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Natural polyisoprene is a biopolymer consisting of isoprene units (C(5)H(8)) that is used commercially in household, medical, and industrial materials. For the management of natural polyisoprene production, the selection of high-yield polyisoprene-producing trees, and an understanding of polyisoprene biosynthesis, a high-throughput and highly sensitive screening method for the quantification of polyisoprene is required. In this study, we examined pyrolysates from polyisoprenes, polyprenols, carotenoids, ubiquinone (CoQ-10), and sterols by pyrolysis gas chromatography/mass spectrometry (PyGC/MS) and determined that the amounts of isoprene and limonene released from polyprenols and polyisoprenes were dependent upon their molecular weights. Based on these results, we developed a relative quantification method for polyisoprene in leaves by direct analysis of 1 mg of leaves using PyGC/MS. This novel quantification method eliminated extraction steps and can be used in the measurement of polyisoprene contents in Eucommia ulmoides and Hevea brasiliensis.

Beach changes along the coast between Kazusa-minato Port and Isone Point facing the Uraga Strait were investigated. Along the coastline not only the shoreline recession caused by the spatial imbalance of longshore sand transport, but also erosion of the sea cliffs composed of unconsolidated loose sand layers occurred. The long-term shoreline evolution in the area under study was investigated using past aerial photographs. The interaction between the shoreline recession and the erosion of sea cliffs was studied. On the basis of the field data, a model for predicting shoreline changes and the recession of sea cliffs composed of unconsolidated sand layers was developed.

Lignan is a large class of plant secondary metabolites, which has long attracted pharmacological interest because of its anti-tumor and estrogenic activities. Forsythia plants are known to produce a wide variety of lignans, such as (−)-matairesinol, (−)-secoisolariciresinol, (+)-pinoresinol, and (+)-phillygenin. The majority of such lignans are accumulated in glucoside forms. However, their glucosylation mechanisms largely remain to be elucidated. Here we describe the sequence, enzymatic activities, and gene expression profiles of UDP-sugar dependent-glycosyltransferases (UGT) from Forsythia koreana through a reverse-genetic approach. A Forsythia UGT, UGT71A18 protein, expressed in E. coli, preferentially glucosylated furofuran-class lignans, including (+)-pinoreisnol, (+)-epipinoreisnol, and (+)-phylligenin. Moroeover, the recombinant UGT71A18 exhibited specificity to UDP-glucose as a glycosyl donor. Gene expression analysis revealed that UGT71A18 is expressed predominantly in leaves and the suspension cell culture of F. koreana, and that the UGT71A18 transcript is upregulated in the transgenic cell culture expressing the RNAi construct of the pinoresinol lariciresinol reductase (PLR) gene, compared to non-transformants. These results are consistent with the remarkable elevation of pinoresinol glucosides in the PLR-RNAi lines. Collectively, the present data strongly suggests that UGT71A18, in part, is responsible for glucosylation of furofuran-class lignans, including (+)-pinoresinol and/or structurally related lignans in vivo.

Aims: We have evaluated the therapeutic efficacy and safety of pulse steroid therapy for ulcerative colitis (UC) in a Japanese pediatric population by means of a survey. Methods: A questionnaire on UC patients treated with therapy between 2002 and 2006 was sent to 37 members of the Japanese Society for Pediatric Inflammatory Bowel Disease. Results: 21 of 62 cases in 6 of 19 centers registered in this study had been treated with pulse steroid therapy. The success rate of remission induction with this treatment was 55%, and improvement was observed in all cases in which remission was not achieved. There were no reports of any obvious side effects. The most common reason for using pulse steroid therapy was for remission induction in relapsed cases. Over the course of 12 (or fewer) months, the number of cases in which remission was maintained was only 1 in 4. However, the amount of concomitant steroid use had significantly decreased after 1 year. Conclusions: This survey shows that in Japan, pulse steroid therapy is used for a relatively large number of children with UC and is as an effective method of remission induction that has few side effects.

