Search

The main goal of this work is to characterize the biochemical profile of three halophilous microalgal strains isolated from extreme ecosystems in northern Chile to assess their potential as possible sources of technological and commercial advantages. A procedure for the validation of a routine method for determining the total protein, carbohydrate, lipid and chlorophyll a (Chl-a) contents in Chlorella fusca, Dunaliella salina and Spirulina sp. has been developed. Detection limits of 10.8 μg/mg, 2.7 μg/mg and 5.6 μg/mg for total protein content and 3.5 μg/mg, 1.8 μg/mg and 8.8 μg/mg for total carbohydrate content were determined in C. fusca, D. salina and Spirulina sp., respectively. Most of the biological macromolecule levels measured in the microalgae were in accordance with the magnitude of previously reported data for other strains of the same taxa. However, lower levels than expected of lipids and Chl-a were measured in C. fusca and Spirulina sp., which may be associated with an imbalance between specific growth rates and the rate of macromolecule synthesis. The protein values measured in Spirulina sp. (52.3 ± 2.2 μg/mg DW) were close to the lower limit of the range reported in the literature for non-halophilous strains of the same genus. Except for the Chl-a quantification procedure, the analytical methods for macromolecules had a high degree of repeatability and reproducibility. The variability among repeated measurements of Chl-a was associated with auto-degradation processes during pigment extraction.

Extended-spectrum AmpC cephalosporinases (ESACs) have been reported in Enterobacteriaceae, Pseudomonas aeruginosa and Acinetobacter baumannii. Here, we characterize a new AmpC variant presenting a broadened substrate activity towards fourth-generation cephalosporins, selected in vivo following cefepime treatment for Enterobacter aerogenes.Two consecutive clonally related isolates of E. aerogenes were evaluated. Screening for ESAC production was performed using plates containing 200 mg/L cloxacillin. MICs were determined by microdilution (CLSI guidelines). bla(AmpC) genes were cloned into a pCR-Blunt II-TOPO vector and expressed in Escherichia coli. The ampC genes were cloned into vector pGEX-6P-1 for protein purification.Isolate Ea595 was resistant to two fourth-generation cephalosporins, cefepime and cefpirome; using plates containing cloxacillin, susceptibility to ceftazidime and cefepime was restored, suggesting overproduction of the ESAC β-lactamase. Sequencing identified a new AmpC β-lactamase variant presenting one amino acid substitution, Val291Gly, inside the H-10 helix. Recombinant plasmids harbouring this ESAC β-lactamase conferred a broadened resistance profile to cefepime and cefpirome, with resistance levels increasing from 16- to 32-fold in E. coli. AmpC-Ea595 hydrolysed ceftazidime, cefepime and cefpirome at high levels, presenting a lower K(m) and enabling us to classify the enzyme as an ESAC. Homology modelling suggested that the size of the active site could have increased.We characterized an ESAC β-lactamase selected in vivo and conferring a high level of resistance to fourth-generation cephalosporins in E. aerogenes. The broadened spectrum was caused by a new modification to the H-10 helix, which modified the active site.

The plasma membrane distribution and related biological activity of nucleoside transporter proteins (NTs) were investigated in human syncytiotrophoblast from term placenta using a variety of approaches, including nucleoside uptake measurements into vesicles from selected plasma membrane domains, NT immunohistochemistry, and subcellular localization (basal, heavy, and light apical membranes as well as raft-enriched membranes from the apical domain). In contrast with other epithelia, in this epithelium, we have identified the high-affinity pyrimidine-preferring human concentrative nucleoside transporter (hCNT) 1 as the only hCNT-type protein expressed at both the basal and apical membranes. hCNT1 localization in lipid rafts is also dependent on its subcellular localization in the apical plasma membrane, suggesting a complex cellular and regional expression. Overall, this result favors the view that the placenta is a pyrimidine-preferring nucleoside sink from both maternal and fetal sides, and hCNT1 plays a major role in promoting pyrimidine salvage and placental growth. This finding may be of pharmacological relevance, because hCNT1 is known to interact with anticancer nucleoside-derived drugs and other molecules, such as nicotine and caffeine, for which a great variety of harmful effects on placental and fetal development, including intrauterine growth retardation, have been reported.

The human placental syncytiotrophoblast (hSTB) is a polarized epithelial structure, without paracellular routes, forming the main barrier for materno–fetal exchange. There is ample evidence suggesting the presence of potassium (K + ) channels in the placental apical membrane; which could contribute to membrane potential and volume regulation. We have therefore examined the K + currents of isolated apical membranes from human term placenta using electrophysiological methods: reconstitution of ion channels from apical membranes into giant liposomes (single channel recordings, patch clamp method) or their functional transplantation into Xenopus laevis oocytes (total currents recording, voltage clamp method). Single channel recording experiments show the presence of K + channels in the hSTB microvillous membrane sensitive to Tetraethylammonium (TEA) and Barium (Ba +2 ). Patch current activity was diminished 50% and 70% by 20 mmol/L TEA and 5 mmol/L Ba +2 respectively. The more frequent conductance was approximately 73 pS, however several levels of current were detected suggesting the presence of more than one type of K + channel. In addition, sodium (Na + ) sensitivity was detected in the patch current thus, over 10 mmol/L Na + reduced the seal current to 38%. These results were corroborated by the total current experiments where the K + current elicited in injected oocytes with apical purified membrane was blocked by Ba +2 and TEA. The total current was also affected by Na + , becoming larger when a Na + -free solution was used. Our results show the existence of at least two types of Ba +2 -sensitive K + channels including a TEA sensitive sub-population, and some of them Na + sensitive K + channels. These channels could be the conductive pathways proposed previously for this cation in placental hSTB. Our novel contribution has been to successfully obtain K + channel recordings in systems suitable for electrophysiological studies of isolated apical membranes.

