyses of macrophage functions using a small single cell sample

Microbial pathogens such as bacillus Calmette-Guérin (BCG) induce the activation of macrophages. Activated macrophages can be characterized by the increased production of reactive oxygen and nitrogen metabolites, generated via NADPH oxidase and inducible nitric oxide synthase, respectively, and by the increased expression of major histocompatibility complex class II molecules (MHC II). Multiple microassays have been developed to measure these parameters. Usually each assay requires 2-5 x 10(5) cells per well. In some experimental conditions the number of cells is the limiting factor for the phenotypic characterization of macrophages. Here we describe a method whereby this limitation can be circumvented. Using a single 96-well microassay and a very small number of peritoneal cells obtained from C3H/HePas mice, containing as little as <=2 x 10(5) macrophages per well, we determined sequentially the oxidative burst (H2O2), nitric oxide production and MHC II (IAk) expression of BCG-activated macrophages. More specifically, with 100 µl of cell suspension it was possible to quantify H2O2 release and nitric oxide production after 1 and 48 h, respectively, and IAk expression after 48 h of cell culture. In addition, this microassay is easy to perform, highly reproducible and more economical.

Pipeline for macro- and microarray analyses

The pipeline for macro- and microarray analyses (PMmA) is a set of scripts with a web interface developed to analyze DNA array data generated by array image quantification software. PMmA is designed for use with single- or double-color array data and to work as a pipeline in five classes (data format, normalization, data analysis, clustering, and array maps). It can also be used as a plugin in the BioArray Software Environment, an open-source database for array analysis, or used in a local version of the web service. All scripts in PMmA were developed in the PERL programming language and statistical analysis functions were implemented in the R statistical language. Consequently, our package is a platform-independent software. Our algorithms can correctly select almost 90% of the differentially expressed genes, showing a superior performance compared to other methods of analysis. The pipeline software has been applied to 1536 expressed sequence tags macroarray public data of sugarcane exposed to cold for 3 to 48 h. PMmA identified thirty cold-responsive genes previously unidentified in this public dataset. Fourteen genes were up-regulated, two had a variable expression and the other fourteen were down-regulated in the treatments. These new findings certainly were a consequence of using a superior statistical analysis approach, since the original study did not take into account the dependence of data variability on the average signal intensity of each gene. The web interface, supplementary information, and the package source code are available, free, to non-commercial users at http://ipe.cbmeg.unicamp.br/pub/PMmA.

Cloning, sequence analysis, and expression of cDNA coding for the major house dust mite allergen, Der f 1, in Escherichia coli

Our objective was to clone, express and characterize adult Dermatophagoides farinae group 1 (Der f 1) allergens to further produce recombinant allergens for future clinical applications in order to eliminate side reactions from crude extracts of mites. Based on GenBank data, we designed primers and amplified the cDNA fragment coding for Der f 1 by nested-PCR. After purification and recovery, the cDNA fragment was cloned into the pMD19-T vector. The fragment was then sequenced, subcloned into the plasmid pET28a(+), expressed in Escherichia coli BL21 and identified by Western blotting. The cDNA coding for Der f 1 was cloned, sequenced and expressed successfully. Sequence analysis showed the presence of an open reading frame containing 966 bp that encodes a protein of 321 amino acids. Interestingly, homology analysis showed that the Der p 1 shared more than 87% identity in amino acid sequence with Eur m 1 but only 80% with Der f 1. Furthermore, phylogenetic analyses suggested that D. pteronyssinus was evolutionarily closer to Euroglyphus maynei than to D. farinae, even though D. pteronyssinus and D. farinae belong to the same Dermatophagoides genus. A total of three cysteine peptidase active sites were found in the predicted amino acid sequence, including 127-138 (QGGCGSCWAFSG), 267-277 (NYHAVNIVGYG) and 284-303 (YWIVRNSWDTTWGDSGYGYF). Moreover, secondary structure analysis revealed that Der f 1 contained an a helix (33.96%), an extended strand (17.13%), a ß turn (5.61%), and a random coil (43.30%). A simple three-dimensional model of this protein was constructed using a Swiss-model server. The cDNA coding for Der f 1 was cloned, sequenced and expressed successfully. Alignment and phylogenetic analysis suggests that D. pteronyssinus is evolutionarily more similar to E. maynei than to D. farinae.

Cloning, bioinformatics analysis, and expression of the dust mite allergen Der f 5 of Dermatophagoides farinae

Crude extracts of house dust mites are used clinically for diagnosis and immunotherapy of allergic diseases, including bronchial asthma, perennial rhinitis, and atopic dermatitis. However, crude extracts are complexes with non-allergenic antigens and lack effective concentrations of important allergens, resulting in several side effects. Dermatophagoides farinae (Hughes; Acari: Pyroglyphidae) is one of the predominant sources of dust mite allergens, which has more than 30 groups of allergen. The cDNA coding for the group 5 allergen of D. farinae from China was cloned, sequenced and expressed. According to alignment using the VECTOR NTI 9.0 software, there were eight mismatched nucleotides in five cDNA clones resulting in seven incompatible amino acid residues, suggesting that the Der f 5 allergen might have sequence polymorphism. Bioinformatics analysis revealed that the matured Der f 5 allergen has a molecular mass of 13604.03 Da, a theoretical pI of 5.43 and is probably hydrophobic and cytoplasmic. Similarities in amino acid sequences between Der f 5 and allergens of other domestic mite species, viz. Der p 5, Blo t 5, Sui m 5, and Lep d 5, were 79, 48, 53, and 37%, respectively. Phylogenetic analysis indicated that Der f 5 and Der p 5 clustered together. Blo t 5 and Ale o 5 also clustered together, although Blomia tropicalis and Aleuroglyphus ovatus belong to different mite families, viz. Echimyopodidae and Acaridae, respectively.

Bioinformatics analysis of biomarkers and transcriptional factor motifs in Down syndrome

In this study, biomarkers and transcriptional factor motifs were identified in order to investigate the etiology and phenotypic severity of Down syndrome. GSE 1281, GSE 1611, and GSE 5390 were downloaded from the gene expression ominibus (GEO). A robust multiarray analysis (RMA) algorithm was applied to detect differentially expressed genes (DEGs). In order to screen for biological pathways and to interrogate the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database, the database for annotation, visualization, and integrated discovery (DAVID) was used to carry out a gene ontology (GO) function enrichment for DEGs. Finally, a transcriptional regulatory network was constructed, and a hypergeometric distribution test was applied to select for significantly enriched transcriptional factor motifs. CBR1, DYRK1A, HMGN1, ITSN1, RCAN1, SON, TMEM50B, and TTC3 were each up-regulated two-fold in Down syndrome samples compared to normal samples; of these, SON and TTC3 were newly reported. CBR1, DYRK1A, HMGN1, ITSN1, RCAN1, SON, TMEM50B, and TTC3 were located on human chromosome 21 (mouse chromosome 16). The DEGs were significantly enriched in macromolecular complex subunit organization and focal adhesion pathways. Eleven significantly enriched transcription factor motifs (PAX5, EGR1, XBP1, SREBP1, OLF1, MZF1, NFY, NFKAPPAB, MYCMAX, NFE2, and RP58) were identified. The DEGs and transcription factor motifs identified in our study provide biomarkers for the understanding of Down syndrome pathogenesis and progression.