Density assessment and reporting for Phlebotomus perniciosus and other sand fly species in periurban residential estates in Spain.

Green periurban residential areas in Mediterranean countries have flourished in the last decades and become foci for leishmaniasis. To remedy the absence of information on vector ecology in these environments, we examined phlebotomine sand fly distribution in 29 sites in Murcia City over a 3-year period, including the plots of 20 detached houses and nine non-urbanized sites nearby. We collected 5,066 specimens from five species using "sticky" interception and light attraction traps. The relative frequency of the main Leishmania infantum vector Phlebotomus perniciosus in these traps was 32% and 63%, respectively. Sand fly density was widely variable spatially and temporally and greatest in non-urbanized sites, particularly in caves and abandoned buildings close to domestic animal holdings. Phlebotomus perniciosus density in house plots was positively correlated with those in non-urbanized sites, greatest in larger properties with extensive vegetation and non-permanently lived, but not associated to dog presence or a history of canine leishmaniasis. Within house plots, sand fly density was highest in traps closest to walls. Furthermore, the study provides a guideline for insect density assessment and reporting and is envisioned as a building block towards the development of a pan-European database for robust investigation of environmental determinants of sand fly distribution.

Neutrophil CD64 expression, procalcitonin and presepsin are useful to differentiate infections from flares in SLE patients with SIRS.

Background/Objective Differentiating systemic lupus erythematosus (SLE) activity from infections in febrile patients is difficult because of similar initial clinical presentation. The aim of this study is to evaluate the usefulness of a number of biomarkers for differentiating infections from activity in SLE patients admitted with systemic inflammatory response (SIRS). Methods Patients with SLE and SIRS admitted to the emergency room were included in this study. Measurements of different markers including procalcitonin, neutrophil CD64 expression and presepsin, were performed. Infection was considered present when positive cultures and/or polymerase chain reaction were obtained. Sensitivity and specificity were calculated for all biomarkers. Results Twenty-seven patients were admitted, 23 women (82.5%), mean age 33.2 years. An infectious disease was confirmed in 12 cases. Markers for SLE activity including anti-DNA titers by IIF ( p = 0.041) and enzyme-linked immunosorbent assay ( p = 0.009) were used for differentiating SLE flares from infection. On the contrary, increased procalcitonin ( p = 0.047), neutrophil CD64 expression by flow cytometry ( p = 0.037) and presepsin ( p = 0.037) levels were observed in infected SLE patients. Conclusions High neutrophil CD64 expression, presepsin and procalcitonin levels are useful to differentiate infections from activity in SLE patients. In most cases, a positive bioscore that includes these three markers demonstrate the presence of an infectious disease.

Rheological properties of ethyl cellulose-monoglyceride-candelilla wax oleogel vis-a-vis edible shortenings.

The gelation process, elasticity, and mechanical recovery after shear were studied in mixed oleogels of ethylcellulose (EC), monoglycerides (MG), and candelilla wax (CW). EC oleogels produced without MG showed grainy texture due to incomplete dissolution of crystalline fractions of raw EC in the vegetable oil (150 °C). These fractions were eliminated by dissolving the raw EC/MG mixture in ethanol, evaporating the solvent, dispersing, and dissolving the solid residue in the vegetable oil (150 °C) prior gelation. The EC polymeric network, and MG, and CW crystals had a positive interaction on the elasticity of mixed oleogels. Mixed oleogels produced under static conditions showed a 100 % of elasticity recovery after shearing, a phenomenon associated with an EC interchain hydrogen bonding mediated by hydroxyl groups of MGs. This tentatively resulted from the formation of junction zones of the type EC-[MG]n-EC. The rheological behavior of these olegels was remarkably close to that of commercial shortenings.

Temperature effect in physicochemical and bioactive behavior of biogenic hydroxyapatite obtained from porcine bones.