On the Akashi coast, beach nourishment using gravel with the grain size ranging between 3 and 15 mm was begun in November 2008. The beach changes were monitored using the photographs from several fixed points. It was found that fine sand was deposited over the gravel layer under the calm wave conditions, whereas the nourished gravel was transported onshore and deposited in front of the seawall under the storm wave conditions. Assuming that the equilibrium slope changes in response to the change in thickness of sand layer, a model for predicting the deposition of fine sand over the gravel layer in beach nourishment using gravel and the shoreward movement of gravel during storms was developed on the basis of the contour-line-change model.

Lignans are a large class of secondary metabolites in plants, with numerous biological effects in mammals, including antitumor and antioxidant activities. Sesamin, the most abundant furofuran-class lignan in sesame seeds (Sesamum plants), is produced by the cytochrome P450 enzyme CYP81Q1 from the precursor lignan, pinoresinol. In contrast, Forsythia plants produce dibenzylbutyrolactone-class lignans, such as matairesinol, from pinoresinol via the catalysis of pinoresinol/lariciresinol reductase (PLR) and secoisolariciresinol dehydrogenase. Here we present the engineering of lignan biosynthesis in Forsythia cell suspension cultures for the development of an efficient production method of beneficial lignans. A suspension cell culture prepared from leaves of Forsythia koreana produced lignans, mainly pinoresinol and matairesinol glucosides, at levels comparable with that obtained from the leaves. In an attempt to increase the pinoresinol content in Forsythia, we generated a transgenic cell line overexpressing an RNA interference (RNAi) construct of PLR (PLR-RNAi). Down-regulation of PLR expression led to a complete loss of matairesinol and an accumulation of approximately 20-fold pinoresinol in its glucoside form in comparison with the non-transformant. Moreover, the Forsythia transgenic cells co-expressing CYP81Q1 and PLR-RNAi exhibited production of sesamin as well as accumulation of pinoresinol glucoside. These data suggest Forsythia cell suspension to be a promising tool for the engineering of lignan production. To the best of our knowledge, this is the first report on transgenic production of an exogenous lignan in a plant species.

Polyethylene wear particles induce macrophages to release cytokines, which can lead to osteolysis and aseptic loosening of total joint prostheses1. The generation of polyethylene wear particles is one of the most important factors that affects the midterm and long-term clinical results associated with total knee arthroplasty2,3. The generation of polyethylene wear particles is correlated with the activity level of the patient4, and greater demands are placed on a total knee prosthesis when it is implanted in a younger, more active patient5. Therefore, to achieve better long-term results for patients who have higher activity levels, modifications of materials (i.e., highly cross-linked polyethylene and alumina ceramic) and design (i.e., mobile bearing and medial pivot) have been developed to reduce polyethylene wear after total knee arthroplasty. It takes decades to evaluate the long-term results of newly introduced total knee prostheses. Thus, it is particularly important to examine in vivo polyethylene wear generation in new prostheses before they come into widespread use. It is difficult to determine the in vivo polyethylene wear of total joint prostheses (with the exception of total hips) with use of postoperative radiographs. To measure polyethylene wear in vivo, we developed and employed a method in which we isolated and analyzed polyethylene wear particles in the synovial fluid of knees that had a well-functioning total knee prosthesis6-9. The number, size, and shape of polyethylene wear particles have been reported to be critical factors in the development of osteolysis. Greater volume, submicrometer size, and an elongated shape of polyethylene wear particles all stimulate an increased macrophage response10-12. Our hypothesis was that modification of total knee arthroplasty materials and designs can influence the generation of polyethylene wear particles in vivo. In the present study, …

A new concept of IgG4-related disease characterized by a high serum IgG4 level and tissue infiltration of IgG4-positive plasmacytes that can involve salivary glands has been proposed. In this article, 2 patients with IgG4-related sclerosing sialadenitis involving the submandibular glands are reported. One patient presented with bilateral and painless swelling of the submandibular glands. He had already been treated with systemic prednisolone owing to the occurrence of retrobulbar neuritis. Laboratory examinations showed high serum IgG4 concentrations, and a biopsy of the submandibular gland revealed the infiltration of IgG4-positive plasmacytes. Abdominal computerized tomography demonstrated tumefaction in the tail of the pancreas, thus suggesting localized autoimmune pancreatitis. The other patient also showed bilateral and painless swelling of the submandibular glands, but there was no involvement of any other organs. These patients were effectively treated with corticosteroids, which resulted in a reduction of the swelling of the submandibular gland and increased saliva. When a patient is suspected of having sclerosing sialadenitis, it is important to consider that the patient may have a systemic IgG4-related plasmacytic disease.