4 páginas, 1 tabla, 5 esquemas., The preparation of carbohydrate mimics in which the endocyclic oxygen has been replaced by a guanidine-type nitrogen atom is reported. The synthetic strategy involves the furanose --> piperidine rearrangement of 5-deoxy-5-guanidino-L-idose precursors. The reaction proceeds through elimination of water to give 3-oxopiperidines, which were isolated as the corresponding hydrates. Biological evaluation of the new glycomimetics evidenced a strong influence of the nature of the substituents at the nitrogen atoms on the glycosidase inhibitory properties., We thank the Spanish Ministerio de Educación y Ciencia (contract nos. CTQ2007-61180/PPQ and CTQ2006-15515-C02-01/BQU) and the Junta de Andalucía for financial support.

Human chorionic gonadotropin (hCG) is a key autocrine/paracrine regulator of placental syncytiotrophoblast, the transport epithelium of the human placenta. Syncytiotrophoblast hCG secretion is modulated by the partial pressure of oxygen (pO2), reactive oxygen species (ROS) and potassium (K+) channels. Here we test the hypothesis that K+ channels mediate the effects of pO2 and ROS on hCG secretion. Placental villous explants from normal term pregnancies were cultured for 6 days at 6% (normoxia), 21% (hyperoxia) or 1% (hypoxia) pO2. On days 3-5, explants were treated with 5mM 4-aminopyridine (4-AP) or tetraethylammonium (TEA), blockers of pO2-sensitive voltage-gated K+ (KV) channels, or ROS (10-1000μM H2O2). hCG secretion and lactate dehydrogenase (LDH) release, a marker of necrosis, were determined daily. At day 6, hCG and LDH were measured in tissue lysate and 86Rb (K+) efflux assessed to estimate syncytiotrophoblast K+ permeability. hCG secretion and 86Rb efflux were significantly greater in explants maintained in 21% pO2 than normoxia. 4-AP/TEA inhibited hCG secretion to a greater extent at 21% than 6% and 1% pO2, and reduced 86Rb efflux at 21% but not 6% pO2. LDH release and tissue LDH/hCG were similar in 6%, 21% and 1% pO2 and unaffected by 4-AP/TEA. H2O2 stimulated 86Rb efflux and hCG secretion at normoxia but decreased 86Rb efflux, without affecting hCG secretion, at 21% pO2. 4-AP/TEA-sensitive K+ channels participate in pO2-sensitive hCG secretion from syncytiotrophoblast. ROS effects on both hCG secretion and 86Rb efflux are pO2-dependent but causal links between the two remain to be established.

Con el objetivo de esclarecer algunos de los aspectos más significativos de los análisis fenomenológicos del dolor físico, en este trabajo pretendo mostrar que la fenomenología del dolor es un caso particular de análisis dentro del marco general de la fenomenología de la corporeidad. Para ello consideraré, por un lado, la relación entre las modalidades atencionales y la aparición del dolor y, por otro lado, la relación entre la aparición del dolor y la perturbación del esquema corporal. Por último, a fin de justificar mi tesis, trataré la relación entre atención y esquema corporal. With the aim of elucidating some important aspects of the phenomenological analysis of physical pain, I will show in this paper that Phenomenology of Pain constitutes a singular case within the general context of the Phenomenology of Body. In order to show that thesis, I will consider, first, the relationship between attentional modes and the birth of pain and, second, the relationship between birth of pain and body schema disturbance. Finally, with the objective of justifying mi own thesis, I will consider the relationship between attention and body schema.

Although the importance of DNA methylation-dependent gene expression to neuronal plasticity is well established, the dynamics of methylation and demethylation during the induction and expression of synaptic plasticity have not been explored. Here, we combined electrophysiological, pharmacological, molecular, and immunohistochemical approaches to examine the contribution of DNA methylation and the phosphorylation of Methyl-CpG-binding protein 2 (MeCP2) to synaptic plasticity. We found that, at twenty minutes after theta burst stimulation (TBS), the DNA methylation inhibitor 5-aza-2-deoxycytidine (5AZA) impaired hippocampal long-term potentiation (LTP). Surprisingly, after two hours of TBS, when LTP had become a transcription-dependent process, 5AZA treatment had no effect. By comparing these results to those in naive slices, we found that, at two hours after TBS, an intergenic region of the RLN gene was hypomethylated and that the phosphorylation of residue S80 of MeCP2 was decreased, while the phosphorylation of residue S421 was increased. As expected, 5AZA affected only the methylation of the RLN gene and exerted no effect on MeCP2 phosphorylation patterns. In summary, our data suggest that tetanic stimulation induces critical changes in synaptic plasticity that affects both DNA methylation and the phosphorylation of MeCP2. These data also suggest that early alterations in DNA methylation are sufficient to impair the full expression of LTP.