Biogenic hydroxyapatite (BHAp) is a widely used material in the biomedical area due to its similarities with the bone tissue mineral phase. Several works have been spotlighted on the thermal behavior of bone. However, little research has focused on determining the influence of calcination temperature in the physicochemical and bioactive properties of BHAp. In this work, a study of the physicochemical properties' changes and bioactive response of BHAp produced from porcine femur bones using calcination temperatures between 900 to 1200 °C was conducted. The samples' structural, morphological, and compositional changes were determined using XRD, SEM, and FTIR techniques. XRD results identified three temperature ranges, in which there are structural changes in BHAp samples and the presence of additional phases. Moreover, FTIR results corroborated that B-type substitution is promoted by increasing the heat treatment temperature. Likewise, samples were immersed in a simulated biological fluid (SBF), following the methodology described by Kokubo and using ISO 23317:2014 standard, for 3 and 7 days. FTIR and SEM results determined that the highest reaction velocity was reached for samples above 1000 °C, due to intensity increasing of phosphate and carbonate bands and bone-like apatite morphologies, compared to other temperatures evaluated.

Salivary D-dimer in pigs: Validation of an automated assay and changes after acute stress.

D-dimer is a peptide found in serum and is derived from the degradation of blood clots. Even though it has been analysed in human saliva, D-dimer has not been previously evaluated in the saliva of any veterinary species, and its source and role remain unknown. The objectives of this research were firstly, to validate the use of an automated method for the measurement of D-dimer in porcine saliva, and secondly, to evaluate whether D-dimer concentration changes in pig saliva after an acute stress stimulus. For this purpose, a complete analytical validation of a commercially-available immunoturbidimetric assay was carried out. In addition, an experimental acute stress model was induced in 11 pigs based on a technique involving restraint by nose-snare immobilisation for 1 min. Saliva samples were subsequently collected at different times and D-dimer, salivary alpha-amylase (sAA) and cortisol were assessed in order to evaluate changes in its concentrations after the stress induction. The D-dimer automated assay showed adequate reproducibility and sensitivity, with coefficients of variation below 10% and a limit of quantification of 0.167 µg/mL fibrinogen equivalent units (FEU). It also showed a high accuracy, determined by linearity under dilution and recovery tests. In the stress model, a significant increase (P < 0.05) in salivary D-dimer 15 min after the stress stimulus and a positive correlation between D-dimer and sAA (r = 0.51; P < 0.001) were observed. These results indicate that D-dimer can be measured in porcine saliva with an automated method and suggest that its concentration can be influenced by stressful conditions.

Water-in-oil organogel based emulsions as a tool for increasing bioaccessibility and cell permeability of poorly water-soluble nutraceuticals.

The use of organogels in food and pharmaceutical sciences has several technical problems related with restricted diffusion of the drugs and lack of a proper gelator molecule. These features are important into the new product design. An alternative to improve technological properties in organogels is the use of emulsions. However, there is a lack of knowledge about the behavior on bioaccessibility and permeability of bioactives loaded into organogel-based emulsions. The objective of the present experimental work was to study the physical properties of organogel-based emulsions made with vegetable oil loaded with three different bioactives (betulin, curcumin and quercetin) and the influence on their bioaccessibility. Organogels were made of canola or coconut oils and myverol as gelator (10% w/w). Water-in-oil emulsions (at 5, 10 and 12.5 wt% of water content) were prepared by mixing the melted proper organogel and water (80 °C) under high shear conditions (20,000 rpm). Micrographs, rheological tests (amplitude, frequency, temperature sweeps and creep-compliance measurements), DSC and particle size analysis were performed to samples. In vitro digestion (oral, gastric and intestinal phase), lipolysis assays, bioaccessibility and permeability tests by cell culture of Caco-2 were made. Organogels of coconut oil have shown poor emulsification properties.

Structuration, elastic properties scaling, and mechanical reversibility of candelilla wax oleogels with and without emulsifiers.

The crystal network development, elastic properties scaling behavior, and mechanical reversibility of candelilla wax (CW) oleogels with and without emulsifiers were studied. Saturated monoglycerides (MG) and polyglycerol polyricinoleate (PGPR) were added at 1 or 2 times the critical micelle concentration. Although the micelles of both emulsifiers act as nucleation sites for the mixture of aliphatic acids and alcohols of CW, they did not affect the oleogel's thermodynamic stability. It was established that the crystal network of CW consists of at least two types of crystals, one rich in n-hentriacontane and other rich in aliphatic acids. Both crystals species contributed significantly to the oleogel elasticity. The elastic properties scaling behavior of CW oleogels fitted the fractal model within the weak-link regime. The setting temperature and added emulsifier modified the crystal network fractal dimension. During shearing, oleogels had massive breaking of junction zones, causing the loss of fractality in the crystal network, which in turn decreased the system's elasticity.

