MicroRNA-200 family modulation in distinct breast cancer phenotypes.

The epithelial to mesenchymal transition (EMT) contributes to tumor invasion and metastasis in a variety of cancer types. In human breast cancer, gene expression studies have determined that basal-B/claudin-low and metaplastic cancers exhibit EMT-related characteristics, but the molecular mechanisms underlying this observation are unknown. As the family of miR-200 microRNAs has been shown to regulate EMT in normal tissues and cancer, here we evaluated whether the expression of the miR-200 family (miR-200f) and their epigenetic state correlate with EMT features in human breast carcinomas. We analyzed by qRT-PCR the expression of miR-200f members and various EMT-transcriptional inducers in a series of 70 breast cancers comprising an array of phenotypic subtypes: estrogen receptor positive (ER+), HER2 positive (HER2+), and triple negative (TN), including a subset of metaplastic breast carcinomas (MBCs) with sarcomatous (homologous or heterologous) differentiation. No MBCs with squamous differentiation were included. The DNA methylation status of miR-200f loci in tumor samples were inspected using Sequenom MassArray® MALDI-TOF platform. We also used two non-tumorigenic breast basal cell lines that spontaneously undergo EMT to study the modulation of miR-200f expression during EMT in vitro. We demonstrate that miR-200f is strongly decreased in MBCs compared with other cancer types. TN and HER2+ breast cancers also exhibited lower miR-200f expression than ER+ tumors. Significantly, the decreased miR-200f expression found in MBCs is accompanied by an increase in the expression levels of EMT-transcriptional inducers, and hypermethylation of the miR-200c-141 locus. Similar to tumor samples, we demonstrated that downregulation of miR-200f and hypermethylation of the miR-200c-141 locus, together with upregulation of EMT-transcriptional inducers also occur in an in vitro cellular model of spontaneous EMT. Thus, the expression and methylation status of miR-200f could be used as hypothetical biomarkers to assess the occurrence of EMT in breast cancer.

Non-invasive prenatal diagnosis of multiple endocrine neoplasia type 2A using COLD-PCR combined with HRM genotyping analysis from maternal serum.

The multiple endocrine neoplasia type 2A (MEN2A) is a monogenic disorder characterized by an autosomal dominant pattern of inheritance which is characterized by high risk of medullary thyroid carcinoma in all mutation carriers. Although this disorder is classified as a rare disease, the patients affected have a low life quality and a very expensive and continuous treatment. At present, MEN2A is diagnosed by gene sequencing after birth, thus trying to start an early treatment and by reduction of morbidity and mortality. We first evaluated the presence of MEN2A mutation (C634Y) in serum of 25 patients, previously diagnosed by sequencing in peripheral blood leucocytes, using HRM genotyping analysis. In a second step, we used a COLD-PCR approach followed by HRM genotyping analysis for non-invasive prenatal diagnosis of a pregnant woman carrying a fetus with a C634Y mutation. HRM analysis revealed differences in melting curve shapes that correlated with patients diagnosed for MEN2A by gene sequencing analysis with 100% accuracy. Moreover, the pregnant woman carrying the fetus with the C634Y mutation revealed a melting curve shape in agreement with the positive controls in the COLD-PCR study. The mutation was confirmed by sequencing of the COLD-PCR amplification product. In conclusion, we have established a HRM analysis in serum samples as a new primary diagnosis method suitable for the detection of C634Y mutations in MEN2A patients. Simultaneously, we have applied the increase of sensitivity of COLD-PCR assay approach combined with HRM analysis for the non-invasive prenatal diagnosis of C634Y fetal mutations using pregnant women serum.

Role of fibronectin in the adhesion of Acinetobacter baumannii to host cells.

Adhesion to host cells is an initial and important step in Acinetobacter baumannii pathogenesis. However, there is relatively little information on the mechanisms by which A. baumannii binds to and interacts with host cells. Adherence to extracellular matrix proteins, such as fibronectin, affords pathogens with a mechanism to invade epithelial cells. Here, we found that A. baumannii adheres more avidly to immobilized fibronectin than to control protein. Free fibronectin used as a competitor resulted in dose-dependent decreased binding of A. baumannii to fibronectin. Three outer membrane preparations (OMPs) were identified as fibronectin binding proteins (FBPs): OMPA, TonB-dependent copper receptor, and 34 kDa OMP. Moreover, we demonstrated that fibronectin inhibition and neutralization by specific antibody prevented significantly the adhesion of A. baumannii to human lung epithelial cells (A549 cells). Similarly, A. baumannii OMPA neutralization by specific antibody decreased significantly the adhesion of A. baumannii to A549 cells. These data indicate that FBPs are key adhesins that mediate binding of A. baumannii to human lung epithelial cells through interaction with fibronectin on the surface of these host cells.