Issue Section: Research Letters., This work was supported by the Ministerio de Sanidad y Consumo, Instituto de Salud Carlos III (projects PI11-00934 and PI14/00940) and the Consejería de Innovación Ciencia y Empresa, Junta de Andalucía (P11-CTS-7730), Spain, the Plan Nacional de I+D+i 2008-2011 and the Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Economía y Competitividad, the Spanish Network for Research in Infectious Diseases (REIPI RD12/0015)—co-financed by the European Development Regional Fund ‘A Way to Achieve Europe’.

The aim of the study was to determine the interplay between the plasmid-mediated qepA2 gene and multiple chromosomally mediated fluoroquinolone resistance determinants in the development of fluoroquinolone resistance in Escherichia coli and its influence on bacterial fitness.E. coli ATCC 25922 and derived isogenic strains harbouring different chromosomally mediated fluoroquinolone resistance determinants were electroporated with pBK-CMV vector encoding QepA2. The MICs of fluoroquinolones were determined by standardized microdilution. The mutant prevention concentration (MPC) was evaluated. Bacterial fitness was analysed using ΔlacZ system competition assays.The ciprofloxacin MIC for strains harbouring the qepA2 gene was 4- to 8-fold higher compared with strains without the qepA2 gene. The qepA2 gene also increased the MPC of ciprofloxacin 4- to 16-fold. Combination of the qepA2 gene plus two to three additional mechanisms conferred a clinically relevant resistance level. The presence of the qepA2 gene was associated with fitness costs in strains with mutations in the gyrA and/or parC genes, although the presence of an additional deletion of the marR gene compensated for this fitness cost by increasing bacterial fitness by 5%-23%.The additive effect of chromosomally mediated fluoroquinolone resistance mechanisms and the qepA2 gene led to clinical levels of fluoroquinolone resistance. Under competitive conditions, the qepA2 gene had a biological cost in E. coli that was compensated for by the presence of an additional deletion in the marR gene.

A practical synthesis of reducing isourea-derived azasugar glycomimetics related to the indolizidine and trehazolin glycosidase inhibitor families with different pK(a) values is disclosed. The polyhydroxylated bicyclic system was built from readily accessible hexofuranose derivatives through a synthetic scheme that involves the preparation of a 5-deoxy-5-carbodiimido adduct by triphenylphosphine-mediated tandem Staudinger--aza-Wittig-type coupling of azide and isothiocyanate precursors, intramolecular cyclization of a transient vic-hydroxycarbodiimide derivative, and nucleophilic addition of the endocyclic nitrogen atom of the generated 2-amino-2-oxazoline intermediate, with a pseudo-C-nucleoside structure, to the masked aldehyde group of the monosaccharide. The last step is pH-dependent so that the final compounds can pivot between the furanose and the 2-oxaindolizidine forms. Nevertheless, the indolizidine tautomer having the R configuration at the aminoacetalic center, fitting the anomeric effect, was the only species detected in solution at neutral or slightly acidic pH when starting from solutions at basic pH. Glycosidase inhibition tests (K(i) values down to 1.9 microM) showed a marked dependence of the selectivity and potency toward alpha- and beta-glucosidases upon the nature of the substituent at the exocyclic isourea nitrogen, shifting from alpha- to beta-selectivity when going from hydrophilic to hydrophobic substituents. Enzyme inhibition is also pH dependent, supporting a dominant role for the uncharged form of the polyhydroxyiminoindolizidine system in the inhibition of beta-glucosidases.

The title indolizidine, a ring-modified analog of the potent mannosidase inhibitor (-)-swainsonine, has been prepared by intramolecular nucleophilic addition of the sp 2 -type nitrogen atom of a preformed 1,3-oxazine-2-thione heterocycle to the masked aldehyde group in a D-mannose precursor; the stereochemistry of the generated aminoacetalic stereocenter is governed by the generalized anomeric effect, the pseudoanomeric hydroxyl group adopting an axial orientation that mimics that of α-D-mannopyranoside aglycons.

Resumen Introduccion La resistencia combinada a quinolonas y β-lactamicos es frecuente en enterobacterias. La aparicion en enterobacterias de metalo-β-lactamasas y determinantes de resistencia a quinolonas mediada por plasmidos constituyen un problema emergente en nuestro pais. Metodos La sensibilidad se determino mediante tiras de E-test. Los genes de resistencia se detectaron mediante PCR y se procedio a la caracterizacion de los plasmidos portadores. Resultados En este estudio se describen 2 cepas (1 Klebsiella oxytoca y 1 Klebsiella pneumoniae) portadoras de los genes blaVIM-1 y qnrS2 en un plasmido transferible de 70-Kb aisladas en cultivos de vigilancia en el Hospital Universitario Virgen Macarena de Sevilla. Conclusion Esta es la primera asociacion de blaVIM-1 y qnrS2 en un mismo plasmido no tipable en nuestro centro.

