Halogen Bonds in Clathrate Cages: A Real Space Perspective

We thank the Spanish MINECO, grant CTQ2015-65790-P, and theFICYT, gran

Development of peptide-based lineage-specific serology for chronic Chagas disease: geographical and clinical distribution of epitope recognition.

BACKGROUND: Chagas disease, caused by infection with the protozoan Trypanosoma cruzi, remains a serious public health issue in Latin America. Genetically diverse, the species is sub-divided into six lineages, known as TcI-TcVI, which have disparate geographical and ecological distributions. TcII, TcV, and TcVI are associated with severe human disease in the Southern Cone countries, whereas TcI is associated with cardiomyopathy north of the Amazon. T. cruzi persists as a chronic infection, with cardiac and/or gastrointestinal symptoms developing years or decades after initial infection. Identifying an individual's history of T. cruzi lineage infection directly by genotyping of the parasite is complicated by the low parasitaemia and sequestration in the host tissues. METHODOLOGY/PRINCIPAL FINDINGS: We have applied here serology against lineage-specific epitopes of the T. cruzi surface antigen TSSA, as an indirect approach to allow identification of infecting lineage. Chagasic sera from chronic patients from a range of endemic countries were tested by ELISA against synthetic peptides representing lineage-specific TSSA epitopes bound to avidin-coated ELISA plates via a biotin labelled polyethylene glycol-glycine spacer to increase rotation and ensure each amino acid side chain could freely interact with their antibodies. 79/113 (70%) of samples from Brazil, Bolivia, and Argentina recognised the TSSA epitope common to lineages TcII/TcV/TcVI. Comparison with clinical information showed that a higher proportion of Brazilian TSSApep-II/V/VI responders had ECG abnormalities than non-responders (38% vs 17%; p<0.0001). Among northern chagasic sera 4/20 (20%) from Ecuador reacted with this peptide; 1/12 Venezuelan and 1/34 Colombian samples reacted with TSSApep-IV. In addition, a proposed TcI-specific epitope, described elsewhere, was demonstrated here to be highly conserved across lineages and therefore not applicable to lineage-specific serology. CONCLUSIONS/SIGNIFICANCE: These results demonstrate the considerable potential for synthetic peptide serology to investigate the infection history of individuals, geographical and clinical associations of T. cruzi lineages

SPROUTY-2 represses the epithelial phenotype of colon carcinoma cells via upregulation of ZEB1 mediated by ETS1 and miR-200/miR-150

SPROUTY-2 (SPRY2) is a modulator of tyrosine kinase receptor signaling with receptor- and cell type-dependent inhibitory or enhancing effects. Studies on the action of SPRY2 in major cancers are conflicting and its role remains unclear. Here we have dissected SPRY2 action in human colon cancer. Global transcriptomic analyses show that SPRY2 downregulates genes encoding tight junction proteins such as claudin-7 and occludin and other cell-to-cell and cell-to-matrix adhesion molecules in human SW480- ADH colon carcinoma cells. Moreover, SPRY2 represses LLGLL2/HUGL2, PATJ1/INADL and ST14, main regulators of the polarized epithelial phenotype, and ESRP1, an epithelial-to-mesenchymal transition (EMT) inhibitor. A key action of SPRY2 is the upregulation of the major EMT inducer ZEB1, as these effects are reversed by ZEB1 knock-down by means of RNA interference. Consistently, we found an inverse correlation between the expression level of claudin-7 and those of SPRY2 and ZEB1 in human colon tumors. Mechanistically, ZEB1 upregulation by SPRY2 results from the combined induction of ETS1 transcription factor and the repression of microRNAs (miR-200 family, miR-150) that target ZEB1 RNA. Moreover, SPRY2 increased AKT activation by epidermal growth factor (EGF) whereas AKT and also Src inhibition reduced the induction of ZEB1. Altogether, these data suggest that AKT and Src are implicated in SPRY2 action. Collectively, these results show a tumorigenic role of SPRY2 in colon cancer that is based on the dysregulation of tight junction and epithelial polarity master genes via upregulation of ZEB1. The dissection of the mechanism of action of SPRY2 in colon cancer cells is important to understand the upregulation of this gene in a subset of patients with this neoplasia that have poor prognosis.This study was supported by the Ministerio de Economía y Competitividad of Spain and Fondo Europeo de Desarrollo Regional (FEDER) (grant SAF2013-43468-R to A.M., SAF2011-29530 to F.X.R.); FEDERInstituto de Salud Carlos III (RD12/0036/0021 to A.M. and J.M.R., RD12/0036/0034 to F.X.R., RD12/0036/0016 to M.S., RD12/0036/0012 to H.G.P., RD06/0020/0003, PS09/00562 and PI13/00703 to J.M.R.); Comunidad de Madrid (S2010/BMD-2344 Colomics2 to A.M.); Fundación Científica de la Asociación Española contra el Cáncer (to J.M.R.); U.S. Department of Defense (CA093471 and CA110602 to E.H.); National Institutes of Health/National Cancer Institute (1R01CA155234-01 to E.H.); National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (1R21AR062239-01 to E.H.); and the Melanoma Research Alliance (to E. H.)