The involvement of thaumatin-like proteins in plant food cross-reactivity: a multicenter study using a specific protein microarray

Cross-reactivity of plant foods is an important phenomenon in allergy, with geographical variations with respect to the number and prevalence of the allergens involved in this process, whose complexity requires detailed studies. We have addressed the role of thaumatin-like proteins (TLPs) in cross-reactivity between fruit and pollen allergies. A representative panel of 16 purified TLPs was printed onto an allergen microarray. The proteins selected belonged to the sources most frequently associated with peach allergy in representative regions of Spain. Sera from two groups of well characterized patients, one with allergy to Rosaceae fruit (FAG) and another against pollens but tolerant to food-plant allergens (PAG), were obtained from seven geographical areas with different environmental pollen profiles. Cross-reactivity between members of this family was demonstrated by inhibition assays. Only 6 out of 16 purified TLPs showed noticeable allergenic activity in the studied populations. Pru p 2.0201, the peach TLP (41%), chestnut TLP (24%) and plane pollen TLP (22%) proved to be allergens of probable relevance to fruit allergy, being mainly associated with pollen sensitization, and strongly linked to specific geographical areas such as Barcelona, Bilbao, the Canary Islands and Madrid. The patients exhibited >50% positive response to Pru p 2.0201 and to chestnut TLP in these specific areas. Therefore, their recognition patterns were associated with the geographical area, suggesting a role for pollen in the sensitization of these allergens. Finally, the co-sensitizations of patients considering pairs of TLP allergens were analyzed by using the co-sensitization graph associated with an allergen microarray immunoassay. Our data indicate that TLPs are significant allergens in plant food allergy and should be considered when diagnosing and treating pollen-food allergy.

Graph based study of allergen cross-reactivity of plant lipid transfer proteins (LTPs) using microarray in a multicenter study.

The study of cross-reactivity in allergy is key to both understanding. the allergic response of many patients and providing them with a rational treatment In the present study, protein microarrays and a co-sensitization graph approach were used in conjunction with an allergen microarray immunoassay. This enabled us to include a wide number of proteins and a large number of patients, and to study sensitization profiles among members of the LTP family. Fourteen LTPs from the most frequent plant food-induced allergies in the geographical area studied were printed into a microarray specifically designed for this research. 212 patients with fruit allergy and 117 food-tolerant pollen allergic subjects were recruited from seven regions of Spain with different pollen profiles, and their sera were tested with allergen microarray. This approach has proven itself to be a good tool to study cross-reactivity between members of LTP family, and could become a useful strategy to analyze other families of allergens.

Targeting aquaporin function: potent inhibition of aquaglyceroporin-3 by a gold-based compound.

Aquaporins (AQPs) are membrane channels that conduct water and small solutes such as glycerol and are involved in many physiological functions. Aquaporin-based modulator drugs are predicted to be of broad potential utility in the treatment of several diseases. Until today few AQP inhibitors have been described as suitable candidates for clinical development. Here we report on the potent inhibition of AQP3 channels by gold(III) complexes screened on human red blood cells (hRBC) and AQP3-transfected PC12 cells by a stopped-flow method. Among the various metal compounds tested, Auphen is the most active on AQP3 (IC(50) = 0.8±0.08 µM in hRBC). Interestingly, the compound poorly affects the water permeability of AQP1. The mechanism of gold inhibition is related to the ability of Au(III) to interact with sulphydryls groups of proteins such as the thiolates of cysteine residues. Additional DFT and modeling studies on possible gold compound/AQP adducts provide a tentative description of the system at a molecular level. The mapping of the periplasmic surface of an homology model of human AQP3 evidenced the thiol group of Cys40 as a likely candidate for binding to gold(III) complexes. Moreover, the investigation of non-covalent binding of Au complexes by docking approaches revealed their preferential binding to AQP3 with respect to AQP1. The high selectivity and low concentration dependent inhibitory effect of Auphen (in the nanomolar range) together with its high water solubility makes the compound a suitable drug lead for future in vivo studies. These results may present novel metal-based scaffolds for AQP drug development.