Tandem Staudinger–aza-Wittig reaction of primary azidodeoxy sugars with triphenylphosphine–carbon disulfide affords the corresponding primary deoxyisothiocyanato sugars in high yield. No products arising from O→N acyl migration or formation of dimeric carbodiimides were observed. Interestingly, a polymer-supported triarylphosphine can advantageously replace triphenylphosphine, thus limiting the purification step to a simple filtration process. The reaction also allows the preparation of 5-deoxy-5-isothiocyanato sugars, a hitherto unknown class of compounds, from the corresponding azide precursors. Secondary sugar azides bearing the azido group at an endocyclic carbon atom afforded much lower isothiocyanation yields under these reaction conditions.

The aim of this study is to evaluate the effect of preeclampsia on the level of lipid peroxidation, activity and expression of both plasma membrane Ca(2+)- and Na(+), K(+)-ATPases in syncytiotrophoblast.The level of lipid peroxidation was estimated by measuring TBARS. ATPase activities were quantified by a colorimetric method measuring the amount of inorganic phosphate during the assay. Expression of Ca(2+)- and Na(+), K(+)-ATPases in syncytiotrophoblast plasma membranes and term placenta tissue sections was investigated using Western blot and immunohistochemistry, respectively.Our results show a higher level of lipid peroxidation of syncytiotrophoblast plasma membranes from preeclamptic, as compared to uncomplicated pregnant women. Preeclampsia also significantly reduced the activity of Ca(2+)- and Na(+), K(+)-ATPases; however, expression of both ATPases was unaffected.Our findings suggest that the reduction of Ca(2+)- and Na(+), K(+)-ATPase activities during preeclampsia could be at least partially due to an increased level of lipid peroxidation of the syncytiotrophoblast plasma membranes.

Objective. Assess transient gene expression of GUS in cassava (Manihot esculenta Crantz) leaves using Agrobacterium tumefaciens infiltration. Materials and methods. A. tumefaciens strains GV3101 and AGL1 containing pCAMBIA1305.2 were used to evaluate transient gene expression of β-glucuronidase (GUS). A. tumefaciens infiltration (agroinfiltration) was made using both leaves from in vitro and 1 month old greenhouse plants. Leaves were incubated in X-GLUC buffer, stained and photographed to detect GUS activity. Results. Agroinfiltration assays showed GUS transient expression in leaves of cassava varieties widely cultivated in the north coast and eastern savannah, MCOL2215 (Venezuelan) and CM6438-14 (Vergara), respectively. A. tumefaciens agressive strain AGL1 showed high efficiency inducing GUS expression in cassava leaves. Conclusions. We recommend using A. tumefaciens agressive strain AGL1 for agroinfiltration to assess transient expression in cassava leaves.

The aim of this study was to analyse the interplay among plasmid-mediated qnr genes, alone or in combination with multiple chromosomal-mediated fluoroquinolone (FQ) resistance determinants, susceptibility to FQs and bacterial fitness in an isogenic Escherichia coli collection.E. coli ATCC 25922 was used to modify or delete chromosomal genes. qnr genes were cloned into the pBK-CMV vector. The MICs of FQs were determined by microdilution. Mutant prevention concentration and frequency of mutants were evaluated. Bacterial fitness was analysed using ΔlacZ system competition assays using in vitro and in vivo models.The relationships between the number of resistance mutations and bacterial fitness were complex. With specific combinations of resistance mechanisms the addition of a new resistance mutation was shown to improve bacterial fitness. qnrA1 caused a decrease in fitness (7%-21%) while qnrS1 caused an increase in fitness (9%-21%) when combined with chromosomal mutations. We identified susceptible triple mutants in which the acquisition of a fourth resistance mutation significantly increased fitness and at the same time reached the clinical resistance level (the acquisition of qnrS1 in a S83L + D87N + ΔmarR genetic background). A strong correlation with the production of reactive oxygen species, as well as changes in susceptibility, was observed following treatment with ciprofloxacin.Our data indicate that there may be critical stages (depending on the genotype) in resistance development, including chromosomal- and plasmid-mediated mechanisms, at which some low-fitness mutants below the resistance breakpoint are able to evolve clinical resistance with just one or two mutations, and show increased fitness.

The quality of water for human consumption has been correlated with the occurrence of different diseases. Studying the relationship between these parameters would allow determining the impact of water quality on human health, and to direct preventative measures and promote environmental health.To analyze the quality of water intended for human consumption and its association with morbimortality in Colombia, 2008-2012.The database for surveillance of water quality was analyzed by means of descriptive statistics of the principal indicators (total coliforms, Escherichia coli , turbidity, color, pH, free residual chlorine and water quality risk index). The results were correlated with infant mortality and morbidity due to acute diarrheal diseases, foodborne diseases and hepatitis A. A risk map was prepared to identify those municipalities with the highest risk of water contamination and infant mortality.A high percentage of municipalities did not conform to existing standards for water potability values. Problems were identified that were related to presence of E. coli and total coliforms, as well as absence of free residual chlorine, a situation that was exacerbated in rural areas. Water quality showed a high correlation with infant mortality, highlighting its importance for children's health.Water quality was found to have an important impact on infant mortality. Improving water quality in Colombia will require policies that strengthen water supply systems in this country. Strengthening of environmental health surveillance programs is essential to guide actions aimed at improving water quality and exert a positive impact on health.

