Search

Rhodococcus qingshengii N9T-4 can grow on media without added carbon sources. Here, we report the complete nucleotide sequence of the N9T-4 genome, consisting of a chromosome (6,439,972 bp), a linear plasmid (pN9T4-1 [565,206 bp]), and two circular plasmids (pN9T-4-2 [99,662 bp] and pN9T-4-3 [30,419 bp]).

Nicotinamide mononucleotide (NMN), an intermediate in nicotinamide adenine dinucleotide biosynthesis, is recently attracting much attention for its pharmacological and anti-aging efficacies. However, current commercial products containing NMN are very high-priced because efficient and facile methods for industrial NMN production are limited. In this study, aiming for its nutraceutical application, we attempted to screen lactic acid bacteria for intracellular and/or extracellular NMN production. Using a bioassay system with an auxotrophic yeast that requires nicotinamide riboside (NR; dephosphorylated NMN), three candidates were obtained from a library of 174 strains of facultative anaerobic lactic acid bacteria. All three candidates belonged to the genus Fructobacillus and produced NR in the culture media (0.8–1.5 mg/l). Lactic acid bacteria of the genus Fructobacillus are known to use d-fructose as an electron acceptor in anaerobic lactic acid fermentation; addition of d-fructose to the medium caused intracellular accumulation of NMN and NR, but no extracellular production of these compounds was observed. Draft genome sequencing for one of the candidates suggested that nicotinamide phosphoribosyltransferase, which exists commonly in mammals but is less reported in microorganisms, is a key enzyme for NMN and NR production in the fructophilic bacteria.

Two genes, aldA, and mnoA, encoding an NAD-dependent aliphatic dehydrogenase and N,N′-dimethyl-4-nitrosoaniline-dependent methanol dehydrogenase, respectively, are strongly expressed when Rhodococcus erythropolis N9T-4 is grown under oligotrophic conditions. In this study, we found a transcriptional regulator required for the transcription of both aldA and mnoA. The transcriptional regulator was also found to be essential for the oligotrophic growth of N9T-4.

Two genes

We can relatively easily isolate “super” oligotrophs from various environments, which can grow on minimal medium without additional carbon and energy sources. The super oligotrophs isolated to date have all belonged to the genera Rhodococcus and Streptomyces. This chapter primarily describes the genetics and biochemistry of one of the super oligotrophs, R. erythropolis N9T-4, which was isolated from crude oil. It is suggested that C2 metabolism starting from acetaldehyde is the key to oligotrophic carbon metabolism of N9T-4. Intriguingly, this bacterium is also oligotrophic for nitrogen and sulfur sources. The most unequivocal evidence for the oligotrophy of N9T-4 relates to the utilization of nitrogen, in which ammonium transporter is involved in the incorporation of atmospheric ammonia under oligotrophic conditions. A unique intracellular structure found in the oligotrophically grown super oligotroph is also introduced.

Rhodococcus erythropolis N9T-4, isolated from stored crude oil, shows extremely oligotrophic features and can grow on a basal medium without any additional carbon, nitrogen, sulfur, and energy sources, but requires CO2 for its oligotrophic growth. Transmission electron microscopic observation showed that a relatively large and spherical compartment was observed in a N9T-4 cell grown under oligotrophic conditions. In most cases, only one compartment was observed per cell, but in some cases, it was localized at each pole of the cell, suggesting that it divides at cell division. We termed this unique bacterial compartment an oligobody. The oligobody was not observed or very rarely observed in small sizes under nutrient rich conditions, whereas additional carbon sources did not affect oligobody formation. Energy dispersive X-ray spectroscopy analysis revealed remarkable peaks corresponding to phosphorus and potassium in the oligobody. The oligobodies in N9T-4 cells could be stained by Toluidine blue, suggesting that the oligobody is composed of inorganic polyphosphate and is a type of acidocalcisome. Two genes-encoding polyphosphate kinases, ppk1 and ppk2, were found in the N9T-4 genome: ppk1 disruption caused a negative effect on the formation of the oligobody. Although it was suggested that the oligobody plays an important role for the oligotrophic growth, both ppk-deleted mutants showed the same level of oligotrophic growth as the wild-type strain.

An extreme oligotroph, Rhodococcus erythropolis N9T-4, showed intracellular accumulation of trehalose and glycogen under oligotrophic conditions. No trehalose accumulation was observed in cells grown on the rich medium. Deletion of the polyphosphate kinase genes enhanced the trehalose accumulation and decreases the intracellular glycogen contents, suggesting an oligotrophic relationship between among the metabolic pathways of trehalose, glycogen, and inorganic polyphosphate biosyntheses.