Moisture Compatibility of Portland Stones and other Oolitic Limestones

Approximately 30% of Europe's building stock comprises historic buildings, with the UK having a significant contribution of Portland stone structures. However, difficulties in sourcing the original material have led to the need for compatible material substitutes. Currently, the level of compatibility required between original and substitute material has not been quantified, and the approach has been to evaluate each case separately. This research aims to answer the question of how compatible is compatible enough, specifically in relation to moisture risk, and provide a guiding framework for the heritage conservation sector. The project uses a sensitivity analysis to quantify the compatibility of Portland Stone and other Oolitic limestones, considering porosity and age of the original/substitute stone as sensitive parameters when selecting compatible strategies for repair and reconstructio

Culture-free genome-wide locus sequence typing (GLST) provides new perspectives on Trypanosoma cruzi dispersal and infection complexity

El análisis del polimorfismo genético es una poderosa herramienta para la vigilancia epidemiológica y investigar. Sin embargo, la inferencia poderosa de la variación genética del patógeno es a menudo restringido por el acceso limitado al ADN objetivo representativo, especialmente en el estudio de especies parásitas obligadas para las cuales el cultivo ex vivo requiere muchos recursos o es propenso a sesgos. Los métodos modernos de captura de secuencias permiten analizar directamente la variación genética de los patógenos del material del huésped/vector, pero a menudo son demasiado complejos y costosos para entornos de escasos recursos donde prevalecen las enfermedades infecciosas. Este estudio propone un método sencillo y rentable Herramienta de tipificación de secuencias de locus de todo el genoma (GLST) basada en la amplificación paralela masiva de puntos críticos de información en todo el genoma del patógeno objetivo. el multiplexado La reacción en cadena de la polimerasa amplifica cientos de objetivos genéticos diferentes definidos por el usuario en un único tubo de reacción y la posterior limpieza basada en gel de agarosa y código de barras completan la preparación de la biblioteca por menos de 4 USD por muestra. Nuestro estudio genera un modelo flexible Flujo de trabajo de diseño de panel de imprimación GLST para Trypanosoma cruzi, el agente parásito de Chagas enfermedad. Aplicamos con éxito nuestro panel GLST de 203 objetivos a extractos nómicos metagénicos directos y sin cultivo de vectores triatominos que contienen un mínimo de 3,69 pg/μl de ADN de T. cruzi y elaborar más sobre el rendimiento del método mediante la secuenciación de bibliotecas GLST de T. cruzi clones de referencia que representan unidades de tipificación discretas (DTU) TcI, TcIII, TcIV, TcV y TcVI. Los 780 sitios SNP que identificamos en el conjunto de muestras distinguen parásitos de forma repetitiva infectar vectores simpátricos y detectar correlaciones entre distancias genéticas y geográficas a escala regional (< 150 km), así como continental. Los marcadores también separan claramente TcI, TcIII, TcIV y TcV + TcVI y parecen distinguir infecciones multiclonales dentro de TcI. Discutimos las ventajas, limitaciones y perspectivas de nuestro método a través de un espectro de la investigación epidemiológica.Analysis of genetic polymorphism is a powerful tool for epidemiological surveillance and research. Powerful inference from pathogen genetic variation, however, is often restrained by limited access to representative target DNA, especially in the study of obli gate parasitic species for which ex vivo culture is resource-intensive or bias-prone. Mod ern sequence capture methods enable pathogen genetic variation to be analyzed directly from host/vector material but are often too complex and expensive for resource-poor set tings where infectious diseases prevail. This study proposes a simple, cost-effective ‘genome-wide locus sequence typing’ (GLST) tool based on massive parallel amplifica tion of information hotspots throughout the target pathogen genome. The multiplexed polymerase chain reaction amplifies hundreds of different, user-defined genetic targets in a single reaction tube, and subsequent agarose gel-based clean-up and barcoding com pletes library preparation at under 4 USD per sample. Our study generates a flexible GLST primer panel design workflow for Trypanosoma cruzi, the parasitic agent of Chagas disease. We successfully apply our 203-target GLST panel to direct, culture-free metage nomic extracts from triatomine vectors containing a minimum of 3.69 pg/μl T. cruzi DNA and further elaborate on method performance by sequencing GLST libraries from T. cruzi reference clones representing discrete typing units (DTUs) TcI, TcIII, TcIV, TcV and TcVI. The 780 SNP sites we identify in the sample set repeatably distinguish parasites infecting sympatric vectors and detect correlations between genetic and geographic dis tances at regional (< 150 km) as well as continental scales. The markers also clearly sep arate TcI, TcIII, TcIV and TcV + TcVI and appear to distinguish multiclonal infections within TcI. We discuss the advantages, limitations and prospects of our method across a spectrum of epidemiological research