The placenta mediates maternal-fetal exchange and has historically been regarded as a passive conduit for nutrients. However, emerging evidence suggests that the placenta actively responds to nutritional and metabolic signals from the mother and the fetus. We propose that the placenta integrates a multitude of maternal and fetal nutritional cues with information from intrinsic nutrient-sensing signaling pathways to match fetal demand with maternal supply by regulating maternal physiology, placental growth, and nutrient transport. This process, which we have called placental nutrient sensing, ensures optimal allocation of resources between the mother and the fetus to maximize the chances for propagation of parental genes without jeopardizing maternal health. We suggest that these mechanisms have evolved because of the evolutionary pressures of maternal undernutrition, which result in decreased placental growth and down-regulation of nutrient transporters, thereby limiting fetal growth to ensure maternal survival. These regulatory loops may also function in response to maternal overnutrition, leading to increased placental growth and nutrient transport in cases of maternal obesity or gestational diabetes. Thus, placental nutrient sensing modulates maternal-fetal resource allocation to increase the likelihood of reproductive success. This model implies that the placenta plays a critical role in mediating fetal programming and determining lifelong health.

Recent technical advances have opened up new avenues in molecular science with great impact on both in vitro and in vivo applications. Next generation sequencing has become available to a greater extent and advanced molecular techniques and methods within RNA/DNA-editing, barcoding and generation of high diversity DNA libraries are constantly evolving, giving rise to novel applications and discoveries.The aim of this study was to establish an unbiased, totally randomized screening assay for RNA-editing that can be universally utilized. By creating high resolution DNA-libraries from genomic sequences where each fragment is uniquely labelled with a DNA-barcode, we were able to map the biological function of each complete sequence. Here, we have utilized this approach to identify the most promising intronic sequences for RNA-editing. The screening assay was performed by deep sequencing of a lentiviral library, where each fragment was linked to a corresponding barcode. mRNA from transduced cells in vitro and in vivo was sequenced to identify and count the expressed barcodes, thus we could identify specific fragments suitable for RNA-editing. View Large Image | Download PowerPoint SlideRNA-editing was achieved via trans-splicing, where two different pre-mRNAs (one endogenous and one virally derived) hybridize through complementary sequences and form one mRNA. In trans-splicing, only the downstream segment of the mRNA is replaced, thus the gene is still under the control of its native promoter and other endogenous regulatory elements remain unaltered. Trans-splicing is inherently specific since the part of the mRNA that is delivered requires endogenous expression to exert an effect. View Large Image | Download PowerPoint SlideBy combining advanced cloning methods, DNA-barcoding and Next generation sequencing, we could screen extensive DNA libraries and select fragments highly efficient in RNA-editing. Our method can be used for correction of mutations, protein tracing and cell type identification.

Regulation of human placental syncytiotrophoblast renewal by cytotrophoblast migration, aggregation/fusion and differentiation is essential for successful pregnancy. In several tissues, these events are regulated by intermediate conductance Ca2+-activated K+ channels (IKCa), in part through their ability to regulate cell volume. We used cytotrophoblasts in primary culture to test the hypotheses that IKCa participate in the formation of multinucleated syncytiotrophoblast and in syncytiotrophoblast volume homeostasis. Cytotrophoblasts were isolated from normal term placentas and cultured for 66 h. This preparation recreates syncytiotrophoblast formation in vivo, as mononucleate cells (15 h) fuse into multinucleate syncytia (66 h) concomitant with elevated secretion of human chorionic gonadotropin (hCG). Cells were treated with the IKCa inhibitor TRAM-34 (10 µM) or activator DCEBIO (100 µM). Culture medium was collected to measure hCG secretion and cells fixed for immunofluorescence with anti-IKCa and anti-desmoplakin antibodies to assess IKCa expression and multinucleation respectively. K+ channel activity was assessed by measuring 86Rb efflux at 66 h. IKCa immunostaining was evident in nucleus, cytoplasm and surface of mono- and multinucleate cells. DCEBIO increased 86Rb efflux 8.3-fold above control and this was inhibited by TRAM-34 (85%; p

Objectives: The aim of this study was to evaluate the impact of qnrA1, qnrB1 and qnrS1 on the in vivo efficacies of ciprofloxacin and levofloxacin in an experimental model of pneumonia caused by Escherichia coli. Methods: Two isogenic groups of E. coli transformants, based on two ATCC 25922 strains, with or without the GyrA mutation Ser83Leu, and carrying qnrA1, qnrB1 or qnrS1, were used in an experimental pneumonia model. The efficacies of ciprofloxacin (40 mg/kg/day) and levofloxacin (50 and 150 mg/kg/day) were evaluated. Results: For the pneumonia caused by the parental strains lacking qnr genes, both fluoroquinolones significantly (P,0.05) reduced the bacterial lung concentration by .7 log10 cfu/g against E. coli ATCC/pBK and between 5.09 and 6.34 log10 cfu/g against E. coli ATCC-S83L/pBK. The presence of any qnr genes in the strains of both isogenic groups diminished the reduction of bacterial lung concentration with any therapy (P,0.05). Furthermore, all therapeutic schemes reduced the percentage of positive blood cultures in both isogenic groups (P,0.05). Finally, the survival results suggest a higher mortality with the strains expressing qnr genes. Conclusions: The presence of qnrA1, qnrB1 and qnrS1 in E. coli reduced the efficacy of ciprofloxacin and levofloxacin in a murine pneumonia model.