The phytochrome photoreceptors regulate plant growth and development throughout their life cycle. Rice (Oryza sativa) possesses three phytochromes, phyA, phyB, and phyC. Physiological, genetic, and biochemical analyses of null mutants of each phytochrome have revealed the function of each in rice. However, few studies have investigated the relationship between phytochrome signaling and metabolism. In the present study, non-targeted metabolite analysis by gas chromatography time-of-flight mass spectrometry (GC/TOF-MS) and targeted metabolite analysis by capillary electrophoresis electrospray ionization mass spectrometry (CE/ESI-MS) were employed to investigate metabolic changes in rice phyA phyB phyC triple mutants. Distinct metabolic profiles between phyA phyB phyC triple mutants and the wild type (WT), as well as those between young and mature leaf blades, could be clearly observed by principal component analysis (PCA). The metabolite profiles indicated high accumulation of amino acids, organic acids, sugars, sugar phosphates, and nucleotides in the leaf blades of phyA phyB phyC triple mutants, especially in the young leaves, compared with those in the WT. Remarkable overaccumulation of monosaccharide, such as glucose (53.4-fold), fructose (42.5-fold), and galactose (24.5-fold), was observed in young leaves of phyA phyB phyC triple mutants. These metabolic phenotypes suggest that sugar metabolism, carbon partitioning, sugar transport, or some combination of these is impaired in the phyA phyB phyC triple mutants, and conversely, that phytochromes have crucial roles in sugar metabolism.

In recent plant hormone research, there is an increased demand for a highly sensitive and comprehensive analytical approach to elucidate the hormonal signaling networks, functions, and dynamics. We have demonstrated the high sensitivity of a comprehensive and quantitative analytical method developed with nanoflow liquid chromatography-electrospray ionization-ion trap mass spectrometry (LC-ESI-IT-MS/MS) under multiple-reaction monitoring (MRM) in plant hormone profiling. Unlabeled and deuterium-labeled isotopomers of four classes of plant hormones and their derivatives, auxins, cytokinins (CK), abscisic acid (ABA), and gibberellins (GA), were analyzed by this method. The optimized nanoflow-LC-ESI-IT-MS/MS method showed ca. 5-10-fold greater sensitivity than capillary-LC-ESI-IT-MS/MS, and the detection limits (S/N=3) of several plant hormones were in the sub-fmol range. The results showed excellent linearity (R(2) values of 0.9937-1.0000) and reproducibility of elution times (relative standard deviations, RSDs

In Hevea brasiliensis, laticifers produce and accumulate rubber particles. Despite observation using histochemical methods, development stage structure and structures with ceasing functions have rarely been described. Spectral confocal laser scanning microscopy with Nile red staining simplifies laticifer structure observation in tangential sections while enhancing the resolution. Laticifer and ray images were extracted from unmixed images and used to monitor changes during growth. A laticifer network structure developed from increased anastomoses between adjoining laticifers outside of the conducting phloem, but because of increased radial division and growth of rays, the network structure ruptured and disintegrated. We also investigated immunohistochemical localization of two rubber particle-associated proteins in the laticifers: small rubber particle protein (SRPP) and rubber elongation factor (REF). Mature bark test results show that SRPP is localized only in the laticifer layers in the conducting phloem; REF is localized in all laticifer layers. Because SRPP plays a positive role in rubber biosynthesis, results show that the rubber biosynthesis capability of laticifers is concentrated where rays and the sieve tube actively transport metabolites.