Rhodococcus erythropolis N9T-4, which is an extremely oligotrophic bacterium, can survive in a completely inorganic medium with no additional carbon source. This bacterium utilizes atmospheric CO2, but does not require any additional energy source such as light and hydrogen gas, required by autotrophic microorganisms. However, its CO2 fixation and energy-acquisition systems in the oligotrophic growth remain unrevealed. We expected N9T-4 to have the transporter(s) that imports essential compound(s) for its oligotrophic growth. Three putative ATP-binding cassette (ABC) transporters were found to be highly upregulated under oligotrophic conditions. We constructed the gene-deletion mutants of a gene encoding the substrate-binding protein for each ABC transporter (∆sbp1, ∆sbp2, and ∆sbp3). Among these mutants, ∆sbp1 showed growth defects on oligotrophic medium without carbon source. We examined the growth of the mutants on the oligotrophic medium containing 1% trehalose as a sole carbon source. The results exhibited worse growth of ∆sbp3 than that of the control strain (∆ligD), whereas intracellular trehalose content of all mutants decreased compared with that of ∆ligD. It was reported that trehalose functions as the mycolate carrier to the arabinogalactan layer in the cell wall of Mycobacterium tuberculosis. Transmission electron microscopic analysis of ∆sbp1 cells showed that an outermost envelope of the ∆sbp1 cell diminished, which was expected to be mycolate layer. From these results, we suggest that the same trehalose-recycling system as that in a Mycobacterium cell functions in the oligotrophic growth of N9T-4, and the ABC transporter comprising Sbp1 as the substrate-binding protein is strongly involved in the oligotrophic growth of N9T-4.

Corrosion under insulation (CUI) is an aging degradation issue in long-term service vessels and pipes made of carbon steel and low-alloy steel. One of the problems in managing CUI is an equivocal evaluation technique of thinning detected by inspection. A replacement period with more accuracy can be evaluated more appropriately by performing fitness-for-service (FFS) assessment in this equipment. It is important to verify the validity of the FFS assessment using actually corroded pipes in order to promote the spread of FFS assessment in the domestic industry. In the present paper, pipes with complicated metal-loss due to CUI that were used in a chemical plant are burst in burst tests. An estimated burst pressure, which is calculated based on the assessment of metal-loss and through finite element analysis (FEA) based on FFS assessment, is compared with experimentally obtained burst pressures in order to validate the integrity evaluation. The burst pressure is then estimated by FEA, in which the ductile fracture under the multiaxial stress condition is considered. The burst pressure estimated by FFS approximately matched the burst pressure obtained based on experimental results. Fitness-for-service is sufficiently valid for investigating the remaining strength or burst pressure of corroded pipe. In addition, the burst pressure estimated by FEA that considered the ductile fracture under the multiaxial stress condition agreed with the experimental results and is valid so long as the remaining strength factor (RSF) is less than 0.6.

Rhodococcus erythropolis N9T-4 grows on an inorganic solid-state medium with no additional carbon and energy sources; however, it is unable to grow well in a liquid culture medium under the oligotrophic conditions. We examined submerged cultivations of N9T-4 using a polyurethane foam sponge to achieve approximately 10 times of the oligotrophic growth of the bacterium in the liquid culture medium.

We previously developed an industrial production process for novel water-soluble indigestible polysaccharides (resistant glucan mixture, RGM). During the process, an anhydrosugar—levoglucosan —is formed as a by-product and needs to be removed to manufacture a complete non-calorie product. Here, we attempted to isolate thermophilic bacteria that utilize levoglucosan as a sole carbon source, to establish a removing process for levoglucosan at higher temperature. Approximately 800 natural samples were used to isolate levoglucosan-utilizing microorganisms. Interestingly, levoglucosan-utilizing microorganisms—most of which were filamentous fungi or yeasts—could be isolated from almost all samples at 25°C. We isolated three thermophilic bacteria that grew well on levoglucosan medium at 60°C. Two of them and the other were identified as Bacillus smithii and Parageobacillus thermoglucosidasius, respectively, by 16S rDNA sequence analysis. Using B. smithii S-2701M, which showed best growth on levoglucosan, glucose and levoglucosan in 5% (wt/vol) RGM were completely diminished at 50°C for 144 h. These bacteria are known to have a biotechnological potential, given that they can ferment a range of carbon sources. This is the first report in the utilization of levoglucosan by these thermophiles, suggesting that our results expand their biotechnological potential for the unutilized carbon resources.