Data quality is a difficult notion to define precisely, and different communities have different views and understandings of the subject. This causes confusion, a lack of harmonization of data across communities and omission of vital quality information. For some existing data infrastructures, data quality standards cannot address the problem adequately and cannot fulfil all user needs or cover all concepts of data quality. In this study, we discuss some philosophical issues on data quality. We identify actual user needs on data quality, review existing standards and specifications on data quality, and propose an integrated model for data quality in the field of Earth observation (EO). We also propose a practical mechanism for applying the integrated quality information model to a large number of datasets through metadata inheritance. While our data quality management approach is in the domain of EO, we believe that the ideas and methodologies for data quality management can be applied to wider domains and disciplines to facilitate quality-enabled scientific research.

Poly-lactide-co-glycolide (PLGA) microparticles emerged as one of the most promising strategies to achieve site-specific drug delivery. Although these microparticles have been demonstrated to be effective in several wound healing models, their potential in cardiac regeneration has not yet been fully assessed. The present work sought to explore PLGA microparticles as cardiac drug delivery systems. PLGA microparticles were prepared by Total Recirculation One-Machine System (TROMS) after the formation of a multiple emulsion. Microparticles of different size were prepared and characterized to select the most suitable size for intramyocardial administration. Next, the potential of PLGA microparticles for administration in the heart was assessed in a MI rat model. Particle biodegradation over time and myocardial tissue reaction were studied by routine staining and confocal microscopy. Results showed that microparticles with a diameter of 5 μm were the most compatible with intramyocardial administration in terms of injectability through a 29-gauge needle and tissue response. Particles were present in the heart tissue for up to three months post-implantation and no particle migration towards other solid organs was observed, demonstrating good myocardial retention. CD68 immunolabeling revealed 31%, 47% and below 4% microparticle uptake by macrophages one week, one month and three months after injection, respectively (P

All participating laboratories are members of ProteoRed-ISCIII.-- et al., The Spanish team of the Human Proteome Project (SpHPP) marked the annotation of Chr16 and data analysis as one of its priorities. Precise annotation of Chromosome 16 proteins according to C-HPP criteria is presented. Moreover, Human Body Map 2.0 RNA-Seq and Encyclopedia of DNA Elements (ENCODE) data sets were used to obtain further information relative to cell/tissue specific chromosome 16 coding gene expression patterns and to infer the presence of missing proteins. Twenty-four shotgun 2D-LC–MS/MS and gel/LC–MS/MS MIAPE compliant experiments, representing 41% coverage of chromosome 16 proteins, were performed. Furthermore, mapping of large-scale multicenter mass spectrometry data sets from CCD18, MCF7, Jurkat, and Ramos cell lines into RNA-Seq data allowed further insights relative to correlation of chromosome 16 transcripts and proteins. Detection and quantification of chromosome 16 proteins in biological matrices by SRM procedures are also primary goals of the SpHPP. Two strategies were undertaken: one focused on known proteins, taking advantage of MS data already available, and the second, aimed at the detection of the missing proteins, is based on the expression of recombinant proteins to gather MS information and optimize SRM methods that will be used in real biological samples. SRM methods for 49 known proteins and for recombinant forms of 24 missing proteins are reported in this study., This work was supported by: ProteoRed and the Carlos III National Health Institute Agreement, ProteoRed-ISCIII; the agreement between FIMA and the “UTE project CIMA”; grants SAF2011-29312 from Ministerio de Ciencia e Innovación and ISCIII-RETIC RD06/0020 to FJC and EU FP7 grant ProteomeXchange [grant number 260558]. APM and DTM have been funded by Spanish grants from Ministerio de Ciencia e Innovación BIO2010-17527 and the Government of Madrid (P2010/BMD-2305). BBVA Foundation for its support to HUPO initiatives.

Myocardial infarction (MI) is the leading cause of death worldwide and extensive research has therefore been performed to find a cure. Neuregulin-1 (NRG) is a growth factor involved in cardiac repair after MI. We previously described how biocompatible and biodegradable microparticles, which are able to release NRG in a sustained manner, represent a valuable approach to avoid problems related to the short half-life after systemic administration of proteins. The effectiveness of this strategy could be improved by combining NRG with several cytokines involved in cardiac regeneration. The present study investigates the potential feasibility of using NRG-releasing particle scaffold combined with adipose derived stem cells (ADSC) as a multiple growth factor delivery-based tissue engineering strategy for implantation in the infarcted myocardium. NRG-releasing particle scaffolds with a suitable size for intramyocardial implantation were prepared by TROMS. Next, ADSC were adhered to particle scaffolds and their potential for heart administration was assessed in a MI rat model. NRG was successfully encapsulated reaching encapsulation efficiencies of 92.58 ± 3.84 %. NRG maintained its biological activity after the microencapsulation process. ADSC cells adhered efficiently to particle scaffolds within a few hours. The ADSC-cytokine delivery system developed proved to be compatible with intramyocardial administration in terms of injectability through a 23-gauge needle and tissue response. Interestingly, ADSC-scaffolds were present in the peri-infarted tissue two weeks after implantation. This proof of concept study provides important evidence required for future effectiveness studies and for the translation of this approach.

