Transient cell proliferation with polyethylenimine-cationized N-terminal domain of simian virus 40 large T-antigen

Polyethylenimine (PEI) cationization is a powerful strategy for protein transduction into cells. In this study, we attempted the artificial regulation of cell proliferation by protein transduction of the N-terminal domain (1-132 amino acids) of the simian virus 40 large T-antigen (SVLT-N), which inactivates retinoblastoma family proteins but not p53. To deliver SVLT-N into cells, we employed an indirect cationization method by forming a complex of biotynylated SVLT-N through disulfide bonds (biotin-SS-SVLT-N) and PEI-cationized avidin (PEI600-avidin). Using this complex, SVLT-N was transduced into the nucleus of confluent and quiescent Balb/c 3T3 cells and was found to be complexed with a cellular target protein, pRb. Furthermore, SVLT-N transduction induced cell proliferation in spite of confluent conditions. Because SVLT-N thus transduced into cells gradually degraded and was not detectable after a 4-d incubation, transiently transformed cells were obtained by this method. These results suggest that oncogene protein transduction technology has great potential for in vitro regulation of cell proliferation.     

Bending fatigue strength of case-carburized helical gears with large helix angles up to 40 degrees

Case-carburizing of helical gears with large helix angles may form too large hardened layers near the tooth width end on the acute angle side (ACUTE-END), and adversely affect the bending fatigue strength. We investigated the bending fatigue strength of casecarburized helical gears with large helix angles up to approximately 40° through a bending fatigue test, hardness test, and residual stress measurement. We found that the case-carburizing formed large hardened layers near ACUTE-END, reduced the compressive residual stress near ACUTE-END, and restricted the improvement of the bending fatigue strength in a meshing state where tooth root stress became large near ACUTE-END. Based on the obtained bending fatigue limits, we revealed that ISO 6336-3:2006 overestimated the rate of increase of the permissible circumferential loads for helix angles exceeding approximately 30°, and ISO/DIS 6336-3:2018 underestimated this rate for helix angles near 30°.

     

Structure-Based Design,Synthesis, and Biological Evaluation of Imidazo[4,5-b]Pyridin-2-one-Based p38 MAP Kinase Inhibitors: Part 2

We identified novel potent inhibitors of p38 mitogen-activated protein (MAP) kinase using a structure-based design strategy, beginning with lead compound, 3-(butan-2-yl)-6-(2,4-difluoroanilino)-1,3-dihydro-2H-imidazo[4,5-b]pyridin-2-one ( 1 ). To enhance the inhibitory activity of 1 against production of tumor necrosis factor-α (TNF-α) in human whole blood (hWB) cell assays, we designed and synthesized hybrid compounds in which the imidazo[4,5-b]pyridin-2-one core was successfully linked with the p-methylbenzamide fragment. Among the compounds evaluated, 3-(3-tert-butyl-2-oxo-2,3-dihydro-1H-imidazo[4,5-b]pyridin-6-yl)-4-methyl-N-(1-methyl-1H-pyrazol-3-yl)benzamide ( 25 ) exhibited potent p38 inhibition, superior suppression of TNF-α production in hWB cells, and also significant in vivo efficacy in a rat model of collagen-induced arthritis (CIA). In this paper, we report the discovery of potent, selective, and orally bioavailable imidazo[4,5-b]pyridin-2-one-based p38 MAP kinase inhibitors.     

Evaluation of a nitric acid-based partial-digestion method for selective determination of inorganic arsenic in rice

Arsenic (As) uptake in human occurs via the food chain mainly. The Joint FAO/WHO Expert Committee on Food Additives has established the provisional tolerable weekly intake level for As as an inorganic As (iAs) value, because iAs in food is much more toxic than organic As. In this study, we studied an acid based partial-digestion method for the complete extraction of arsenicals from rice. HPLC/ICP-MS was used to determine the concentration of iAs selectively. The conditions adopted to extract arsenicals from a 0.5 g of finely ground rice sample were addition of 2 mL of 0.15 mol/L nitric acid and heating at 80 degrees C for 2 hr. The LOD and LOQ for iAs were 0.0024 and 0.0079 mg/kg dry weight, respectively. Recovery studies showed good accuracy. When the method was applied to ten short-grain brown rice samples, the iAs concentrations were 0.108-0.227 mg/kg dry weight and the total As concentrations were 0.118-0.260 mg/kg dry weight. Although dimethylarsinic acid was also detected in most samples, the percentage of iAs content in total As content was 62.2-96.3%. Thus, iAs was the principal As species in the short-grain brown rice samples tested.     

Detection of Verotoxigenic Escherichia coli O157 and O26 in food by plating methods and LAMP method: a collaborative study

In order to establish a rapid and sensitive method for the detection of Verotoxigenic Escherichia coli O157 and O26, a collaborative study was conducted focusing on a comparison of the efficiency of loop-mediated amplification (LAMP) assay targeting the Verocytotoxin (also called Shiga toxin) gene, utilizing a direct plating method and a plating method with immunomagnetic separation (IMS-plating method) using various agar media. In combination with enrichment with the modified EC supplemented with novobiocin, E. coli O157 was detected in most samples of ground beef and alfalfa sprouts by LAMP assay, the direct plating method and the IMS-plating method. E. coli O26 was detected in approximately 100% of the food samples by LAMP assay. However, the IMS-plating and direct plating methods recovered 80 and 50% in ground beef samples, respectively. As a result, it was demonstrated the LAMP assay is superior to the IMS-plating method. Based on these results, it appears LAMP assay is effective as a screening assay to detect E. coli O157 and O26 from positive samples.     