The production of oat (Avena sativa L.) phytoalexins, avenanthramides, occurs in response to elicitor treatment with oligo-N-acetylchitooligosaccharides. In this study, avenanthramides production was investigated by techniques that provide high spatial and temporal resolution in order to clarify the process of phytoalexin production at the cellular level. The amount of avenanthramides accumulation in a single mesophyll cell was quantified by a combination of laser micro-sampling and low-diffuse nanoflow liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) techniques. Avenanthramides, NAD(P)H and chlorophyll were also visualized in elicitor-treated mesophyll cells using line-scanning fluorescence microscopy. We found that elicitor-treated mesophyll cells could be categorized into three characteristic cell phases, which occurred serially over time. Phase 0 indicated the normal cell state before metabolic or morphological change in response to elicitor, in which the cells contained abundant NAD(P)H. In phase 1, rapid NAD(P)H oxidation and marked movement of chloroplasts occurred, and this phase was the early stage of avenanthramides biosynthesis. In phase 2, avenanthramides accumulation was maximized, and chloroplasts were degraded. Avenanthramides appear to be synthesized in the chloroplast, because a fluorescence signal originating from avenanthramides was localized to the chloroplasts. Moreover, our results indicated that avenanthramides biosynthesis and the hypersensitive response (HR) occurred in identical cells. Thus, the avenanthramides production may be one of sequential events programmed in HR leading to cell death. Furthermore, the phase of the defense response was different among mesophyll cells simultaneously treated with elicitor. These results suggest that individual cells may have different susceptibility to the elicitor.

A model for predicting bathymetric and grain size changes considering equilibrium slopes corresponding to the composition of grain size and each grain size was applied to predict the beach changes on the Chigasakinaka coast. The cross-shore sorting of sand, in which the grain size gradually decreases with increasing depth, and the formation of a gradually changing longitudinal slope were reproduced well. The diffusion of colored sand with different grain sizes placed on the shoreline and at a depth of 4 m was predicted and compared with the results of a field observation using colored sand on the Chigasakinaka coast. Both results were in good agreement.

In the southern part of Sanur Beach, Bali, beach erosion occurred between groins after the extensive beach nourishment, resulting in shoreline rotation between groins. The cause of shoreline changes were investigated by the field observation of bathymetric survey and sampling of bottom material, and numerical simulation of beach changes were carried out using Noshi et al.' (2008) model. It was found that the shoreline rotation was triggered by the dredging of reef flat, which caused the change in wave direction on the reef flat.

A model for predicting the formation and deformation of the multiple bars is developed. The model developed by Shimizu et al. (2008), which can predict 2-D formation of the multiple bars applying the contour-line-change model, is further expanded to predict 3-D formation and deformation of the multiple bars. Taking the beach changes around Fukude fishing port, where multiple bars developed before the construction of the breakwater, but it disappeared after the construction of the breakwater obstructing longshore sand transport as an example, numerical simulations were carried out. The formation and deformation of the multiple bars were predicted by the model.

近畿地方北部の敦賀・今津観測点においてボアホール型3成分歪計による観測が行われている。両観測点で得られた1996年から12.5年間のデータについて調べた。今津のデータに見られる季節変動は並行観測されている地下水位との相関が高く、敦賀のデータに見られる季節変動は降水量を入力としたタンクモデルを使って説明することができた。設置後の安定化過程を指数関数で近似し、これを除去した。季節変動、指数関数的減衰過程を除去した歪変化には、2000年頃と2005年頃に歪速度変化が見られる。この歪速度変化は機器の感度変化、および降水量の長期的変化では必ずしも説明できない。東海地域で同時期に発生したゆっくりすべりによって生じうる歪を計算して観測された非定常的な歪変化と比較したところ、歪の大きさは観測値の方が1桁大きいが主歪の方向は整合的だった。

Eucommia ulmoides Oliver is a deciduous tree distributed in China, and is the only species in its genus, and the only genus in its family. It grows in temperate zones, and produces long-chain trans-polyisoprene throughout the entire plant. This trans-polyisoprene (TPI) derived from Eucommia is called Eucommia rubber (EU-rubber), and is a hydrocarbon with a carbon molecular weight of a few hundred thousand. EU-rubber is a hard rubber with thermoplasticity, and has properties similar to plastic. In terms of practical use, these plant-derived polymers are attracting attention as an industrial raw material to substitute for petroleum, and demand is expected to grow in the future. Hydrocarbon is also a fuel which has energy with the same value as petroleum.

