Expression of aldehyde dehydrogenase gene increases hydrogen production from low concentration of acetate by Rhodobacter sphaeroides

Rhodobacter sphaeroides RV (RV) is a hydrogen-producing bacterium exhibiting the highest yield of hydrogen production from organic acids such as lactate and acetate, which are the byproducts of hydrogen fermentation by hydrogen-producing anaerobic bacteria. Co-fermentation of the RV strain with anaerobic bacteria is an efficient method of hydrogen production. However, less than 21 mM acetate is produced by the anaerobic bacteria, which is too low for efficient hydrogen production by the RV strain; it requires approximately 75 mM acetate. In this study, 2 distinct isozymes of aldehyde dehydrogenase from Rhodospirillum rubrum were separately overexpressed in the RV strain. The recombinant RV strains that were designated as RVAD1 and RVAD2, exhibited 13-fold higher ALDH activities than the wild-type RV strain. Hydrogen yields of both of the recombinant strains were 1.4-fold higher than that of the RV strain in 21 mM acetate. In 43 mM acetate, the RVAD1 strain showed higher yield, though the RVAD2 strain showed lower yield as compared to that of the RV strain. In 64 mM acetate and all concentrations of lactate tested (21, 43 and 64 mM), the yields of the recombinant strains were lower than those of the RV strain. The intact (empty) expression plasmid increased the ALDH activity and had little effect on the hydrogen production in acetate, however, it decreased the production in lactate. At the beginning of the fermentation process, when very little hydrogen had been produced, the recombinant strains expressing the ALDH gene consumed smaller amounts of acetate compared to the wild-type strain. We have discussed the effects of ALDH on hydrogen production in this report.     

Burnett Method with Absolute Pressure Transducer and Measurements for PVT Properties of Nitrogen and Hydrogen up to 473 K and 100?MPa

A measurement method for PVT properties of high-temperature and high-pressure gases was developed by simplifying the Burnett method and revising the data acquisition procedure. Instead of a differential pressure transducer, which is traditionally used, an absolute pressure transducer is used in the present method, and the measurement of pressure becomes easier. However, the absolute pressure transducer is placed outside the constant temperature bath because of the difficulty of its use in high-temperature surroundings, and some parts with different temperatures from the sample vessels exist as dead space. The present method takes into account the effect of the dead space in the data acquisition procedure. Nitrogen was measured in the temperature range from 353 K to 473 K and at pressures up to 100 MPa to determine the apparatus constants, and then, hydrogen was measured at 473 K and up to 100 MPa. The determined densities are in agreement within uncertainties of 0.07% to 0.24% (k = 2), both with the latest equation of state and existing measured data.     

Controlled transformation of carbon coating on silicon nanochains into nanotubes of carbon by Joule heating

A method of carbon coating of silicon nanochains is developed for the controlled transformation of silicon nanochains into nanotubes of carbon: the surface of silicon nanochains is coated with hexadecanoic acid by annealing after plasma cleaning. Individual silicon nanochains are heated by applying an electric current using a micro-manipulator for the transformation. The transformation is observed in situ by means of transmission electron microscopy.     

Effects of cyclic vertical loading on bearing and pullout capacities of piles with continuous helix wing

Laboratory and field tests were conducted to investigate the bearing and pullout capacities of steel piles with a continuous helix wing during cyclic loading. Both continuous helix and straight-sided piles were subjected to monotonic compressive, monotonic tensile, and cyclic reversal loading in the laboratory, while only the continuous helix pile was tested in the field. Both the laboratory and the field tests showed that the bearing and pullout capacities of the continuous helix pile under cyclic reversal loading decreased to approximately 60–80% of those of the pile under monotonic loading, with a larger reduction seen in the laboratory tests. The decrease in resistance was mainly due to the reduction in shaft friction, which was likely to be the result of soil disturbance and loosening around the pile with cyclic loading. The laboratory tests also showed that the tip resistance of the straight-sided pile under cyclic reversal loading was reduced, similarly due to the loosening of the soil, particularly underneath the pile tip. The tip resistance of the continuous helix pile, in contrast, did not degrade with cyclic loading, owing to the presence of the wing immediately above the pile tip that inhibited the loosening of the soil. These findings were supported by similar field test observations.     

Cold Model Study on Mg Desulfurization of Hot Metal Under Mechanical Stirring

The new method of in-situ desulfurization with mechanical stirring of new type impellers was introduced,in which the bubble′s dispersion and disintegration of magnesium vapor were the key to boosting the desulfurization efficiency and increasing the utilization rate of magnesium.Effects of different new type of impellers on bubble dispersion and disintegration were studied through bubble image analysis,gas-liquid mass transfer,and power consumption levels of different impeller structures.The results showed that the sloped swept-back blade impeller-2produces optimal bubble′s dispersion and disintegration,as well as higher volumetric mass transfer coefficient and CO2gas utilization while consuming the least power.Numerical simulation result with Fluent software also showed that the sloped swept-back blade impeller-2has higher turbulent kinetic energy and better velocity distribution than the other two impellers.     