Rhodococcus erythropolis N9T-4 shows extremely oligotrophic growth requiring atmospheric CO2 and forms its colonies on an inorganic basal medium (BM) without any additional carbon source. Screening of a random mutation library constructed by a unique genome deletion method that we established indicated that the aceA, aceB, and pckG genes encoding isocitrate lyase, malate synthase, and phosphoenolpyruvate carboxykinase, respectively, were requisite for survival on BM plates. The aceA- and aceB deletion mutants and the pckG deletion mutant grew well on BM plates containing L-malate and D-glucose, respectively, suggesting that the glyoxylate (GO) shunt and gluconeogenesis are essential for the oligotrophic growth of N9T-4. Interestingly, most of the enzyme activities in the TCA cycle were observed in the cell-free extract of N9T-4, with perhaps the most important exception being α-ketoglutarate dehydrogenase (KGDH) activity. Instead of the KGDH activity, we detected a remarkable level of α-ketoglutarate decarboxylase (KGD) activity, which is the activity exhibited by the E1 component of the KGDH complex in Mycobacterium tuberculosis. The recombinant KGD of N9T-4 catalyzed the decarboxylation of α-ketoglutarate to form succinic semialdehyde (SSA) in a time-dependent manner. Since N9T-4 also showed a detectable SSA dehydrogenase activity, we concluded that N9T-4 possesses a variant TCA cycle, which uses SSA rather than succinyl-CoA. These results suggest that oligotrophic N9T-4 cells utilize the GO shunt to avoid the loss of carbons as CO2 and to conserve CoA units in the TCA cycle.

Rhodococcus erythropolis N9T-4 shows extremely oligotrophic growth and requires CO2 for its growth. In this report, nitrogen sources for the oligotrophic growth of N9T-4 were examined. As is true for most other bacteria, N9T-4 preferred ammonium salt to nitrate as the nitrogen source on an inorganic minimum medium without carbon sources. Interestingly, N9T-4 could also grow on the minimal medium solidified by agarose or silica gel without carbon and nitrogen sources, suggesting that this bacterium is also oligotrophic for nitrogen. We can rule out the possibility of diazotrophic growth of this bacterium, because nitrogenase activity was not detected in the cells and the putative gene encoding nitrogenase was not found in N9T-4 genome. DNA microarray analysis revealed that one of the ammonium transporter genes (amtB) was strongly upregulated 40-50 fold higher under oligotrophic conditions than under heterotrophic conditions. Disruption of amtB led to a growth defect under nitrogen-limiting conditions. Furthermore, additional ammonia vapor enhanced the growth of N9T-4 on the minimum medium without nitrogen sources in a closed culture system. These results suggest that N9T-4 utilizes the trace amount of atmospheric ammonia as the nitrogen source.

AC susceptibilities (real χ ′ and imaginary χ ″) of Sr 0.6 K 0.4 Fe 2 As 2 (122 type) polycrystalline with Ag addition are analysed by the grained Bean model. A variety of characteristics, double peak in χ ″ and shoulder transition in χ ′, appear in the model simulation. Comparing the measured χ ′ and χ ″ with the model allows more clear insight on the polycrystalline structure. Estimated critical current densities J cg and J cl of the grain and the link in the iron-based pnictides show that the addition of 20 wt.% Ag increases J cl 9 times larger. Improvement of intergrain characteristics with Ag addition is clearly indicated.

The proline analogue cis-4-hydroxy-L-proline (CHOP), which inhibits the biosynthesis of collagen, has been clinically evaluated as an anticancer drug, but its water solubility and low molecular weight limits its therapeutic potential since it is rapidly excreted. In addition, CHOP is too toxic to be practical as an anticancer drug, due primarily to its systematic effects on noncollagen proteins. To promote CHOP's retention in blood and/or to decrease its toxicity, N-acetylation of CHOP might be a novel approach as a prodrug. The present study was designed to achieve the microbial production of N-acetyl CHOP from L-proline by coexpression of L-proline cis-4-hydroxylases converting L-proline into CHOP (SmP4H) from the Rhizobium Sinorhizobium meliloti and N-acetyltransferase converting CHOP into N-acetyl CHOP (Mpr1) from the yeast Saccharomyces cerevisiae. We constructed a coexpression plasmid harboring both the SmP4H and Mpr1 genes and introduced it into Escherichia coli BL21(DE3) or its L-proline oxidase gene-disrupted (ΔputA) strain. M9 medium containing L-proline produced more N-acetyl CHOP than LB medium containing L-proline. E. coli ΔputA cells accumulated L-proline (by approximately 2-fold) compared to that in wild-type cells, but there was no significant difference in CHOP production between wild-type and ΔputA cells. The addition of NaCl and L-ascorbate resulted in a 2-fold increase in N-acetyl CHOP production in the L-proline-containing M9 medium. The highest yield of N-acetyl CHOP was achieved at 42 h cultivation in the optimized medium. Five unknown compounds were detected in the total protein reaction, probably due to the degradation of N-acetyl CHOP. Our results suggest that weakening of the degradation or deacetylation pathway improves the productivity of N-acetyl CHOP.