DNA binding and selective gene induction by different forms of the p53 protein.

P53 is a tumor suppressor gene that plays a crucial role in suppressing tumorigenesis by inducing either cell cycle arrest or apoptosis in cells with DNA damage. In more than 50% of tumors p53 is inactivated by gene mutations. However, there have also been reports of tumor cells in which p53 remains wild type and is present in elevated concentrations. Here we utilized a set of mutant cell lines, which, unlike the parental A1-5 cell line, which expresses a mouse tsp53 and becomes growth arrested at 32 degrees C, are capable of growth at this same incubation temperature. We found that the tsp53 in the two cell lines, ALTR-17 and ALTR-24, was identical to the parental A1-5s and concentrated in the nucleus at 32 degrees C. Examination of both lines revealed that p21 was induced at 32 degrees C, although to a lesser extent than in parental cells and that the p21 genes were not mutated. Interestingly, evaluation of the conformation of tsp53 using conformation-specific antibodies showed that the protein existed in different forms, which were found to bind DNA using chromatin immunoprecipitation assays and which we showed could induce expression of a p21 reporter construct. We conclude that the tsp53 may exist in various forms capable of binding DNA.

Disposition kinetics and pharmacokinetics--pharmacodynamic integration of difloxacin against Staphylococcus aureus isolates from rabbits.

The pharmacokinetics of difloxacin were studied following intravenous (IV), subcutaneous (SC) and oral administration of 5mg/kg to healthy white New Zealand rabbits (n = 6). Difloxacin concentrations were determined by HPLC assay with fluorescence detection. Minimal inhibitory concentrations (MICs) assay of difloxacin against different strains of S. aureus from different european countries was performed in order to compute the main pharmacodynamic surrogate markers. The plasma difloxacin clearance (Cl) for the IV route was (mean +/- SD) 0.41 +/- 0.05 L/h kg. The steady-state volume of distribution (V(ss)) was 1.95 +/- 0.17 L/kg. The terminal half-life [Formula: see text] was (mean+/-SD) 4.19+/-0.34 h, 7.53 +/- 1.32 h and 8.00 +/- 0.45 h after IV, IM and oral, respectively. From this data, it seems that a 5 mg/kg dose difloxacin would be effective by SC and oral routes in rabbits against bacterial isolates with MIC0.1 microg/mL.

The physical association of multiple molecular chaperone proteins with mutant p53 is altered by geldanamycin, an hsp90-binding agent.

Wild-type p53 is a short-lived protein which turns over very rapidly via selective proteolysis in the ubiquitin-proteasome pathway. Most p53 mutations, however, encode for protein products which display markedly increased intracellular levels and are associated with positive tumor-promoting activity. The mechanism by which mutation leads to impairment of ubiquitination and proteasome-mediated degradation is unknown, but it has been noted that many transforming p53 mutants are found in stable physical association with molecular chaperones of the hsp70 class. To explore a possible role for aberrant chaperone interactions in mediating the altered function of mutant p53 and its intracellular accumulation, we examined the chaperone proteins which physically associate with a temperature-sensitive murine p53 mutant. In lysate prepared from A1-5 cells grown under mutant temperature conditions, hsp70 coprecipitated with p53Val135 as previously reported by others, but in addition, other well-recognized elements of the cellular chaperone machinery, including hsp90, cyclophilin 40, and p23, were detected. Under temperature conditions favoring wild-type p53 conformation, the coprecipitation of chaperone proteins with p53 was lost in conjunction with the restoration of its transcriptional activating activity. Chaperone interactions similar to those demonstrated in A1-5 cells under mutant conditions were also detected in human breast cancer cells expressing two different hot-spot mutations. To examine the effect of directly disrupting chaperone interactions with mutant p53, we made use of geldanamycin (GA), a selective hsp90-binding agent which has been shown to alter the chaperone associations regulating the function of unliganded steroid receptors. GA treatment of cells altered heteroprotein complex formation with several different mutant p53 species. It increased p53 turnover and resulted in nuclear translocation of the protein in A1-5 cells. GA did not, however, appear to restore wild-type transcriptional activating activity to mutant p53 proteins in either A1-5 cells or human breast cancer cell lines.