Stabilization of flavylium dye by incorporation into the pore of protonated zeolites

Flavylium dye, a model compound of natural anthocyanin, has a defect that its color stability is very poor. We have complexed the flavylium dye with protonated zeolites and succeeded to enhance both the thermal and chemical stability of the dye. The aqueous dispersion of the flavylium/zeolite complex maintains its color even at 353 K or at pH 9, under which conditions the aqueous solution of the flavylium loses its color immediately. The stability is enhanced when the dye is incorporated into the pore of the zeolites, and a large stabilization effect is obtained by a tight adsorption with a narrow pore, as long as the dye is incorporated into the pore. Because of its enhanced color stability, the flavylium dye incorporated in the zeolite pore can be a candidate for an environmentally friendly coloring material.     

Cytotoxic activity to acute myeloid leukemia cells by Antp-TPR hybrid peptide targeting Hsp90

We previously reported that Antp-TPR hybrid peptide inhibited the interaction of Hsp90 with TPR2A and had selective cytotoxic activity discriminating between normal and cancer cells to induce cancer cell death. In this study, we investigated the cytotoxic activity of Antp-TPR peptide toward acute myeloid leukemia (AML) cells. It was demonstrated that Antp-TPR peptide induced AML cell death in cell lines such as U937, K562, THP-1, and HL-60 via activation of caspases 3 and 7, and disruption of mitochondrial membrane potential. Conversely, Antp-TPR peptide did not reduce the viability of normal cells including peripheral blood mononuclear cells (PBMCs), although both geldanamycin and 17-AAG, small-molecule inhibitors of Hsp90, mediated cytotoxicity to these normal cells at low concentrations. In addition, mutation analysis of TPR peptide demonstrated that the highly conserved amino acids Lys and Arg were critical to the cytotoxic activity. These results indicated that Antp-TPR hybrid peptide would provide potent and selective therapeutic options in the treatment of AML.     

Accumulative gene integration into a pre-determined site using Cre/loxP

Site-specific gene recombination systems, such as Cre/loxP, have been used for genetic modification of cells and organisms in both basic and applied research. We previously developed an accumulative gene integration system (AGIS), in which target gene cassettes could be repeatedly integrated into a pre-determined site on a plasmid or cellular genome by recombinase-mediated cassette exchange (RMCE), using Cre and mutated loxPs. In the present study, we designed a simplified AGIS. For gene integration into a target site, the previous system used two loxP sites in the acceptor DNA, whereas the new system uses a single loxP site. The gene integration reactions were repeated four times in vitro using Cre protein and specific plasmids. The expected integration reactions mediated by Cre occurred at the loxP sites, resulting in integration of four target genes. The system was also used for genomic integration of reporter genes using Chinese hamster ovary (CHO) cells. The reporter genes were efficiently introduced into the CHO genome in a Cre-dependent manner, and transgene expression was detected after the integration reaction. The expression levels of the reporter genes were enhanced, corresponding to the increase of transgene copy number. Recombinase-mediated AGIS provides a useful tool for the modification of cellular genomes.     

Construction of heterotypic cell sheets by magnetic force-based 3-D coculture of HepG2 and NIH3T3 cells

Heterotypic 3-D coculture is essential to mimic tissues and organs, because cell-cell interaction between various types of cells is believed to be important for the activation of cellular functions. In this study, magnetic force was applied to construct a 3-D coculture system of HepG2 and NIH3T3 cells as a model of hepatocytes and mesenchymal cells. Magnetite cationic liposomes (MCLs) were used to label target cells. NIH3T3 cells labeled with MCLs were seeded onto ultralow-attachment plates, whose surface is composed of a covalently bound hydrogel layer that is hydrophilic and neutrally charged. When a magnet was placed under the plate, cells accumulated on the bottom of the well. After a 24-h incubation period, the cells formed a multilayered cell sheet, which contained the major mesenchymal extracellular matrix (ECM) components (fibronectin and type I collagen), suggesting that the use of stromal NIH3T3 cells gave sufficient strength to cell sheets. Both NIH3T3 and HepG2 cells were labeled with MCLs, and cocultured by two methods: NIH3T3 cell sheets were constructed and HepG2 cells were subsequently seeded onto NIH3T3 cell sheets, and then allowed to form layered cell sheets by applying magnetic force; or NIH3T3 and HepG2 cells were mixed and then allowed to form mixed cell sheets by applying magnetic force. These heterotypic multilayered cell sheets were successfully constructed and an enhanced albumin secretion by HepG2 cells was observed. These results suggest that the new tissue engineering technique using magnetite nanoparticles and magnetic force, to which we refer to as magnetic force-based tissue engineering (Mag-TE), is a promising approach to construct multilayered cell sheets consisting of heterotypic cocultured cells.