Applications of metabolomics techniques along with chemometrics provide an understanding in the relationship between metabolome of green tea and its quality. A coupled of ultra-performance liquid chromatography with time-of-flight mass spectrometry (UPLC/TOF MS) allowed a high-throughput and comprehensive analysis with minimal sample preparation. Using this technique, a wide range of metabolites were investigated. Data analysis was rapid, considering that the fingerprinting technique was performed. A set of green tea samples from 2006 tea contest of the Kansai area was analyzed to prove usefulness of the developed technique. Green tea with different qualities were discriminated through principal component analysis (PCA). Consequently, projection to latent structure by means of partial least-squares (PLS) was performed to create a constructive quality-predictive model by means of metabolic fingerprinting. Beside epigallocatechin, other predominant catechins, including epigallocatechin gallate and epicatechin gallate, detected in green tea were found to be significant biomarkers to the high quality of Japanese green tea (Sencha).

This study was designed to investigate the efficacy of lipid-lowering agents in preventing steroid-induced osteonecrosis and the mechanism by which they do so in a rabbit model.Female Japanese white rabbits were randomly allocated to receive probucol (group P), pravastatin (group PS), simvastatin (group SS), or saline (group C) for 6 weeks (n = 15 in groups P, PS, and SS; n = 30 in group C). Methylprednisolone (20 mg/kg) was injected at 3 weeks after starting treatment, and the femurs were histologically examined bilaterally 3 weeks after methylprednisolone injection. Midazolam clearance was measured before treatment and before methylprednisolone injection to determine hepatic cytochrome P4503A (CYP3A) levels.The incidence of osteonecrosis in the proximal metaphysis of the femurs in groups PS and SS was significantly lower than in group C (P0.05 and P0.0001, respectively), whereas it did not differ between groups P and C. It was significantly lower in group SS than in group PS (P0.05). Plasma concentrations of lipids (low-density lipoprotein, triglyceride, free fatty acid, and total cholesterol) in groups P, PS, and SS were significantly lower than in group C; and hepatic CYP3A levels were significantly higher in group SS than in groups P or PS after treatment (P0.005 for both).Simvastatin and pravastatin significantly reduced the incidence of steroid-induced osteonecrosis in rabbits. Simvastatin was more effective in reducing the incidence of the disease, and increased CYP3A activity is a possible mechanism for this effect.

Hevea brasiliensis Müll. Arg. is a tree that produces natural rubber, an industrially vital isoprenoid polymer. Biosynthesis of natural rubber is known to take place biochemically by a mevalonate (MVA) pathway, but molecular biological characterization of related genes has been insufficient. From H. brasiliensis, we obtained full-length cDNA of genes encoding all of the enzymes that catalyze the six steps of the MVA pathway. Alignment analysis and phylogenetic analysis revealed that in H. brasiliensis there are three acetyl-CoA acetyltransferase genes, two HMG-CoA synthase (HMGS) genes, and four HMG-CoA reductase (HMGR) genes. Gene expression analysis by type of tissue indicated that MVA pathway genes were highly expressed in latex, as compared to other types of tissue and that HMGS and HMGR, which exist in multiple copies, have different expression patterns. Moreover, these MVA pathway genes in H. brasiliensis were found to complement MVA pathway deletion mutations in yeast.

In this paper, we report a novel method for delivering genes into chloroplasts of tobacco cells using laser microablation. The plasmid pLD200-GFP was introduced into chloroplasts of Nicotiana tabacum cv. Xanthi guard cells and transient GFP expression was detected in the chloroplasts after 2-3 d of incubation. The technique uses an argon fluoride (ArF) excimer laser to perforate the cell surface in a 4 mum(2) area in the presence of plasmid coated gold microparticles. Pretreatment of guard cells to promote stomatal closure prior to laser ablation resulted in a significant increase in the survival rate of cells and a transient expression rate of 2-3% in trial number basis was archived. Our method has unique advantages such as avoiding laborious pretreatments that adversely affect cell viability and specific delivery of transgenes into a desired cell in complex leaf tissue. This technique is a potential tool for cell specific transient gene expression studies for elucidation of gene regulation and expression.

