A new family of sugar-inducible expression vectors for Escherichia coli.

A set of 11 expression vectors was constructed, each of them harbouring a cloning cassette under the control of the araB promoter. Some of these vectors enable expression of foreign proteins in the cytoplasm, while others include a synthetic sequence coding for a very efficient secretion signal sequence. Other features are an f1 origin of replication (in plus or minus orientation) and a promoter(up) mutation that enhances the already very high level of expression from these vectors. With such a versatile vector family, cloning, sequencing and site-directed mutagenesis can be performed on the same vector, and the level of expression can be defined according to the specific constraints of a given protein.     

Cardiovascular, respiratory, thermoregulatory, sedative, and analgesic effects of intravenous administration of medetomidine in rhesus macaques (Macaca mulatta).

BACKGROUND AND PURPOSE: Medetomidine is a selective, specific, and potent alpha2-adrenergic receptor agonist that has been utilized successfully as a sedative/analgesic agent in a variety of domestic and nondomestic animals. The objective of this study was to document the physiological effects of the intravenous administration of medetomidine in rhesus macaques (Macaca mulatta). METHODS: Fifteen healthy rhesus macaques (Macaca mulatta), 5 to 15 years old and weighing 5.5 to 11.8 kg, were given four dosages of medetomidine (50, 100, 150, and 200 microg/kg of body weight) intravenously, and cardiovascular, respiratory, thermoregulatory, sedative, and analgesic effects were determined. RESULTS: All four doses of medetomidine induced a similar and significant decrease in mean arterial pressure, as well as a transient but significant increase in respiratory rate followed by a longer-lasting significant decrease. Bradycardia, hypotension, and loss of thermoregulatory ability accompanied by a biphasic respiratory response and inconsistent sedation, analgesia, and muscular relaxation were observed. Heart rate decrease was rapid for all doses, but was significantly lower and of shorter duration after administration of the 50 microg/kg dosage. CONCLUSION: The inconsistency of the anesthetic plane induced by intravenous administration of medetomidine precludes it from being used alone to sedate rhesus macaques.     

Cross-talk between the calcium channel TRPV4 and reactive oxygen species interlocks adhesive and degradative functions of invadosomes

Invadosomes support cell invasion by coupling both acto-adhesive and extracellular matrix degradative functions, which are apparently antagonistic. β1-integrin dynamics regulate this coupling, but the actual sensing mechanism and effectors involved have not yet been elucidated. Using genetic and reverse genetic approaches combined with biochemical and imaging techniques, we now show that the calcium channel TRPV4 colocalizes with β1-integrins at the invadosome periphery and regulates its activation and the coupling of acto-adhesive and degradative functions. TRPV4-mediated regulation of podosome function depends on its ability to sense reactive oxygen species (ROS) in invadosomes’ microenvironment and involves activation of the ROS/calcium-sensitive kinase Ask1 and binding of the motor MYO1C. Furthermore, disease-associated TRPV4 gain-of-function mutations that modulate ECM degradation are also implicated in the ROS response, which provides new perspectives in our understanding of the pathophysiology of TRPV4 channelopathies.     

Brote de Salmonella enterica subsp. enterica serovar Give asociado con enfermedad transmitida por alimentos en Vichada,Colombia, 2015

IntroducciónSalmonella enterica subsp. enterica serovar Give se encuentra en mamíferos rumiantes, cerdos, aves y ambientes acuáticos, pero rara vez en humanos. En Colombia este serotipo ocupó el decimoprimer lugar en frecuencia en la vigilancia por laboratorio de la enfermedad diarreica aguda entre el 2000 y el 2013.ObjetivoCaracterizar el fenotipo y el genotipo de S. Give en aislamientos relacionados con un brote de enfermedad transmitida por alimentos en el departamento de Vichada en la quinta semana epidemiológica del 2015.Materiales y métodosSe buscó Salmonella spp. en 37 muestras de materia fecal con el método de estudio del Instituto Nacional de Salud. La muestra de sardinas enlatadas fue procesada según la norma ISO6579:2002 Cor.1:2004. Se determinó el serotipo en los aislamientos confirmados mediante serología o PCR en tiempo real, y se hicieron pruebas de sensibilidad a antimicrobianos y electroforesis en gel de campo pulsado con las enzimas Xbal y BlnI.ResultadosTodos los aislamientos de origen humano (11) y el aislamiento del alimento (1), se identificaron como S. Give y este último presentó resistencia a la tetraciclina. El análisis por PFGE-XbaI agrupó bajo el patrón COIN15JEXX01.0005 diez aislamientos de origen humano y a los restantes bajo el COIN15JEXX01.0006, con un 96,3% de similitud. Los resultados de todos los aislamientos se confirmaron con la enzima BlnI; cuatro de ellos (tres humanos y el del alimento) se agruparon bajo el patrón COIN15JEXA26.002, con un porcentaje de similitud del 95,65%.ConclusiónEl estudio confirmó que las sardinas enlatadas se relacionaron con la transmisión de S. Give en el brote, que es el tercero ocasionado por este serotipo en Colombia.Palabras clave: Salmonella, brotes de enfermedades, enfermedades transmitidas por los alimentos, vigilancia epidemiológica, Colombia     

Environmental and occupational biomonitoring using the Comet assay

Biomonitoring of human populations exposed to potential mutagens or carcinogens can provide an early detection system for the initiation of cell disregulation in the development of cancer. In recent years, the Comet assay, also known as a “single cell gel” (SCG) electrophoresis assay, has become an important tool for assessing DNA damage in exposed populations. This is the method of choice for population-based studies of environmental and occupational exposure to air pollutants, metals, pesticides, radiation, and other xenobiotics as we show in this review. To appreciate the role of the Comet assay in the field of biomonitoring, we review data from 122 studies that employed the assay. These studies evaluated environmental versus occupational exposures and the levels of DNA damage in cells of individuals exposed in each case. Our review of the literature reveals the importance of the need to establish standard methodological conditions that affect unwinding and electrophoresis times and tail values (tail length, tail DNA, tail moment), with the goal of being able to compare data collected in different laboratories throughout the world. The Comet assay is susceptible to subtle artifacts of manipulation depending on the type and timing of sampling performed. Therefore, in the reporting of DNA damage detected by the Comet assay, the context of how the DNA damage was created also needs to be reported and considered in the interpretation of Comet assay results. The success of the Comet assay is reflected by its use over the past 20 years in the field of biomonitoring, and by the increasing number of studies that continue to report its use. As the shortcomings of the assay are identified and considered in the interpretation of DNA damage detection, the Comet assay will continue to provide improved reliability as a biomarker in human biomonitoring studies.     

Binding of [3H]cytochalasin B to tumoral islet cells

Tumoral pancreatic islet cells of the RINm5F line are equipped with two classes of [3H]cytochalasin B binding sites with respective Kd of 0.4 and 7 microM. The binding of the fungal metabolite and its dissociation from the binding sites display rapid time courses. The binding is inhibited by D-glucose, more than by L-glucose, by phlorizin and by cytochalasin E. These findings are considered in the light of the dual action of cytochalasin B upon hexose transport and motile activity in islet cells.