Thermostable and highly specific l-aspartate oxidase from Thermococcus litoralis DSM 5473: cloning,overexpression, and enzymological properties

We successfully expressed the l-aspartate oxidase homolog gene (accession no: OCC_06611) of Thermococcus litoralis DSM 5473 in the soluble fraction of Escherichia coli BL21 (DE3) using a pET21b vector with 6X His tag at its C-terminus. The gene product (Tl-LASPO) showed l-aspartate oxidase activity in the presence of FAD in vitro, and this report is the first that details an l-aspartate oxidase derived from a Thermococcus species. The homologs of Tl-LASPO existed mainly in archaea, especially in the genus of Thermococcus, Pyrococcus, Sulfolobus, and Halobacteria. The quaternary structure of Tl-LASPO was homotrimeric with a subunit molecular mass of 52 kDa. The enzyme activity of Tl-LASPO increased with temperature up to 70 °C. Tl-LASPO was active from pH 6.0 to 9.0, and its highest activity was at pH 8.0. Tl-LASPO was stable at 80 °C for 1 h. The highest k _cat/K _m value was observed in assays at 70 °C. Tl-LASPO was highly specific for l-aspartic acid. Tl-LASPO utilized fumaric acid, 2,6-dichlorophenolindophenol, and ferricyanide in addition to FAD as a cofactor under anaerobic conditions. The absorption spectrum of holo-Tl-LASPO exhibited maxima at 380 and 450 nm. The FAD dissociation constant, K _d, of the FAD-Tl-LASPO complex was determined to be 5.9 × 10⁻⁹ M.

     

Elastin Fiber Accumulation in Liver Correlates with the Development of Hepatocellular Carcinoma

MethodsWe enrolled 189 consecutive patients with hepatitis C and advanced fibrosis. Using Elastica van Gieson-stained whole-slide images of pretreatment liver biopsies, collagen and elastin fibers were evaluated pixel by pixel (0.46 μm/pixel) using an automated computational method. Consequently, fiber amount and cumulative incidences of HCC within 3 years were analyzed.ResultsThere was a significant correlation between collagen and elastin fibers, whereas variation in elastin fiber was greater than in collagen fiber. Both collagen fiber (p = 0.008) and elastin fiber (p < 0.001) were significantly correlated with F stage. In total, 30 patients developed HCC during follow-up. Patients who have higher elastin fiber (p = 0.002) in addition to higher collagen fiber (p = 0.05) showed significantly higher incidences of HCC. With regard to elastin fiber, this difference remained significant in F3 patients. Furthermore, for patients with a higher collagen fiber amount, higher elastin was a significant predictor for HCC development (p = 0.02).ConclusionsComputational analysis is a novel technique for quantification of fibers with the added value of conventional staging. Elastin fiber is a predictor for the development of HCC independently of collagen fiber and F stage.     

1,4-Dioxane degradation potential of members of the genera <Emphasis Type="Italic">Pseudonocardia</Emphasis> and <Emphasis Type="Italic">Rhodococcus</Emphasis>

In recent years, several strains capable of degrading 1,4-dioxane have been isolated from the genera Pseudonocardia and Rhodococcus. This study was conducted to evaluate the 1,4-dioxane degradation potential of phylogenetically diverse strains in these genera. The abilities to degrade 1,4-dioxane as a sole carbon and energy source and co-metabolically with tetrahydrofuran (THF) were evaluated for 13 Pseudonocardia and 12 Rhodococcus species. Pseudonocardia dioxanivorans JCM 13855^T, which is a 1,4-dioxane degrading bacterium also known as P. dioxanivorans CB1190, and Rhodococcus aetherivorans JCM 14343^T could degrade 1,4-dioxane as the sole carbon and energy source. In addition to these two strains, ten Pseudonocardia strains could degrade THF, but no Rhodococcus strains could degrade THF. Of the ten Pseudonocardia strains, Pseudonocardia acacia JCM 16707^T and Pseudonocardia asaccharolytica JCM 10410^T degraded 1,4-dioxane co-metabolically with THF. These results indicated that 1,4-dioxane degradation potential, including degradation for growth and by co-metabolism with THF, is possessed by selected strains of Pseudonocardia and Rhodococcus, although THF degradation potential appeared to be widely distributed in Pseudonocardia. Analysis of soluble di-iron monooxygenase (SDIMO) α-subunit genes in THF and/or 1,4-dioxane degrading strains revealed that not only THF and 1,4-dioxane monooxygenases but also propane monooxygenase-like SDIMOs can be involved in 1,4-dioxane degradation.     

Pretreatment Gastric Lavage Reduces Postoperative Bleeding after Endoscopic Submucosal Dissection for Gastric Neoplasms

Aim

For patients receiving endoscopic submucosal dissection (ESD), there is urgent need pertaining to the prevention of postoperative bleeding. We conducted a retrospective propensity score-matched study that evaluated whether pre-ESD gastric lavage prevents postoperative bleeding after ESD for gastric neoplasms.

Methods

From September 2002 to October 2015, the 760 consecutive patients receiving ESD for gastric neoplasm were enrolled and data regarding them were retrospectively analyzed. All patients received conventional preventive treatment against delayed bleeding after ESD, including the administration of proton pump inhibitor and preventive coagulation of visible vessels, at the end of the ESD procedure.