Commercial development of trans-1,4-polyisoprene from Eucommia ulmoides Oliver (EU-rubber) requires specific knowledge on selection of high-rubber-content lines and establishment of agronomic cultivation methods for achieving maximum EU-rubber yield. The development can be facilitated by high-throughput and highly sensitive analytical techniques for EU-rubber extraction and quantification. In this paper, we described an efficient EU-rubber extraction method, and validated that the accuracy was equivalent to that of the conventional Soxhlet extraction method. We also described a highly sensitive quantification method for EU-rubber by Fourier transform infrared spectroscopy (FT-IR) and pyrolysis-gas chromatography/mass spectrometry (PyGC/MS). We successfully applied the extraction/quantification method for study of seasonal changes in EU-rubber content and molecular weight distribution.

In this study, metabolite profiling was demonstrated as a useful tool to plot a specific metabolic pathway, which is regulated by phytochrome A (phyA). Etiolated Arabidopsis wild-type (WT) and phyA mutant seedlings were irradiated with either far-red light (FR) or white light (W). Primary metabolites of the irradiated seedlings were profiled by gas chromatography time-of-flight mass spectrometry (GC/TOF-MS) to obtain new insights on phyA-regulated metabolic pathways. Comparison of metabolite profiles in phyA and WT seedlings grown under FR revealed a number of metabolites that contribute to the differences between phyA and the WT. Several metabolites, including some amino acids, organic acids, and major sugars, as well as putrescine, were found in smaller amounts in WT compared with the content in phyA seedlings grown under FR. There were also significant differences between metabolite profiles of WT and phyA seedlings during de-etiolation under W. The polyamine biosynthetic pathway was investigated further, because putrescine, one of the polyamines existing in a wide variety of living organisms, was found to be present in lower amounts in WT than in phyA under both light conditions. The expression levels of polyamine biosynthesis-related genes were investigated by quantitative real-time RT-PCR. The gene expression profiles revealed that the arginine decarboxylase 2 (ADC2) gene was transcribed less in the WT than in phyA seedlings under both light conditions. This finding suggests that ADC2 is negatively regulated by phyA during photomorphogenesis. In addition, S-adenosylmethionine decarboxylase 2 and 4 (SAMDC2 and SAMDC4) were found to be regulated by phyA but in a different manner from the regulation of ADC2.

The extraction of acoustic features for robust speech recognition is very important for improving its performance in realistic environments. The bi-spectrum based on the Fourier transformation of the third-order cumulants expresses the non-Gaussianity and the phase information of the speech signal, showing the dependency between frequency components. In this letter, we propose a method of extracting short-time bi-spectral acoustic features with averaging features in a single frame. Merged with the conventional Mel frequency cepstral coefficients (MFCC) based on the power spectrum by the principal component analysis (PCA), the proposed features gave a 6.9% relative lower a word error rate in Japanese broadcast news transcription experiments.

SummaryUnderstanding the mechanisms of nephron repair is critical for the design of new therapeutic approaches to treat kidney disease. The kidney can repair after even a severe insult, but whether adult stem or progenitor cells contribute to epithelial renewal after injury and the cellular origin of regenerating cells remain controversial. Using genetic fate-mapping techniques, we generated transgenic mice in which 94%–95% of tubular epithelial cells, but no interstitial cells, were labeled with either β-galactosidase (lacZ) or red fluorescent protein (RFP). Two days after ischemia-reperfusion injury (IRI), 50.5% of outer medullary epithelial cells coexpress Ki67 and RFP, indicating that differentiated epithelial cells that survived injury undergo proliferative expansion. After repair was complete, 66.9% of epithelial cells had incorporated BrdU, compared to only 3.5% of cells in the uninjured kidney. Despite this extensive cell proliferation, no dilution of either cell-fate marker was observed after repair. These results indicate that regeneration by surviving tubular epithelial cells is the predominant mechanism of repair after ischemic tubular injury in the adult mammalian kidney.