平成23年（2011年）3月11日に発生した東北地方太平洋沖地震による宮城県北部地域の地盤被害状況を把握するため，地盤工学会東北支部・関西支部合同調査団を組織し，2011年4月5日～10日，29～30日にかけて調査を実施した。本論文では，その後の聞き取り調査なども踏まえ，河川堤防，海岸堤防，港湾施設，道路・鉄道の盛土・切土斜面，液状化や地盤沈降などの被害に関する調査結果を報告する。調査結果の一部は既に地盤工学会誌に速報として報告されているが，本論文ではそこで報告された被害事例も含めて被害対象ごとに整理して述べる。

A method to control nanostructure of metallic glass film (MGF) fabricated by electroless alloy plating (ELAP) is proposed. The nanostructure is controlled by density of catalyst core on a substrate, and which is a function of immersion time in catalyst solution. MGF patterning was performed successfully, and the characteristic nanostructure was observed by TEM. For ELAP process, density of catalyst core was decreased with increasing immersion time, while size of the core was enlarged. Nanostructures were refined with increase in catalyst core density. These results show that nanostructure of MGF pattern can be controlled by Pd catalyst density in the ELAP process.

Rhodococcus erythropolis N9T-4 shows extremely oligotrophic growth requiring atmospheric CO₂ without any additional carbon or energy source. We performed a gene expression analysis of the oxidoreductases, which are involved in methanol metabolism, under various growth and induction conditions in N9T-4. A real-time PCR analysis revealed that the genes encoding NAD-dependent formaldehyde dehydrogenase (nFADH) and N,N'-dimethyl-4-nitrosoaniline-dependent methanol dehydrogenase (MDH) were strongly expressed under the oligotrophic conditions at levels of 20-100 fold those under heterotrophic conditions, in which n-tetradecane was used as the sole carbon source, while glucose did not affect the gene expression pattern in a minimum medium. The genes encoding mycothiol-dependent formaldehyde dehydrogenase (mFADH) and formate dehydrogenase were not induced under oligotrophic conditions, although mFADH expression was observed when formaldehyde was added to the induction medium. These results suggest that N9T-4 had three distinct formaldehyde oxidation systems, and that MDH and nFADH were the key enzymes in its oligotrophic growth.

In this paper, the dynamic response of asphalt pave ment containing a hydraulic, graded iron and steel slag (hereafter called HMS) base-course under repeated plate-loading was investigated using a model asphalt pavement and the influence of hydraulicity on th e pavement behavior was discussed. The model pavement constructed was a 4-layer system consis ting of a dense-graded asphalt mix surface layer, a dense-graded asphalt mix binder-course, a HMS base-course and a Masado (heavily-weathered granitic sand) subgrade. A repeated plate-loading test was carri ed out so as to achieve a resilient state. It is shown that surface resilient deflection decreases as curing progresses and after 90 days, the deflection becomes almost half of the initial. Large horizontal tensile strains develop at the bottoms of binder- and base-course, which decrease significantly with curing. It is indicative that HMS base-course behaves like a stiffer plate resulting in a hard-to-deflect state due to the development of hydraulicity.

This paper discusses resilient Poisson's ratio of hydraulic, graded iron and steel slag base-course material based on a series of repeated loading triaxial compression tests on the specimens compacted with a maximum dry density and an optimum water content a nd cured for 0, 28, 91 and 365 days. It is shown that resilient Poisson's ratio for no cured specimens is stress dependent in such a way that it increases with deviatoric stress and decreases with mean effective principal stress. As curing time increases, however, the magnitude of resilient Poisson's ratio becomes smaller and its stress dependency vanishes: at curing time of one year, resilient Poisson's ratio appears independent on both mean effective principal and deviatoric stresses and can be taken as constant. Resilient Poisson's ratio can be well expressed by a power function of these two stresses. It is indicative that stress-dependency in resilient Poisson's ratio fades away after hydraulicity has sufficiently developed, which may justify the use of a constant value for resilient Poisson's ratio in pavement response analyses.