Del artículo de revisión a la recomendación profesional: La uniformación y material de protección individual para los equipos del ámbito prehospitalario de urgencias y emergencias

The Spanish Society of Nursing Emergency (SEEUE) has several lines documentaries that can be consulted on its web page and that have been spread through various publications: Statutes, scientific recommendations, professional recommendations, statements and allegations, grounds for nursing emergency, guarantees and rules, documents of interest and legislation. Set this that composed the regulatory environment, legal and recommendations which society poses to the collective nurse from the area of the emergency, as well as the rest of actors associated with urgent assistance (institutional and administratively) and through what has been the work of conceptualization and definition in our area the past few years. Part of this documentation offer possibilities for scientific endorsement and professional and invites to continue building knowledge and evidence. It is in this sense in which this work can and should be defined from a literature review approach and under the scheme of "review article". The working Group in Primary Care (PC) of the SEEUE, decided to build a Professional Recommendation (PR) in one of the areas of "uncertainty/variability" in the employment context and historical demand of nurses and other emergency care team on security issues: "The uniform and personal protective equipment for professional teams on prehospital emergency areas”.

Identification of reference genes for quantitative RT-PCR in ascending aortic aneurysms.

Hypertension and congenital aortic valve malformations are frequent causes of ascending aortic aneurysms. The molecular mechanisms of aneurysm formation under these circumstances are not well understood. Reference genes for gene activity studies in aortic tissue that are not influenced by aortic valve morphology and its hemodynamic consequences, aortic dilatation, hypertension, or antihypertensive medication are not available so far. This study determines genes in ascending aortic tissue that are independent of these parameters. Tissue specimens from dilated and undilated ascending aortas were obtained from 60 patients (age ≤70 years) with different morphologies of the aortic valve (tricuspid undilated n = 24, dilated n = 11; bicuspid undilated n = 6, dilated n = 15; unicuspid dilated n = 4). Of the studied individuals, 36 had hypertension, and 31 received ACE inhibitors or AT1 receptor antagonists. The specimens were obtained intraoperatively from the wall of the ascending aorta. We analyzed the expression levels of 32 candidate reference genes by quantitative RT-PCR (RT-qPCR). Differential expression levels were assessed by parametric statistics. The expression analysis of these 32 genes by RT-qPCR showed that EIF2B1, ELF1, and PPIA remained constant in their expression levels in the different specimen groups, thus being insensitive to aortic valve morphology, aortic dilatation, hypertension, and medication with ACE inhibitors or AT1 receptor antagonists. Unlike many other commonly used reference genes, the genes EIF2B1, ELF1, and PPIA are neither confounded by aortic comorbidities nor by antihypertensive medication and therefore are most suitable for gene expression analysis of ascending aortic tissue.

Mechanisms of hydrogen peroxide-induced vasoconstriction in the isolated perfused rat kidney.

The vasoconstrictor effect of hydrogen peroxide (H(2)O(2)) on isolated perfused rat kidney was investigated. H(2)O(2) induced vasoconstriction in the isolated rat kidney in a concentration-dependent manner. The vasoconstrictor effects of H(2)O(2) were completely inhibited by 1200 U/ml catalase. Endothelium-removal potentiated the renal response to H(2)O(2). The H(2)O(2) dose-response curve was not significantly modified by administration of the NO inhibitor L-NAME (10(-4) mol/l), whereas it was increased by the non-specific inhibitor of K+-channels, tetraethylammonium (3.10(-3) mol/l). Separately, removal of extracellular Ca(2+), administration of a mixture of calcium desensitizing agents (nitroprusside, papaverine, and diazoxide), and administration of a protein kinase C (PKC) inhibitor (chelerythrine, 10(-5) mol/l) each significantly attenuated the vasoconstrictor response to H(2)O(2), which was virtually suppressed when they were performed together. The pressor response to H(2)O(2) was not affected by: dimethyl sulfoxide (7.10(-5) mol/l) plus mannitol (3.10(-5) mol/l); intracellular Ca(2+) chelation using BAPTA (10(-5) mol/l); calcium store depletion after repeated doses of phenylephrine (10(-5) g/g kidney); or the presence of indomethacin (10(-5) mol/l), ODYA (2.10(-6) mol/l) or genistein (10(-5) mol/l). We conclude that the vasoconstrictor response to H(2)O(2) in the rat renal vasculature comprises the following components: 1) extracellular calcium influx, 2) activation of PKC, and 3) stimulation of pathways leading to sensitization of contractile elements to calcium. Moreover, a reduced pressor responsiveness to H(2)O(2) in female kidneys was observed.