Using a non-experimental, transverse, co-relational design, we assess the relationship between optimism and quality of life (QoL) in adults aged between 18 and 65 years. Two questionnaires were administered: WHOQOLBREF, which assessed QoL, and an instrument that assessed trait–state optimism. A total of 1190 subjects from various public and private high schools, institutions and commercial centers in the city of Antofagasta, Chile participated. The mean sample age was 40.0 years and 50.0% of respondents were male. There is a relationship between QoL and state optimism, with males demonstrating a higher QoL. Differences in optimism were observed only in age groups in which younger persons scored less than older persons. The results obtained indicate that there is a partial relationship between QoL and optimism, with state optimism having a significant effect on QoL.

Workshops are an important part of the IFPA annual meeting as they allow for discussion of specialized topics. At IFPA meeting 2012 there were twelve themed workshops, four of which are summarized in this report. These workshops related to various aspects of placental biology: 1) epigenetics and imprinting in the placenta; 2) growth factors and villous trophoblast differentiation; 3) role of the placenta in regulating fetal exposure to xenobiotics during pregnancy; 4) infection and the placenta.

The GEOSS Common Infrastructure (GCI) provides a Clearinghouse (the GEOSS registry and metadata catalogue) and a GEOPortal to discover and visualize EO data in an integrated, standardized and interactive way, as well as broadly use it by the scientific community when dealing with representation and modeling of Earth Systems. EO data sources are ideally elaborated following quality assessment procedures, resulting in quality estimates and other related indicators. The objective of this indicators is to allow users deciding about data fitness for a purpose, but in practice systems providing methods to distribute, show and exploit this producer quality information in a standard and interoperable way are rarely used. This work aims to extract information about data quality from GCI metadata and analyze the obtained results. Additionally, an XML specific tool is able to quick and visually punctuating the metadata that refers to quality.

Combined resistance to quinolones and β-lactams is common in Enterobacteriaceae. The appearance in enterobacteria coding for metallo-β-lactamases and determinants of plasmid-mediated quinolone resistance are an emerging problem in our country.The susceptibility was determined by E-test. The resistance genes were detected by PCR and the corresponding plasmids were characterised.This study describes 2 strains (1 Klebsiella oxytoca, 1 Klebsiella pneumoniae) carrying the genes qnrS2 and blaVIM-1 in a transferable plasmid of 70-Kb isolated in surveillance cultures at the University Hospital Virgen Macarena in Seville.This is the first combination of qnrS2 and bla(VIM-1) on the same non-typeable plasmid isolated in our centre.

Workshops are an important part of the IFPA annual meeting. At IFPA Meeting 2010 there were twelve themed workshops, six of which are summarized in this report. 1. The immunology workshop focused on normal and pathological functions of the maternal immune system in pregnancy. 2. The transport workshop dealt with regulation of ion and water transport across the syncytiotrophoblast of human placenta. 3. The epigenetics workshop covered DNA methylation and its potential role in regulating gene expression in placental development and disease. 4. The vascular reactivity workshop concentrated on methodological approaches used to study placental vascular function. 5. The workshop on epitheliochorial placentation covered current advances from in vivo and in vitro studies of different domestic species. 6. The proteomics workshop focused on a variety of techniques and procedures necessary for proteomic analysis and how they may be implemented for placental research.

Background Patients with chronic pain frequently display comorbid depression, but the impact of this concurrence is often underestimated and mistreated. The aim of this study was to determine the prevalence of unrecognized major depression and other mood disorders and comorbid unexplained chronic pain in primary care settings and to explore the associated factors. Also, to compare the use of health services by patients with unexplained chronic pain, both with and without mood disorder comorbidity. Methods A cross-sectional study was carried out in a sample of primary care centers. 3189 patients consulting for "unexplained chronic pain" were assessed by the Visual Analogue Scales (VAS) and the Primary Care Evaluation of Mental Disorders (PRIME-MD) questionnaire. Results We report: a) a high prevalence of unrecognized mood disorders in patients suffering from unexplained chronic pain complaints (80.4%: CI 95%: 79.0%; 81.8%); b) a greater susceptibility of women to mood disorders (OR adjusted = 1.48; CI 95%:1.22; 1.81); c) a direct relationship between the prevalence of mood disorders and the duration of pain (OR adjusted = 1.01; CI 95%: 1.01; 1.02) d) a higher comorbidity with depression if the pain etiology was unknown (OR adjusted = 1.74; CI 95%: 1.45; 2.10) and, e) an increased use of health care services in patients with such a comorbidity (p < 0.0001). Conclusions The prevalence of undiagnosed mood disorders in patients with unexplained chronic pain in primary care is very high, leading to dissatisfaction with treatment processes and poorer outcomes. Consequently, it seems necessary to explore this condition more regularly in general practice in order to reach accurate diagnoses and to select the appropriate treatment.