Relationship between fracture toughness and shear band length (SBL), crack tip opening displacement (CTOD) on the specimen surface and stretched zone width (SZW) on the fracture surface in Zr-, Cu- and Ti-based bulk metallic glasses (BMGs) was investigated in order to examine the mechanisms that the BMGs show a high fracture toughness. Zr50Cu40Al10 and Zr50Cu30Ni10Al10 BMGs fabricated by an ark tilt casting method, and Cu60Zr30Ti10, Ti41.5Zr2.5Hf5Cu42.5Ni7.5Si1 BMGs fabricated by a high-frequency induction casting method were used. The fracture toughness test was conducted based on ASTM E399. Compact type specimen was used (1.0 mm, 2.0 mm and 2.3 mm in thickness, thickness:width=1:4). Fatigue pre-crack was induced under ΔK less than 6 MPa•m1/2. The crack length and SZW were measured on the fractographs by scanning electron microscope (SEM), and the SBL and CTOD were measured on the photographs of specimen surfaces by the SEM. Observed SBL and CTOD in each BMG showed the near values of the plastic zone size and CTOD calculated by the fracture mechanics under the plane stress condition, respectively. By the stereo-matching observation on the fracture surfaces, the crack tip inside the specimen clearly opened and a stretched zone area was shown. Average SZW in front of the fatigue crack in the Zr- and Cu-based BMGs was roughly proportional to the fracture toughness value.

We found 11 genes (FAO1-11) encoding putative oxidoreductases in the Aspergillus oryzae genome, which are similar to fungal fructosyl-amino acid oxidases. The cDNAs corresponding to the genes were cloned and expressed in Escherichia coli. rFao2 had fructosyl-amino acid oxidase activity, whereas rFao1 did not show any enzyme activity, even though the deduced amino acid sequence of Fao1 is identical to that of one of the fructosyl-amino acid oxidase isozymes from Aspergillus oryzae. rFao7 and rFao8 showed oxidase activity toward sarcosine, L-pipecolate, and L-proline. rFao10 was active toward only sarcosine, of the substrates tested. The functions of the other proteins were also predicted from a phylogenetic analysis.

Bacterial communities in crude-oil samples from Japanese oil stockpiles were investigated by 16S rRNA gene cloning, followed by denaturing gradient gel electrophoresis (DGGE) analysis. 16S rRNA genes were successfully amplified by PCR after isooctane treatment from three kinds of crude-oil sample collected at four oil stockpiles in Japan. DGGE profiles showed that bacteria related to Ochrobactrum anthropi, Burkholderia cepacia, Stenotrophomonas maltophilia, Propionibacterium acnes, and Brevundimonas diminuta were frequently detected in most crude-oil samples. The bacterial communities differed in the sampling time and layer. Among the predominant bacteria detected in the crude oil, only three species were found for bacteria isolated on agar plates and were related to Burkholderia, Stenotrophomonas, and Propionibacterium, while Ochrobactrum sp. could not be isolated although this species seemed to be the most abundant bacterium in crude oil from the DGGE profiles. Using an archaea-specific primer set, methanogens were found in crude-oil sludge but not in crude-oil samples, indicating that methanogens might be involved in sludge formation in oil stockpiles.

Estrogen receptor (ER)-positive breast cancer is a distinct subpopulation of breast cancer exhibiting a high response rate to endocrine therapy. However, not all ER-positive patients respond to the therapy, and a subgrouping of ER-positive patients based on the physiology of estrogen signaling is expected to be useful for predicting the prognosis. This study has revealed that selected estrogen-regulated genes (ERGs) are useful in identification of a poor-prognosis population among ER-positive breast cancer patients. First, the expression levels of 11 ERGs, selected based on our earlier microarray study in cultured cells, were analyzed by means of real-time reverse transcription-PCR in 14 ER-positive human breast cancer tissues. The patients were clearly divided into two groups in cluster analysis. Then, we examined the expression levels of two representative ERGs, histone deacetylase 6 (HDAC6) and insulin-like growth factor binding protein 4 (IGFBP-4), in 62 ER-positive patients with immunohistochemistry to assess the impact of ERG expression on prognosis (median follow-up 4409 days). Positive HDAC6 staining was significantly correlated with a lower disease-free survival rate. Moreover, when the expression level of HDAC6 was assessed in combination with IGFBP-4 expression in the nucleus, the poor-prognosis patients were more accurately identified. This study has identified new candidate ERGs for prediction of prognosis, and we suggest that combined assessment of the expression levels of these ERGs will contribute to the clinically useful stratification of ER-positive breast cancer patients.