[Analysis and modelling of safety culture in a Mexican hospital by Markov chains]. / Análisis y modelado de la cultura de seguridad de un hospital mexicano mediante cadenas de Markov.

INTRODUCTION: The objective of this study was to analyse and model the safety culture with Markov chains, as well as predicting and/or prioritizing over time the evolutionary behaviour of the safety culture of the health's staff in one Mexican hospital. METHOD: The Markov chain theory has been employed in the analysis, and the input data has been obtained from a previous study based on the Safety Attitude Questionnaire (CAS-MX-II), by considering the following 6 dimensions: safety climate, teamwork, job satisfaction, recognition of stress, perception of management, and work environment. RESULTS: The results highlighted the predictions and/or prioritisation of the approximate time for the possible integration into the evolutionary behaviour of the safety culture as regards the "slightly agree" (Likert scale) for: safety climate (in 12 years; 24.13%); teamwork (8 years; 34.61%); job satisfaction (11 years; 52.41%); recognition of the level of stress (8 years; 19.35%); and perception of the direction (22 years; 27.87%). The work environment dimension was unable to determine the behaviour of staff information, i.e. no information cultural roots were obtained. CONCLUSION: In general, it has been shown that there are weaknesses in the safety culture of the hospital, which is an opportunity to suggest changes to the mandatory policies in order to strengthen it.

Phase behavior, structure and rheology of candelilla wax/fully hydrogenated soybean oil mixtures with and without vegetable oil.

Vegetable oil organogelation is one of the most promising strategies to eliminate trans fatty acids in plastic fats. Organogels prepared with edible wax are stable at refrigerator and room temperature. Some functional properties (i.e., texture) of wax organogels can be improved by adding saturated triacylglycerols. Mixtures of fully hydrogenated soybean oil (FH) and candelilla wax (CW) were studied with and without the addition of high oleic safflower oil (HOSFO). Crystallization and melting behavior, X-ray diffraction, and crystalline microstructure of the mixtures were analyzed. The elastic modulus (G'), and the structural recovery after shear of the organogels were also assessed. Mixtures without HOSFO formed solid dispersions of CW and FH crystals, where up to ~10% CW crystals were incorporated into the FH crystal lattice. The vegetable oil solutions of FH/CW mixtures crystallized from the melt, developed mixed crystal networks composed of FH crystals in the ß polymorph and CW in an orthorhombic subcell packing. As the systems crystallized in the most stable polymorph, only minor microstructural changes were shown along 28days of storage at 25°C. CW and FH crystals showed a synergistic effect on the elasticity of organogels. This was attributed to the large number FH crystals nucleated on the surface of CW crystals. The structural recovery after shear was superior for mixed organogels composed of CW platelets and grainy FH crystals compared to that of CW organogels. A recovery of up to 65.7% the G' of gels formed under static conditions was observed upon shearing.

A cross-sectional study of the impact of regular use of insecticides in dogs on Canine Leishmaniosis seroprevalence in southeast Spain.

The relationship between Canine Leishmaniosis (CanL) seroprevalence and regular use of topical insecticides was investigated in 800 pet dogs with no visible signs of CanL in Murcia, southeast Spain in 2011. Dogs were clients to 17 veterinary practices and were analyzed for Leishmania infantum antibodies in blood plasma using two commercial ELISAs (Ingezim, Ingenasa®, Spain; Leishcan, Hipra®, Spain). Owners were interviewed to gather data on dog related variables. They included date of birth, home address and frequency, duration and timing of insecticide treatments used to prevent ectoparasite infestations. The dog's residence was georeferenced and environmental data potentially associated with the dog's risk of L. infantum infection was obtained. A mixed logistic regression model was then developed to analyze the relationship between the dog's serological status and insecticidal treatment adjusted for demographic and environmental variables. Overall, CanL seroprevalence (95% confidence limits) was 18% (16-21%) including 11% in dogs not using insecticide treatments (n=60) and 19% in those using them (n=740) (p>0.05). At least 16 different insecticide products were used and 73%, 26% and 1% of dogs received 1, 2 and 3 products a year. The most frequent commercial brands used and the only ones in the market claiming anti-sandfly activity, were Scalibor collars (deltametrin 40mg/g; MSD®), Advantix pipettes (permethrin 500mg/ml and imidacloprid 100mg/ml; Bayer®) and Exspot spot-on pipettes (permethrin 715mg/ml; MSD®). Seroprevalence was 9%, 16%, 20%, 22% and 25% for dogs with Scalibor collars plus Advantix pipettes, Scalibor collars plus ExSpot pipettes, Advantix pipettes alone, Scalibor collars alone and Exspot pipettes alone, respectively. The multivariable model confirmed a significant reduction in the risk of Leishmania spp. seropositivity in dogs using the Scalibor and Advantix combination compared to those using either product alone and provided evidence of greatly increased risk of CanL in rural areas situated at 300-500m altitude and average March-July temperatures of 18.6-19°C. The study highlights the difficulty in controlling CanL infection by means of insecticide use alone and that it could be improved by using the Scalibor and Advantix combination and identifying and targeting specific geographical areas.