This paper describes a practical method to quantify anionic metabolites using contemporary capillary electrophoresis-electrospray ionization-tandem mass spectrometry (CE-ESI-MS/MS). The use of sulfonated capillary permitted a simultaneous analysis of sugar phosphates, organic acids, nucleotides and coenzyme A compounds with only one CE condition. This capillary also improved the repeatability and sensitivity. MS/MS with multiple reaction monitoring (MRM) detection was utilized to achieve significant selectivity and sensitivity. Under optimized CE-ESI-MS/MS system, the detection limits of 53 metabolites at signal-to-noise ratio of 3 were between 0.040 and 8.8 mumol/l. The relative standard deviations (RSDs) for the majority anionic metabolites were better than 0.5% for migration times, and better than 10.0% for peak areas (n=6). Matrix effects by contaminants in sample solution in CE-ESI-MS/MS analysis were removed dramatically by the sample preparation method with liquid-liquid fractionation and ultrafiltration procedure. Furthermore, the developed method was successfully applied to determine anionic metabolites in a cultured cell of Catharanthus roseus. Accumulation of some metabolites including shikimate, malate, and sedoheptulose 7-phosphate by elicitation of methyl jasmonate was observed. The result would show shikimate, tricarboxylic acid and pentose phosphate pathways were activated. Our method will be useful for detailed analysis of primary metabolism dynamics.

Objectives: This study was designed to compare professional oral care (POC) by a dental hygienist with tooth brushing and mouth rinsing by patients themselves according to the instructions of a nurse (control). Methods: Forty patients were randomly assigned to either the POC group (n = 20) or control group (n = 20). The presence of plaque and bacteria was assessed clinically. Results: One patient in the POC group and three patients in the control group dropped out because of exacerbation of underlying disease or death. Plaque control record scores were significantly lower in the POC group than in the control group on the fifth hospital day and the day of discharge. There was no significant difference between the groups in the detection rate of Candida species; and nosocomial pathogens on either day. Conclusions: Professional oral care by a dental hygienist is more effective than tooth brushing and mouth rinsing by patients themselves according to the instructions of a nurse.

A 77-year-old man with a painless and bilateral submandibular swelling was referred to our hospital. He had already been treated with systemic prednisolone for retrobulbar neuritis. Laboratory examinations showed high serum IgG4 concentrations, and a biopsy of the submandibular gland revealed the infiltration of IgG4-positive plasmacytes. Abdominal CT demonstrated tumor formation in the tail of the pancreas. These findings suggested the patient to have Mikulicz's disease associated with localized autoimmune pancreatitis. Treatment with prednisolone improved the swelling of the submandibular gland. As a result, when a paticent is suspected to have Mikulicz's disease, it is important to consider the possibility of systemic IgG4-related plasmacytic disease.

Two-dimensional quantitative analysis of microbeam X-ray diffraction (XRD) was performed using a transmission optical system to examine biological apatite (BAp) orientation in an isolated trabecula of a human fourth lumber vertebral body. The incident X-ray beam is 20 μm in diameter, which is small enough for the isolated trabecula despite a slight beam divergence of 0.2°. Integrated intensities of (002) and (310) are obtained separately by different incident angles and detector positions. Distribution of the preferential orientation of the BAp c-axis is finally calculated quantitatively as an integrated intensity ratio of (002)/(310) in a plane containing the trabecular direction. Preferential alignment of the BAp c-axis was finally determined to be perfectly parallel to the fiber direction in a rod-shaped trabecula, since accurate one-dimensional alignment is different from the alignment in the femoral cortical bone as a long bone that shows the local maximum of preferential alignment perpendicular to the longitudinal bone axis. For example, the integrated intensity ratio of (002)/(310) has a maximum value of 16 along the trabecular fiber and a minimum value of 0.09 in the perpendicular direction. Using this method, the anisotropy of BAp orientation in the trabecular bone can be quantitatively evaluated in the plane including the trabecular fiber. Thus, we successfully obtained a methodology that two-dimensionally analyzes the distribution of the BAp c-axis along all axes within a plane in a bone specimen.