In this paper, a staged compression-immersion-direct shear test was conducted on compacted samples of crushed mudstone aggregates and its compressive and shear behavior are discussed with a focus on cementation effects. Compression behavior of the compacted samples was influenced significantly by the compaction degree as expected, as were shear behavior and shear strength. Immersion caused an additional compression and a reduction in mobilized shear stress and in the dilatant nature during shear at low applied pressure levels. Moreover, immersion significantly reduced the peak shear strength parameter c with only a little change in phi. The compression and critical state lines of the nonimmersed and immersed specimens seem to parallel each other, and the compression line of the nonimmersed and the critical state line of the immersed form the upper and lower bounds, respectively. A gap between shear stress-void ratio lines of the specimens with and without immersion is considered to represent a combined effect of cementation retained in a crushed mudstone aggregate itself and an interlocking effect of aggregates.

Prostaglandin H synthase (PHS) is a key enzyme in the synthesis of prostaglandins (PGs). Recently, enhanced expression of PHS-2 in brain tumors and the correlation between the PHS-2 level and the histopathological grade of glioma has been reported. Furthermore, in vitro inhibition of glioma cell growth by a specific PHS-2 inhibitor, NS398, has been demonstrated. It has also been shown that prostaglandin E2 (PGE2) contributes to colon carcinogenesis by binding to the prostaglandin E receptor subtype EP1. We therefore evaluated the effects of NS398 and two EP1 antagonists, SC51089 and AH6809, on glioma cell lines. To evaluate mechanisms of NS398's action, two glioma cell lines, a PHS-2-positive cell line (KMG4) and a PHS-2-deficient cell line (A 172), were used. NS398 inhibited both the anchorage-dependent and -independent growth of glioma cell lines regardless of PHS-2 expression, suggesting that some PHS-2-independent mechanisms underlie the antineoplastic effect of NS398. However, the antineoplastic effect was attenuated by the addition of PGE2, which is one of the main products of PHS, suggesting the predominant mechanism is PHS-dependent. The EP1 antagonists, SC51089 and AH6809, inhibited the growth of glioma cell lines in vitro. Furthermore, NS398 or SC51089 slowed tumor growth in vivo, which was assessed using KMG4 tumor xenografts on SCID mice. PHS-2 inhibitors and EP1 antagonists might be useful in the prevention and/or treatment of glioma.

水硬性粒度調整スラグ路盤材のレジリエントモジュラスを評価するために, 軸変位及び軸荷重の計測精度を向上させて, AASHTO 試験法で推奨されている繰返し載荷方法に準じて繰返し三軸圧縮試験を実施した. さらに, 西らが採用している載荷方法でも試験を行い, 既往のレジリエントモジュラスとの比較および載荷方法の違いがレジリエントモジュラスに及ぼす影響について考察した. その結果, 水硬性粒度調整スラグ路盤材のレジリエントモジュラスは, 過去の結果と同様に, 平均主応力と偏差応力に依存し養生期間が長いほど大きい値であることを確認した. さらに, 従来のようなセル外部のLVDTによる軸変位計測から算定した値はかなりの過小評価になっており, 試験法の違いの影響は非常に小さいことがわかった.

Summary. Although primary haemophagocytic lymphohistiocytosis (HLH) is a genetic disorder of T lymphocytes, it remains unclear why T lymphocytes of primary HLH patients preferentially infiltrate the central nervous system and peripheral blood, in addition to the reticuloendothelial systems. We engrafted Herpesvirus saimiri (HVS)-immortalized T-lymphocyte lines established from primary HLH patients into severe combined immunodeficient (scid) mice and examined their capacity to infiltrate mouse organs. A diffuse infiltration of human T lymphocytes was detected in each organ of scid mice treated with 1 × 106 T lymphocytes from all four primary HLH patients assessed, whereas no infiltration of T lymphocytes from healthy individuals was observed in any organ. The infiltration of T lymphocytes was mainly observed in the lung, brain and peripheral blood, in association with haemophagocytosis. These cells were positive for HLA-DR, CD3 and either CD8 or CD4, but negative for CD68. Certain markers of proliferation and apoptotic activities were highly positive in these cells. There was no difference between the infiltration pattern of T lymphocytes of primary HLH patients with a perforin deficiency and those without. By Southern blot analysis, T lymphocytes infiltrating mouse organs were observed to be polyclonal. These findings suggest that our murine model implementing HVS-immortalized human T lymphocytes is suitable to clarify the pathogenesis of primary HLH.

CO utilization by a CO(2)-requiring extremely oligotrophic bacterium, Rhodococcus erythropolis N9T-4 was found when CO(2) was removed from the culture environment before cultivation, suggesting that this bacterium can convert CO into CO(2). However, the gene encoding putative CO dehydrogenase large subunit in N9T-4 was not induced by CO.