Unusual manifestation of toscana virus infection, Spain.

This study was supported in part by project 05/305, Junta de Andalucía, Spain

Prescripción enfermera en el contexto de las urgencias y emergencia

The area of the urgencies and emergencies, assisted from all levels and care settings raises, if possible, patterns of work and ways that collaboration between professionals and teamwork make nurse prescription, often pharmacological, a legislative needs in response to increased scientific evidence and through internationally accepted performance algorithms. Enabling the nurse to act according to these concepts and beyond any "doubt and suspicion" of illegality. Taking a consensus necessary training and development and according to professional specialization and differentiation in this area and as arguments to remove any doubt still remains the subject without enclosing any sense and in many sectors.

Toma de decisiones en Enfermería de Urgencias, Emergencias y Catástrofes: Márgenes de Autonomía y Responsabilidad

Fear of achieving higher levels of nurse autonomy must be overcome for a new environment of relationships, change, communication and respect within the multidisciplinary team. The progressive achievement of new skills, the handling of evidence and the implementation of models and ways of working based on the nursing process, through encouraged training courses into the company and by the possibility of resource management in line and technology, foster as the ideal setting for Emergency Nursing, Emergency and Disaster acquires its own identity and development.

Alveolar overdistension as a cause of lung injury: differences among three animal species.

This study analyses characteristics of lung injuries produced by alveolar overdistension in three animal species. Mechanical ventilation at normal tidal volume (10 mL/Kg) and high tidal volume (50 mL/Kg) was applied for 30 min in each species. Data were gathered on wet/dry weight ratio, histological score, and area of alveolar collapse. Five out of six rabbits with high tidal volume developed tension pneumothorax, and the rabbit results were therefore not included in the histological analysis. Lungs from the pigs and rats showed minimal histological lesions. Pigs ventilated with high tidal volume had significantly greater oedema, higher neutrophil infiltration, and higher percentage area of alveolar collapse than rats ventilated with high tidal volume. We conclude that rabbits are not an appropriate species for in vivo studies of alveolar overdistension due to their fragility. Although some histological lesions are observed in pigs and rats, the lesions do not appear to be relevant.

Strategies for the successful implementation of viral laboratory automation.

It has been estimated that more than 70% of all medical activity is directly related to information providing analytical data. Substantial technological advances have taken place recently, which have allowed a previously unimagined number of analytical samples to be processed while offering high quality results. Concurrently, yet more new diagnostic determinations have been introduced - all of which has led to a significant increase in the prescription of analytical parameters. This increased workload has placed great pressure on the laboratory with respect to health costs. The present manager of the Clinical Laboratory (CL) has had to examine cost control as well as rationing - meaning that the CL's focus has not been strictly metrological, as if it were purely a system producing results, but instead has had to concentrate on its efficiency and efficacy. By applying re-engineering criteria, an emphasis has had to be placed on improved organisation and operating practice within the CL, focussing on the current criteria of the Integrated Management Areas where the technical and human resources are brought together. This re-engineering has been based on the concepts of consolidating and integrating the analytical platforms, while differentiating the production areas (CORE Laboratory) from the information areas. With these present concepts in mind, automation and virological treatment, along with serology in general, follow the same criteria as the rest of the operating methodology in the Clinical Laboratory.

Pro-adrenomedullin as prognostic biomarker in the sepsis.

Measurement of biomarkers is a potential approach to early assessment and prediction of mortality in septic patients. The purpose of this study was to ascertain the prognostic value of pro-adrenomedullin (pADM), measured in all patients admitted to the ICU of our hospital with a diagnosis of severe sepsis or septic shock during 1 year.