We assessed agreement among 3 assays for determining susceptibility to imipenem (IMP) and meropenem (MPM) of Acinetobacter baumannii (Ab) heteroresistant to carbapenems (Hr-CP).Thirty Ab clinical isolates belonging to 6 clones (REP-PCR) were studied, in which IMP and MPM MICs were betweenor = 1 andor = 8 mg/l by Wider system. MICs determined by Wider were compared with those obtained by microdilution (MD) and E-test. Errors in the clinical category were determined considering MD as the reference method.Twenty-five Ab were Hr-CP (growth of resistant colonies within the inhibition area of disks and E-test strips). Agreement for the MICs (+/- 1log(2)) in Hr-CP colonies was as follows: Wider vs. MD (96% IMP, 100% MPM), Wider vs. E-test (50% IMP, 64% MPM), MD vs. E-test (64% IMP, 60% MPM). Major errors were not observed. Minor errors and moderate errors for IMP included Wider vs. MD (40% and 0%), E-test vs. MD (40% and 12%), and disks vs. MD (36% and 8%). Agreement for the MICs considering colonies growing within the inhibition areas (E-test and disks) was Wider vs. E-test (8% IMP, 12% MPM), and MD vs. E-test (8% IMP and MPM). Major errors were not observed in this case either; minor errors and moderate errors for IMP were seen in E-test vs. MD (40% and 48%), and disks vs. MD (40% and 48%).Susceptibility testing methods based on microdilution (Wider system and MD) are not useful to detect subpopulations of Ab Hr-CP.

It is known that human syncytiotrophoblast (hSCT) actively transports more than 80% of the Ca2+ that goes from maternal to fetal circulation. Transepithelial transport of Ca2+ is carried out through channels, transporters and exchangers located in both microvillous (MVM) and basal (BM) plasma membranes. The plasma membrane Ca-ATPase (PMCA) is the most important mechanism of Ca2+ homeostasis control in the human placenta. In this work, we reexamined the distribution of PMCA in isolated hSCT of term placenta. The PMCA activity was determined in isolated hSCT plasma membranes. A partial characterization of the PMCA activity was performed, including an evaluation of the sensitivity of this enzyme to an in vitro induced lipid peroxidation. Expression of the PMCA in hSCT plasma membranes and tissue sections was investigated using Western blots and immunohistochemistry, respectively. Our study demonstrates, for the first time, a correlation between the activity and structural distribution of PMCA in both MVM and BM of hSCT. It also demonstrates a higher PMCA activity and expression in MVM as compared to BM. Finally, PMCA4 seems to be preferentially distributed in both hSCT plasma membranes, while PMCA1 is shown to be present in the hSCT homogenate. However, the membrane fractions did not show any PMCA1 labeling. Our results must be taken into account in order to propose a new model for the transport of calcium across the hSCT.

The functional expression of calcium channels has been scarcely studied in human placental syncytiotrophoblast. We have presently sought to characterize Ca 2+ currents of the healthy syncytiotrophoblast basal membrane using purified basal membranes reconstituted in giant liposomes subjected to patch-clamp recordings. We detected presence of channels with high permeability to Ca 2+ (relative PCa/PK up to 99.5) using K + solutions in symmetric conditions. Recordings performed in Ba 2+ gradients showed Ba 2+ -conducting channels in 100% of experiments. Ba 2+ total patch currents were consistently blocked by addition of NiCl 2 , Nifedipine (L-type voltage-gated calcium channel blocker) or Ruthenium Red (TRPV5–TRPV6 channel blocker); Nifedipine and Ruthenium Red exerted a synergic blocking effect on Ba 2+ total patch currents. Immunohistochemistry of placental villi sections evidenced presence of α 1 subunit of voltage-gated calcium channels and TRPV5–TRPV6 channels in basal and apical syncytiotrophoblast plasma membranes; these three calcium channels were also detected in purified basal and apical fractions using Western blot. These results show the presence of three types of calcium channels in the syncytiotrophoblast basal membrane by both functional and molecular means. These basal membrane calcium channels would not be directly involved in mother-to-fetus Ca 2+ transport, but could participate in other relevant trophoblast processes, such as exocytosis and Ca 2+ transport regulation.

11 páginas, 2 figuras, 1 tabla, 4 esquemas., The feasibility of the intramolecular nucleophilic addition of the nitrogen atom in cyclic (thio)carbamates with a pseudo-C-nucleoside structure to the masked carbonyl group in aldose precursors in the synthesis of reducing (i.e., 5-hydroxy)6-oxaindolizidine frameworks is illustrated by the preparation of the 6-epi, 7-epi, 8-epi and 6,8a-di-epi diastereomers of the potent glycosidase inhibitor (+)-castanospermine. In all cases, the increased anomeric effect caused by the high sp2 character of the pseudoamide-type nitrogen atom resulted in the pseudoanomeric hydroxy group being anchored in an axial orientation in aqueous solution, as in the aglycons in α-glycosides. These analogs of the natural alkaloid showed a higher selectivity in the inhibition of α-glucosidases. Structure/glycosidase inhibitory activity studies indicated that inversion of any hydroxy group resulted in a dramatic decrease in the inhibition potency, confirming the critical importance of a correct hydroxylation profile. In the case of (+)-8-epi-6-oxacastanospermine derivatives, with a hydroxylation profile with a structural complementarity to that of D-galactose, a moderate but very selective inhibition of α-galactosidase was observed, supporting the importance of a defined configuration at pseudoanomeric centres for anomeric specificity., We thank the Spanish Ministerio de Educación y Ciencia for financial support (contract numbers BQU2003–00937 and CTQ2004–05854/BQU).