Especially in the analysis of environmental samples, it is necessary to perform rapid, simple and multiple analyses because it highly desirable to determine the results quickly as well as to analyze numerous samples simultaneously. We use 2,6-dichlorophenolindophenol (DCIP) as a redox color indicator for the Biochemical Oxygen Demand (BOD) determination in this report. The absorbance of DCIP decreases due to the metabolism of organic substances in aqueous samples by Pseudomonas fluoresens biovar V. Our technique using a microplate reader is able to measure 96 samples simultaneously. Thus real samples, blanks, standards and replicates can be determined concurrently, ensuring the accuracy of this method. The process requires approximately 20 min per determination, compared with 5 days for the standard BOD method. This method gives a linear response (r 2 = 0.943) to OECD synthetic sewage samples from 50 to 430 mg L −1.

The clinical outcome of childhood aplastic anaemia (AA) with aberrant cytogenetic clones at diagnosis was surveyed. Among 198 children with newly diagnosed AA registered with the AA Committee of the Japanese Society of Paediatric Hematology between 1994 and 1998, cytogenetic studies of bone marrow (BM) cells were completed in 159 patients. Apart from one Robertsonian translocation, seven patients (4.4%) showed clonal chromosomal abnormalities in hypoplastic BM without myelodysplastic features. The patients included six girls and one boy with a median age of 11 years (range 5-14 years). Six patients had del(6), del(5), del(13), del(20), or -7, and one showed add(9). Four patients responded to the first immunosuppressive therapy (IST: cyclosporin A plus anti-thymocyte globulin) and one obtained a spontaneous remission. Cytogenetic abnormalities remained in two patients with an IST response. On the other hand, two patients showed no IST response. One did not respond to repeat IST and died of acute graft-versus-host disease after an unrelated-BM transplant. Another obtained a complete response after a successful BM transplant. No haematological findings at diagnosis predicted the treatment response. No significant morphological changes developed during the course of the illness. A literature review revealed that half of 24 AA patients with chromosomal abnormalities responded to the first IST, and that +6 was the sole predictable marker for IST unresponsiveness. These results suggest that IST can be applied as the initial therapy for AA with cytogenetic abnormalities in the absence of completely matched donors.

The correlation between infant leukemia and in utero exposure to topoisomerase II (topo-II) inhibitor has been clarified. We examined the in vitro effect of topo-II inhibitor (etoposide) on cleavage of the MLL gene in cord and peripheral blood mononuclear cells (MNCs). Southern blot analysis showed cleavage of the MLL gene in peripheral blood MNCs of infants when the MNCs were exposed to etoposide. MNCs were incubated with etoposide at various concentrations (1 to 50 microM), and a ligation-mediated polymerase chain reaction (LM-PCR) was used to detect double strand breaks (DSBs) of DNA in intron 8 of the MLL breakpoint cluster region. PCR products obtained with LM-PCR were subcloned and sequenced to identify the breakpoint in the MLL gene. The PCR products indicated DSBs of the MLL gene were obtained without any difference in the incidence between 3 different samples (cord and peripheral blood from infants and children). Sequencing analysis showed that the DSBs occurred on the telomeric side of intron 8 and near exon 9. There was no evidence that the cord blood was more susceptible to MLL DNA breakage by topo-II inhibitor than were other cells. Instability of the partner gene during the fetal period could be associated with the pathogenesis of infant leukemia.

Le statut de « bourgeois » (chōnin) dans le Japon de l’epoque premoderne est le produit de la construction, au tournant des xvie et xviie siecles, d’une multitude de villes nouvelles ou se concentra la population des marchands et des artisans. Selon les principes d’organisation sociale et politique de cette epoque, le statut de bourgeois garantissait aux citadins un droit de propriete fonciere en echange de l’accomplissement d’un « service » (yaku) envers l’autorite publique seigneuriale, impose au bourgeois dans le cadre de communautes autonomes de rue ou de quartier. A partir de l’exemple des deux grandes metropoles shogounales d’Edo et d’Ōsaka au xviie siecle, l’article se penche sur l’evolution de quelques rues, en montrant les changements rapides qui affecterent le statut de bourgeois, du fait des modifications de la structure de la propriete dues aux transactions foncieres, mais aussi de la montee de la population locative.