Conformational phenotype of p53 is linked to nuclear translocation.

P53 is inactivated in tumors by mechanisms other than mutations in the p53 gene itself. To gain insight into the mechanisms by which this inactivation occurs, we chemically mutagenized A1-5 cells expressing high levels of temperature sensitive p53val135 (tsp53) and selected for clones that were capable of growth at the permissive temperature for p53 activation. We expanded 22 clones (ALTR cells for A1-5 Low Temperature Resistant) that could grow at the permissive temperature. Most exhibited cytoplasmic sequestration as the mechanism by which p53 was inactivated. We show here that this cytoplasmically sequestered tsp53 protein is maintained in a mutant conformation. Only in clones with nuclear localized p53 is it also expressed in the wild-type conformation suggesting that subcellular localization of tsp53 is important in determining the conformation of the protein. Consistent with this, we show that the changes in conformation of p53 in A1-5 and SK-N-SH cells induced by ionizing radiation also correlate with nuclear translocation of p53. We suggest that nuclear translocation of p53 can result in a change in the conformation from mutant to wild-type but that these may be two separable events. Oncogene (2000) 19, 4042 - 4049.

Selective binding of different p53 response elements by p53 containing complexes.

The p53 tumor suppressor protein binds two copies of a ten base pair motif that is degenerate in eight out of ten bases and conforms to the sequence, 5'PuPuPuC(A/T)(T/A)GPyPyPy-3'. As a consequence of this high degree of degeneracy, p53 response elements show a great deal of variation and it has been speculated that the variation aids in the selective activation of p53 responsive genes by specific stimuli. Here, we examined the DNA binding characteristics of several different p53 protein complexes present in nuclear extracts prepared from a cell line expressing the murine temperature sensitive p53 protein, p53val135. Interestingly, the complexes exhibited a distinct preference for binding to some p53 response elements and not others. A critical determinant of this specificity was the sequence at the center of the ten base pair motif and alteration of a single base within this region was sufficient to alter the set of complexes that associated with the oligonucleotide. In addition, thermal denaturation experiments demonstrated that some complexes could bind DNA even though the p53val135 protein had a mutant conformation. Our results are consistent with the hypothesis that p53 can distinguish between various response elements and suggest that this selectivity is manifested, in part, by the sequence of the motif and conformation of the p53 protein.

Regulation of DNA binding and transactivation in p53 by nuclear localization and phosphorylation.

Compelling evidence indicates that p53 acts as a transcription factor and that this activity is regulated by several factors including subcellular localization and phosphorylation status of the protein. To learn more about how these two processes determine whether p53 becomes activated, we studied the temperature sensitive murine p53, p53val135. At nonpermissive temperatures, p53val135 remains sequestered in the cytoplasm of cells which express it. Electrophoretic mobility shift assays demonstrated that, under these conditions, the protein lacked DNA binding activity. However, by shifting to the permissive temperature, p53val135 became concentrated in the nucleus, hyperphosphorylated, and had acquired the ability to bind DNA in a sequence specific manner. This was accompanied by the induction of two p53 regulated genes, mdm2 and p21waf1, which indicated that p53val135 had become an active transcription factor. Two dimensional gel electrophoresis and tryptic peptide mapping showed that entry into the nucleus resulted in the appearance of new phosphorylated isoforms and that the protein had become extensively phosphorylation at the N-terminus. Notably, phosphorylation at the N-terminus occurred only in the nucleus, whereas phosphorylation at the C-terminus could occur in both the cytoplasm and the nucleus. Based on these observations, we suggest that phosphorylation of p53's N-terminus is compartmentally restricted.