Four cation-selective transporters contribute to apical uptake and accumulation of metformin in Caco-2 cell monolayerss

Metformin is the frontline therapy for type II diabetes mellitus. The oral bioavailability of metformin is unexpectedly high, between 40 and 60%, given its hydrophilicity and positive charge at all physiologic pH values. Previous studies in Caco-2 cell monolayers, a cellular model of the human intestinal epithelium, showed that during absorptive transport metformin is taken up into the cells via transporters in the apical (AP) membrane; however, predominant transport to the basolateral (BL) side occurs via the paracellular route because intracellular metformin cannot egress across the BL membrane. Furthermore, these studies have suggested that the AP transporters can contribute to intestinal accumulation and absorption of metformin. Transporter-specific inhibitors as well as a novel approach involving a cocktail of transporter inhibitors with overlapping selectivity were used to identify the AP transporters that mediate metformin uptake in Caco-2 cell monolayers; furthermore, the relative contributions of these transporters in metformin AP uptake were also determined. The organic cation transporter 1, plasma membrane monoamine transporter (PMAT), serotonin reuptake transporter, and choline high-affinity transporter contributed to approximately 25%, 20%, 20%, and 15%, respectively, of the AP uptake of metformin. PMAT-knockdown Caco-2 cells were constructed to confirm the contribution of PMAT in metformin AP uptake because a PMAT-selective inhibitor is not available. The identification of four intestinal transporters that contribute to AP uptake and potentially intestinal absorption of metformin is a significant novel finding that can influence our understanding of metformin pharmacology and intestinal drug-drug interactions involving this highly prescribed drug

Organic Cation Transporter 1 (OCT1/mOct1) Is Localized in the Apical Membrane of Caco-2 Cell Monolayers and Enterocytes

Organic cation transporters (OCTs) are members of the solute carrier 22 family of transporter proteins that are involved in absorption, distribution, and excretion of organic cations. OCT3 is localized in the apical (AP) membrane of enterocytes, but the literature is ambiguous about OCT1 (mOct1) localization, with some evidence suggesting a basolateral (BL) localization in human and mouse enterocytes. This is contrary to our preliminary findings showing AP localization of OCT1 in Caco-2 cell monolayers, an established model of human intestinal epithelium. Therefore, this study aims at determining the localization of OCT1 (mOct1) in Caco-2 cells, and human and mouse enterocytes. Functional studies using OCT1-specific substrate pentamidine showed transporter-mediated AP but not BL uptake in Caco-2 cells and human and mouse intestinal tissues. OCT1 inhibition decreased AP uptake of pentamidine by ∼50% in all three systems with no effect on BL uptake. A short hairpin RNA-mediated OCT1 knockdown in Caco-2 cells decreased AP uptake of pentamidine by ∼50% but did not alter BL uptake. Immunostaining and confocal microscopy in all three systems confirmed AP localization of OCT1 (mOct1). Our studies unequivocally show AP membrane localization of OCT1 (mOct1) in Caco-2 cells and human and mouse intestine. These results are highly significant as they will require reinterpretation of previous drug disposition and drug-drug interaction studies where conclusions were drawn assuming BL localization of OCT1 in enterocytes. Most importantly, these results will require revision of the regulatory guidance for industry in the United States and elsewhere because it has stated that OCT1 is basolaterally localized in enterocytes

Differential effects of natural palm oil, chemically- and enzymatically-modified palm oil on weight gain, blood lipid metabolites and fat deposition in a pediatric pig model