In Saccharomyces cerevisiae , when a rich nitrogen source such as ammonium is added to the culture medium, the general amino acid permease Gap1p is ubiquitinated by the yeast Nedd4-like ubiquitin ligase Rsp5p, followed by its endocytosis to the vacuole. The arrestin-like Bul1/2p adaptors for Rsp5p specifically mediate this process. In this study, to investigate the downregulation of Gap1p in response to environmental stresses, we determined the intracellular trafficking of Gap1p under various stress conditions. An increase in the extracellular ethanol concentration induced ubiquitination and trafficking of Gap1p from the plasma membrane to the vacuole in wild-type cells, whereas Gap1p remained stable on the plasma membrane under the same conditions in rsp5 A401E and Δ end3 cells. A 14 C-labeled citrulline uptake assay using a nonubiquitinated form of Gap1p (Gap1p K9R/K16R ) revealed that ethanol stress caused a dramatic decrease of Gap1p activity. These results suggest that Gap1p is inactivated and ubiquitinated by Rsp5p for endocytosis when S. cerevisiae cells are exposed to a high concentration of ethanol. It is noteworthy that this endocytosis occurs in a Bul1/2p-independent manner, whereas ammonium-triggered downregulation of Gap1p was almost completely inhibited in Δ bul1/2 cells. We also found that other environmental stresses, such as high temperature, H 2 O 2 , and LiCl, also promoted endocytosis of Gap1p. Similar intracellular trafficking caused by ethanol occurred in other plasma membrane proteins (Agp1p, Tat2p, and Gnp1p). Our findings suggest that stress-induced quality control is a common process requiring Rsp5p for plasma membrane proteins in yeast.

This paper summarizes the features of ground damage toresidential fills due to the 1995 Hyogo-ken NanbuEarthquake in Japan. Many residential lands sufferedground damage to various extents in the earthquakeevent. Permanent deformation took place ingently-sloping residential fills. Some slopes asgentle as four degrees exhibited landslide-likedownward movements from dozens to hundreds ofcentimeters, so on. It is suggested that liquefactionmight be a dominant factor for displacements ingently-inclined residential fills.

A compact optical device and disposable strips for the simultaneous determination of the biochemical oxygen demand (BOD) of multiple samples have been developed. The compact optical device was constructed using three pairs of light emitting diodes (LEDs) and silicon photodiodes (Si-PDs), and the disposable strips were fabricated using inexpensive, transparent polycarbonate plates. Pseudomonas fluorescens bacteria were immobilizedin a layer at the bottom of each reaction cell on the sensor strip using a photocrosslinkable resin (ENT-2000). Using the redox electron acceptor 2,6-dichlorophenolindophenol sodium salt as the chromophore, a linear relationship was observed between the response of the Si-PDs and the BOD value of Organization of Economic Cooperation synthetic wastewater. Real samples from a wastewater treatment plant and a kitchen in our factory were examined, and good correlations were observed between the BOD values derived using this system and those determined by the conventional BOD5 method. In addition, in spite of the sensor response decreasing to approximately half its initial value after 2 weeks, the correlation coefficients (r2) of calibration curves remained above 0.9 upon storage of the strips in 0.1 M NaCl solution at 4°C for up to 6 weeks. In conclusion, this system is transportable and compact as well as being simple to operate and suitable for rapid, on-site measurements. © 2001 John Wiley & Sons, Inc. Field Analyt Chem Technol 5: 222–227, 2001

The pH, electrical conductivity, ammonia-nitrogen, chemical oxygen demand, carbon-to-nitrogen ratio, total organic carbon and biochemical oxygen demand (BOD5) of compost extract solution and the moisture content and organic matter content of solid compost samples were investigated as parameters for monitoring the composting process. It was observed that the BOD5 value of the compost sample decreased dramatically from 355 to 33 mg L−1 during the process. Compared with other parameters, the BOD5 value showed the largest change. However, the conventional method of BOD5 determination is not suitable for process control owing to its time-consuming nature and the complicated procedures required. Thus, compost monitoring using a mediator-type BOD sensor consisting of Pseudomonas fluorescens biovar. V and potassium ferricyanide was investigated. In order to realize rapid and reproducible measurements, we developed a 10-channel potentiostat system incorporating screen-printed disposable chips incorporating microorganisms immobilized in sodium alginate gel. Using sludge extract solutions as standard solutions for calibration, it was found that aqueous extracts of the compost were most suitable for BOD determination. In conclusion, the relative change in BOD sensor values determined using our system corresponded well with the BOD5 values obtained using the standard 5 d method during 58 d of composting.

The local and global critical current densities of polycrystalline bulks Sr 0.6 K 0.4 Fe 2 As 2 were investigated by both the remnant magnetic moment and Campbell’s methods. It was found that the global critical current density was improved by addition of Ag. It was speculated that the reason for the enhancement of the global critical current density was ascribed to the enlarged size of particle and the clean boundaries due to addition of Ag. It was also found that the critical current density estimated from usual magnetization measurement was a type of average value of both local and global critical current densities and can be explained by simple contributions from magnetic moments by local and global shielding currents. Therefore, the size of the grain was estimated by the present model and the agreement with the observation by SEM was obtained.