Bile acid activation of the gadd153 promoter and of p53-independent apoptosis: relevance to colon cancer.

Bile acids are strongly implicated in the etiology of colon cancer. Bile acids also induce apoptosis, and this action may be a key to understanding their role in colon cancer. However the mechanism of bile acid induction of apoptosis is not known. We present evidence of bile acid activation of the gadd153 promoter (a promoter activated by DNA damaging agents). We also show that bile acid induction of apoptosis is p53-independent. In addition, bile salts were found to induce blebbing preceding the actual morphological onset of apoptosis, which indicates early cytoskeletal alterations.

Urinary ferritin and cystatin C concentrations at different stages of kidney disease in leishmaniotic dogs.

Traditional analytes do not detect early renal disease; therefore there is a need to find new early markers of chronic kidney disease (CKD) in dogs to avoid the progression to irreversible renal damage. Our objective was to evaluate the presence of ferritin and cystatin C in urine of dogs with CKD and to relate their concentrations with the severity of the disease. Samples obtained from dogs naturally infected with Leishmania infantum were classified into four groups on the basis of the results of urinary protein/creatinine ratio and serum creatinine. This study shows that ferritin and cystatin C concentrations were increased in the urine of dogs with renal damage. Cystatin C value in urine only increased in severe stages of CKD with serum creatinine values >1.4 mg/dL, while the urinary ferritin concentration increased in dogs with proteinuria and serum creatinine <1.4 mg/dL, being, therefore, a renal biomarker earlier than creatinemia.

Serum paraoxonase 1 (PON1) activity in acute pancreatitis of dogs.

OBJECTIVES: Serum paraoxonase 1 is considered a marker of inflammation and oxidative damage. The aims of this study were to evaluate changes in serum paraoxonase 1 activity in dogs with acute pancreatitis, to correlate serum paraoxonase 1 activity and other analytes known to be altered in dogs with pancreatitis and to assess the relationship between serum paraoxonase 1 activity and disease severity in dogs with acute pancreatitis. MATERIALS AND METHODS: Retrospective analysis of dogs with acute pancreatitis and healthy dogs in which serum paraoxonase 1 activity was measured were compared. RESULTS: Median serum paraoxonase 1 activity was significantly lower in dogs with pancreatitis (n = 19) compared to healthy ones (n = 19). Serum paraoxonase 1 activity was negatively correlated with serum lipase and amylase activities, and C-reactive protein and haptoglobin concentrations and was positively correlated with total cholesterol and glucose concentration. Disease severity was negatively correlated with serum paraoxonase 1 activity and positively correlated with triglyceride and C-reactive protein concentration. CLINICAL SIGNIFICANCE: Serum paraoxonase 1 activity is lower in dogs with acute pancreatitis and together with triglyceride and C-reactive protein concentrations is a potential marker of disease severity.

Okadaic acid induces transcription of junB through a CCAAT box and NF-Y.

The shellfish toxin, okadaic acid (OA), is a potent tumor promoter that induces expression of the proto-oncogene junB in mouse keratinocyte 308 cells. Here we show, through deletion analysis of the junB promoter, that sequences near the TATA box conferred transcriptional induction by OA. Transient transfections of luciferase constructs bearing the junB promoter with single mutations in various cis elements demonstrated that a promoter containing a mutated CCAAT box could not be induced by OA. When this CCAAT box was inserted into a heterologous promoter construct, OA induction was dependent on an intact CCAAT box. Flanking cis elements located near the CCAAT box, although not required for OA inducibility, did play a role in the basal level of transcription. NF-Y was shown by EMSA to bind to the CCAAT box. OA induction from the junB CCAAT box was blocked by dominant negative NF-YA as well as the CCAAT box-dependent anticancer drug, ET-473. Expression of a lexA/NF-YA chimeric protein demonstrated that OA induction was dependent on the binding of NF-Y family members. These studies demonstrate that OA can mediate transcriptional activation of junB through the classical CCAAT box and that transcription factor NF-Y plays a functional role in the induction.