Results

Pre-ESD risk factors for postoperative bleeding included tumor size and no gastric lavage. Using multivariate analysis tumor size >2.0 cm (HR 2.90, 95% CI 1.65–5.10, p = 0.0002) and no gastric lavage (HR 3.20, 95% CI 1.13–9.11, p = 0.029) were found to be independent risk factors. Next, we evaluated the effect of gastric lavage on the prevention of post-ESD bleeding using a propensity score-matching method. A total of 284 subjects (142 per group) were selected. Adjusted odds ratio of gastric lavage for post-ESD bleeding was 0.25 (95% CI 0.071–0.886, p = 0.032).

Conclusion

Pretreatment gastric lavage reduced postoperative bleeding in patients receiving ESD for gastric neoplasm.     

Diets and body condition of polar cod (<Emphasis Type="Italic">Boreogadus saida</Emphasis>) in the northern Bering Sea and Chukchi Sea

To understand trophic responses of polar cod Boreogadus saida (a key species in Arctic food webs) to changes in zooplankton and benthic invertebrate communities (prey), we compared its stomach contents and body condition between three regions with different environments: the northern Bering Sea (NB), southern Chukchi Sea (SC), and central Chukchi Sea (CC). Polar cod were sampled using a bottom trawl, and their potential prey species in the environment were sampled using a plankton net and a surface sediment sampler. Polar cod fed mainly on appendicularians in the NB and SC where copepods were the most abundant in the environment, while they fed on copepods, euphausiids, and gammarids in the CC where barnacle larvae were the most abundant species in plankton samples on average. The stomach fullness index of polar cod was higher in the NB and SC than CC, while their body condition index did not differ between these regions. The lower lipid content of appendicularians compared to other prey species is the most plausible explanation for this inconsistency.     

Ecological impacts of the 2011 Tohoku Earthquake Tsunami on aquatic animals in rice paddies

The 2011 Tohoku Earthquake Tsunami hit the Pacific coast of eastern Japan on March 11, 2011 and disturbed various ecosystems, including rice paddies along the coast. In Miyagi Prefecture, located in the Tohoku Region, a total of 12685 ha, or about 11.5 % of the rice paddies, were inundated and damaged by the tsunami. Although rice paddies are artificially constructed, they have long been used by many kinds of aquatic animals as habitats. To assess impacts of the 2011 Tohoku Earthquake Tsunami on aquatic animals inhabiting the rice paddies, census surveys were conducted at inundated and non-inundated rice paddies at six sites along the coast of Miyagi Prefecture from June to July 2012. This was 30–60 days after the first transplanting of rice seedling in inundated rice paddies following the tsunami. In the surveys a total of 100 aquatic animal taxa, 72 from inundated and 86 from non-inundated rice paddies, were found. Although the taxonomic richness and composition differed among the sites, the total number of observations of walk dispersers and obligatory aquatic animals without resting stages were significantly lower in inundated than in non-inundated rice paddies. However, no notable difference was detected in taxonomic richness and the total number of observations of flight dispersers and obligatory aquatic animals with resting stages between inundated and non-inundated rice paddies. The results showed that the tsunami had a substantial impact on freshwater animals but that restoration of the paddies for rice cultivation enhanced rapid recolonization by aquatic animals with high dispersal abilities and resting stages.     

What factors determine the number of nonmuscle myosin II in the sarcomeric unit of stress fibers?

Biomechanics and Modeling in Mechanobiology - Actin stress fibers (SFs), a contractile apparatus in nonmuscle cells, possess a contractile unit that is apparently similar to the sarcomere of...     

Efficient Endocytic Uptake and Maturation in Drosophila Oocytes Requires Dynamitin/p50

Dynactin is a multi-subunit complex that functions as a regulator of the Dynein motor. A central component of this complex is Dynamitin/p50 (Dmn). Dmn is required for endosome motility in mammalian cell lines. However, the extent to which Dmn participates in the sorting of cargo via the endosomal system is unknown. In this study, we examined the endocytic role of Dmn using the Drosophila melanogaster oocyte as a model. Yolk proteins are internalized into the oocyte via clathrin-mediated endocytosis, trafficked through the endocytic pathway, and stored in condensed yolk granules. Oocytes that were depleted of Dmn contained fewer yolk granules than controls. In addition, these oocytes accumulated numerous endocytic intermediate structures. Particularly prominent were enlarged endosomes that were relatively devoid of Yolk proteins. Ultrastructural and genetic analyses indicate that the endocytic intermediates are produced downstream of Rab5. Similar phenotypes were observed upon depleting Dynein heavy chain (Dhc) or Lis1. Dhc is the motor subunit of the Dynein complex and Lis1 is a regulator of Dynein activity. We therefore propose that Dmn performs its function in endocytosis via the Dynein motor. Consistent with a role for Dynein in endocytosis, the motor colocalized with the endocytic machinery at the oocyte cortex in an endocytosis-dependent manner. Our results suggest a model whereby endocytic activity recruits Dynein to the oocyte cortex. The motor along with its regulators, Dynactin and Lis1, functions to ensure efficient endocytic uptake and maturation.