<p>Abstract</p> <p>Background</p> <p>Increasing prevalence of obesity and overweight in the Western world, continue to be a major health threat and is responsible for increased health care costs. Dietary intervention studies show a strong positive association between saturated fat intake and the development of obesity and cardiovascular disease. This study investigated the effect of positional distribution of palmitic acid (Sn-1, 2 & 3) of palm oil on cardiovascular health and development of obesity, using weaner pigs as a model for young children.</p> <p>Methods</p> <p>Male and female weaner piglets were randomly allocated to 4 dietary treatment groups: 1) pork lard (LRD); 2) natural palm olein (NPO); 3) chemically inter-esterified PO (CPO) and 4) enzymatically inter-esterified PO (EnPO) as the fat source. Diets were formulated with 11% lard or with palm olein in order to provide 31% of digestible energy from fat in the diet and were balanced for cholesterol, protein and energy across treatments.</p> <p>Results</p> <p>From 8 weeks onwards, pigs on EnPO diet gained (P < 0.05) more weight than all other groups. Feed conversion efficiency (feed to gain) over the 12 week experimental period did not vary between treatment groups. Plasma LDL-C content and LDL-C/HDL-C ratio in pigs fed natural PO tended to be lower compared to all other diets. The natural PO lowered (P < 0.02) the plasma triglyceride (TG) content relative to the lard or EnPO diets, but was not different from the CPO diet. The natural PO diet was associated with lower (P < 0.05) saturated fat levels in subcutaneous adipose tissue than the CPO and EnPO diets that had lower saturated fat levels than the lard diet. Female pigs had lower lean and higher fat and fat:lean ratio in the body compared with male pigs. No difference in weight gain or blood lipid parameters was observed between sexes.</p> <p>Conclusions</p> <p>The observations on plasma TG, muscle and adipose tissue saturated fatty acid contents and back fat (subcutaneous) thickness suggest that natural palm oil may reduce deposition of body fat. In addition, dietary supplementation with natural palm oil containing palmitic acid at different positions in meat producing animals may lead to the production of meat and meat products with lower saturated fats. An increase in fat content and a decrease in lean content in female pigs resulted in an increased body fat:lean ratio but gender had no effect on blood lipid parameters or insulin concentrations.</p

Culture-free genome-wide locus sequence typing (GLST) provides new perspectives on Trypanosoma cruzi dispersal and infection complexity.

Analysis of genetic polymorphism is a powerful tool for epidemiological surveillance and research. Powerful inference from pathogen genetic variation, however, is often restrained by limited access to representative target DNA, especially in the study of obligate parasitic species for which ex vivo culture is resource-intensive or bias-prone. Modern sequence capture methods enable pathogen genetic variation to be analyzed directly from host/vector material but are often too complex and expensive for resource-poor settings where infectious diseases prevail. This study proposes a simple, cost-effective 'genome-wide locus sequence typing' (GLST) tool based on massive parallel amplification of information hotspots throughout the target pathogen genome. The multiplexed polymerase chain reaction amplifies hundreds of different, user-defined genetic targets in a single reaction tube, and subsequent agarose gel-based clean-up and barcoding completes library preparation at under 4 USD per sample. Our study generates a flexible GLST primer panel design workflow for Trypanosoma cruzi, the parasitic agent of Chagas disease. We successfully apply our 203-target GLST panel to direct, culture-free metagenomic extracts from triatomine vectors containing a minimum of 3.69 pg/μl T. cruzi DNA and further elaborate on method performance by sequencing GLST libraries from T. cruzi reference clones representing discrete typing units (DTUs) TcI, TcIII, TcIV, TcV and TcVI. The 780 SNP sites we identify in the sample set repeatably distinguish parasites infecting sympatric vectors and detect correlations between genetic and geographic distances at regional (< 150 km) as well as continental scales. The markers also clearly separate TcI, TcIII, TcIV and TcV + TcVI and appear to distinguish multiclonal infections within TcI. We discuss the advantages, limitations and prospects of our method across a spectrum of epidemiological research

The effect of collagen organization on tensile strength loss in anterior cruciate ligament grafts post-reconstruction surgery

Gemstone Team LEGSGrafts used for anterior cruciate ligament (ACL) reconstructions fall short of restoring native mechanics. This study investigated a morphological cause for tension loss by comparing native ACL and two common grafts, bone-patellar tendonbone (BPTB) and semitendinosus/gracilis hamstring tendon (ST/G), in a cadaveric system. Tension loss during continuous passive motion was quantified via force transducer. Microstructural changes were assessed by measuring collagen crimp angles. No significant differences were found for rates of percent tension loss relative to total tension loss among grafts. However, all groups displayed exponential decay, implying rapid tension loss. The crimp angles for the unstressed grafts were significantly different from each other, suggesting innate differences. The percent change experienced by stressed grafts, normalized to their unstressed baselines, showed that ST’s crimp behavior was significantly different from that of ACL and BPTB, implying the BPTB graft is superior for ACL reconstruction because it better mimics the ACL’s morphological behavior