Proteolytic processing and activation of Clostridium perfringens epsilon toxin by caprine small intestinal contents.

Epsilon toxin (ETX), a pore-forming toxin produced by type B and D strains of Clostridium perfringens, mediates severe enterotoxemia in livestock and possibly plays a role in human disease. During enterotoxemia, the nearly inactive ETX prototoxin is produced in the intestines but then must be activated by proteolytic processing. The current study sought to examine ETX prototoxin processing and activation ex vivo using the intestinal contents of a goat, a natural host species for ETX-mediated disease. First, this study showed that the prototoxin has a KEIS N-terminal sequence with a molecular mass of 33,054 Da. When the activation of ETX prototoxin ex vivo by goat small intestinal contents was assessed by SDS-PAGE, the prototoxin was processed in a stepwise fashion into an ~27-kDa band or higher-molecular-mass material that could be toxin oligomers. Purified ETX corresponding to the ~27-kDa band was cytotoxic. When it was biochemically characterized by mass spectrometry, the copresence of three ETX species, each with different C-terminal residues, was identified in the purified ~27-kDa ETX preparation. Cytotoxicity of each of the three ETX species was then demonstrated using recombinant DNA approaches. Serine protease inhibitors blocked the initial proteotoxin processing, while carboxypeptidase inhibitors blocked further processing events. Taken together, this study provides important new insights indicating that, in the intestinal lumen, serine protease (including trypsin and possibly chymotrypsin) initiates the processing of the prototoxin but other proteases, including carboxypeptidases, then process the prototoxin into multiple active and stable species. Importance: Processing and activation by intestinal proteases is a prerequisite for ETX-induced toxicity. Previous studies had characterized the activation of ETX using only arbitrarily chosen amounts of purified trypsin and/or chymotrypsin. Therefore, the current study examined ETX activation ex vivo by natural host intestinal contents. These analyses demonstrated that (i) ETX processing in host intestinal contents occurs in an ordered, stepwise fashion, (ii) processing of prototoxin by host intestinal contents results in higher-molecular-mass material and 3 distinct ~27-kDa ETX species, and (iii) serine proteases, such as trypsin, chymotrypsin, and other proteases, including carboxypeptidases, play a role in the activation of ETX by intestinal contents. These studies provide new insights into the activation and processing of ETX and demonstrate that this process is more complicated than previously appreciated

Heathland creation on improved grassland using sulphur: is there a conflict between optimal application rates for plant and animal communities?

We examined the effectiveness of using elemental sulphur (a soil acidifier) as a method for creating heathland on improved pasture. We determined i) optimal rate of sulphur application to control mesotrophic grasses ii) if invertebrates avoid areas of sulphur application. Results indicated that optimal sulphur application attracted invertebrates

Software projects effort estimation using “use case points” method in the context of Project Management Body of Knowledge

As a contribution to consider software projects to be risk investments, it is important the full standardization of either the planning methodology (as a subset of project management methodology) and the effort estimation method. We think there is important and abundant evidence backing the convenience of the join use of Project Management Body of Knowledge and “Use case points” method in software project planning and control. Software development teams should share information, as it is important to compare the “ex post” analysis of programming/budgeting estimations against project execution data with “real world” software projects. This comparative activity would include teams into a permanent improvement process.IV Workshop de Ingeniería de Software y Base de DatosRed de Universidades con Carreras en Informática (RedUNCI

The Protective Effect of L-carnitine on Ionizing Radiation-induced Free Oxygen Radicals

Ionizing radiation is known to generate reactive oxygen species (ROS) that can be removed by antioxidants.  L-carnitine, a natural component of mammalian tissue, is a necessary factor in the utilization of long-chain  fatty acids to produce energy. Furthermore it has been shown that L-carnitine is an antioxidant which has  a scavenger effect on ROS and a stabilizing effect on damaged cell membranes. The aim of the study was  to evaluate the potential protective effect of L-carnitine on radiation-induced free radicals in hamsters. Lcarnitine  was given by gavage at a dose of 50 mg/kg for 15 consecutive days before irradiation with a single  dose of 8 Gy. 24 h after radiation exposure, the hamsters were sacrificed and samples were taken from  blood and tissues, and the biochemical and histopatological determinations were carried out. In the irradiated  group, there were significant increases in plasma and liver malondialdehyde (MDA) with marked  reduction in glutathione (GSH) levels in the liver, compared with controls. In red blood cells, superoxide  dismutase (SOD) and catalase activities were also reduced. All these effects were reversed by L-carnitine.  In conclusion, L-carnitine with its antioxidant and free radical scavenging properties could play a modulatory  role against the cellular damage produced by free radicals induced by ionizing radiation.

Eddy-Current Measurements of Corrosion-Related Thinning in Aluminuium Lap Splices

The aging commercial aviation fleet requires new methods for detecting and characterizing corrosion. In particular, the need exists for a rapid and reliable method of nondestructively detecting and characterizing corrosion in layered aircraft skins. Aircraft skin consists of thin aluminum sheet. There are many joints at which these sheets are overlapped and attached to the airframe by fasteners that extend through the sheets and into the substructure. The overlapping aluminum sheets are generally separated by a thin gap, which may be filled with a sealant. Because of the overlapping nature of such joints, the second sheet and the substructure are not directly accessible from the surface

Comparison of Protective Effects of Melatonin and Amifostine on Radiation-Induced Renal Oxidative Stress in Rats

ΔΕΝ ΔΙΑΤΙΘΕΤΑΙ ΠΕΡΙΗΨΗIn this study, we aimed to compare the protective effects of melatonin and amifostine on radiation-induced oxidative stress. Fifty female Wistar rats (3-4 months old, weighing 200±25 g) were divided into five groups (with ten rats each) and treated as follows: control (Cont), radiotherapy alone (RT), radiotherapy + amifostine (RT+AMI), radiotherapy + melatonin (RT+MEL), radiotherapy + amifostine + melatonin (RT+AMI+MEL). Rats were irradiated individually with a single dose of 8 Gy and amifostine (200 mg/kg) and melatonin (10 mg/kg) was administered to rats 30 minutes before irradiation. At the end of this follow-up period (72 hours) the rats were sacrificed. Spectrophotometric Analysis has been performed to kidney tissue samples. As a result of statistical comparison between groups after RT, total antioxidant capacity (TAC) decreased, total oxidant status (TOS) and oxidative stress index (OSI) increased, although the statistically significant change was only for OSI (p = 0.030). Addition of AMI or MEL to RT increased TAC and OSI significantly (p = 0.000), but there was no additive effect for TAC and OSI when both drugs were given together (p = 1.000, p = 0.172, respectively). In terms of TOS, statistically significant increasing was only for AMI (p = 0.000). There was protective effect when both drugs were given together against on Radiation-Induced Renal Oxidative Stress