Propagation and termination of an internal crack in continuously cast austenitic stainless steel analyzed by ∑ values and the Schmid factor

The formation mechanism of an internal crack was clarified from the viewpoint of the crystallography and thermal expansion. An inverse pole figure map obtained by EBSD pattern showed that the crack propagated along the grain boundaries having high ∑ values within the columnar zone. After the crack initiation, these positions were considered to undergo cracking followed by propagation toward the equiaxed side. Near the termination position, the grains ahead of crack propagation had a Schmid factor higher than 0.45 consuming elastic strain energy. Thermal expansion measurements showed that the grain with (0 0 1) orientation had the largest expansion while that with (0 1 1) the smallest. The grain boundaries neighboring the combination of (0 0 1) and (0 1 1) grains had the largest thermal stress. Therefore, thermal stress contributed to the initiation of cracking. It was thus proposed to enlarge the equiaxed zone to prevent cracking by discontinuing the crack propagation.     

A short guide to topological terms in the effective theories of condensed matter

This article is meant as a gentle introduction to the topological terms that often play a decisive role in effective theories describing topological quantum effects in condensed matter systems. We first take up several prominent examples, mainly from the area of quantum magnetism and superfluids/superconductors. We then briefly discuss how these ideas are now finding incarnations in the studies of symmetry-protected topological phases, which are in a sense a generalization of the concept of topological insulators to a wider range of materials, including magnets and cold atoms.     

Silver(I)-Ion-Mediated Cytosine-Containing Base Pairs: Metal Ion Specificity for Duplex Stabilization and Susceptibility toward DNA Polymerases

Spectroscopic characterization of Ag^I-ion-mediated C-Ag^I-A and C-Ag^I-T base pairs found in primer extension reactions catalyzed by DNA polymerases was conducted. UV melting experiments revealed that C-A and C-T mismatched base pairs in oligodeoxynucleotide duplexes are specifically stabilized by Ag^I ions in 1:1 stoichiometry in the same manner as a C-C mismatched base pair. Although the stability of the mismatched base pairs in the absence of Ag^I ions is in the order C-A≈C-T>C-C, the stabilizing effect of Ag^I ions follows the order C-C>C-A≈C-T. However, the comparative susceptibility of dNTPs to Ag^I-mediated enzymatic incorporation into the site opposite templating C is dATP>dTTP≫dCTP, as reported. The net charge, as well as the size and/or shape complementarity of the metal-mediated base pairs, or the stabilities of mismatched base pairs in the absence of metal ions, would be more important than the stability of the metallo-base pairs in the replicating reaction catalyzed by DNA polymerases.     

Recovery of residual nutrients from agri-food byproducts using a combination of solid-state fermentation and insect rearing

The present research develops an innovative nutrient recovery strategy for capturing of un-utilised nutrients from agri-food byproducts using a combination of solid state fermentation (SSF) and insect rearing. SSF of borage and flaxseed meals were performed using GRAS organisms to release indigenous nutrients and to produce additional nutrients. Proximate analysis of the SSF-meals showed increases in both lipid and protein contents. Black soldier fly larvae (BSFL) were then grown on these SSF-meals for 12 day. The meals fermented singularly with Lactobacillus plantarum or Aspergillus niger displayed up to 30% enhancement in BSFL biomass, and dual fermentation with L. plantarum and Aspergillus oryzae resulted in an additional 10% enhancement. These examinations showed SFL use over-90% of proteins and lipids in SSF-meals. The results indicate that fermentation of these low-value meals can boost the efficacy of larval growth and the recovery of nutrients from agricultural byproducts as larval biomass.     

In Vivo Simultaneous Analysis of Gene Expression by Dual-Color Luciferases in Caenorhabditis elegans

Both fluorescent and luminescent observation are widely used to examine real-time gene expression patterns in living organisms. Several fluuorescent and luminescent proteins with specific optical properties have been developed and applied for simultaneous, multi-color observation of more than two gene expression profiles. Compared to fluorescent proteins, however, the application of multi-color luminescent imaging in living organisms is still limited. In this study, we introduced two-color luciferases into the soil nematode C. elegans and performed simultaneous analysis of two gene expression profiles. Using a green-emitting luciferase Eluc (emerald luciferase) and red-emitting luciferase SLR (stable luciferase red), the expression patterns of two genes were simultaneously observed in single animals from embryonic to adult stages over its whole life span. In addition, dual gene activities were observed at the single embryo level, with the simultaneous observation of morphological changes. These are the first application of a two-color luciferase system into a whole animal and suggest that precise relationship of expression patterns of multiple genes of interest can be analyzed over the whole life of the animal, dependent on the changes in genetic and/or environmental conditions.