Background and Aims: Azathioprine (AZA) and 6-mercaptopurine (6-MP) have recently been used in Japanese pediatric patients with ulcerative colitis. The aims of this study were to evaluate both the therapeutic efficacy and safety of AZA/6-MP in this group of patients. Methods: Fourteen members of the Japanese Society for Pediatric Inflammatory Bowel Disease reported 35 retrospective cases that received AZA/6-MP and were evaluated for adverse drug effects. In those who tolerated AZA/6-MP, disease activity and corticosteroid doses before and during the first 6 months of therapy were assessed. Results: AZA or 6-MP was safely used in 21 of 35 patients (60%) without adverse drug effects. The disease activity began to decrease from the first month of therapy and the maximum effect was achieved after 3 months. The mean daily prednisolone dose was decreased from 26.9 to 11.6 mg and dose reduction was achieved in 58% of patients after 6 months of therapy. Fourteen of the 35 patients (40%) experienced adverse drug effects, including leukopenia (n = 11), aplastic anemia (n = 1), pancreatitis (n = 1) and liver dysfunction (n = 1). Conclusions: The majority of Japanese children with ulcerative colitis tolerated AZA/6-MP and experienced favorable effects. However, 40% experienced adverse drug effects, mainly myelosuppression.

On the basis of cross-shore sand transport under the action of long period waves and standing waves reflecting from the seabed slope given by Bowen (1980) and Katoh (1984), a model for predicting the development of multi-bar system was proposed by applying the contour-line-change model. The increase in the intervals between the bars and the decay of the relative height of bars in the cross-shore direction were predicted well. The possibility of the recovery of the multi-bar system by beach nourishment using fine sand is also shown.

2004年9月5日に紀伊半島南東沖で発生した地震（Mj7.4）は南海トラフ軸付近で発生した津波を伴う大地震である。この地震の発震機構は南北方向に圧縮軸を持つ逆断層型で、フィリピン海プレート内あるいは上部マントルで発生したものと解釈されている。我々はこの地震の詳細な余震分布とその時空間推移を調査するため、自己浮上式海底地震計（OBS）による余震観測を実施した。余震観測は2004年9月から2005年8月までの期間、断続的に3回にわたって実施された。震源決定に用いた1次元P波の速度構造はOBSの観測海域で実施された速度構造探査結果を参照して求めた。また震源の決定精度を向上させるため、PS変換波を用いた堆積層補正をおこなった。その結果、気象庁一元化震源に比べ詳細かつ高精度な余震分布を得ることができた。OBS観測から求めた余震の震源は気象庁一元化震源と比較すると、深さは20kmほど浅く5～30kmに分布し、震央については全体的に南東方向に10kmほどシフトした。気象庁一元化震源では余震活動は主にトラフ軸より陸側に分布するように見えていたが、主たる余震活動はトラフ軸沿いで発生していることがわかった。また、余震活動は深さ5～10 kmの比較的浅い地震群と、深さ15～30kmの比較的深い地震群の2群に分かれていることが見出された。前者は余震域の中央部から北部にかけてのフィリピン海プレート内およびその上部の付加体内で発生しており、後者は前震および本震の破壊域と思われる海溝軸下の上部マントルで発生していることがわかった。さらに、浅い地震群の中にいくつかの地震クラスターが見出された。これらの地震クラスターは付加体からフィリピン海プレート内までほぼ鉛直に下降しているとみられる。これら余震域の中で発見された地震クラスターは、超低周波地震の発生源となっている付加体の分岐断層との関係において興味が持たれる。

Long-term shoreline changes of the Kita-shitaura and Miura coasts were investigated on the basis of the aerial photographs and field observations. Beach erosion has been severe on the Kita-shitaura coast, because of the recurrent dredging of the navigation channel of Kita-shitaura fishing port and the disposal. In contrast, on the Miura coast, dredged materials were placed on the shoreline, resulting in widening of the foreshore. Thus, in this area, sand balance has been lost due to the anthropogenic factors. To improve this situation, the recycling of dredged sand is required instead of constructing hard structures.

There is little information available regarding the sagittal mechanical axis of the lower extremity of normal subjects under weight-bearing conditions. The purpose of this study was to determine the sagittal alignment of the lower extremity under such conditions. Anteroposterior and lateral radiographs were taken of the 20 lower extremities of 10 healthy male Japanese volunteers (mean age, 27 years) while standing. The coronal mechanical axis passed through 33.9% medial to the proximal tibial articulating surface. The sagittal mechanical axis passed through 38.0% anterior to the distal femoral condyle and 27.9% anterior to the proximal tibial articulating surface, and also passed 5.2 mm anterior to the intercondylar notch. Our study therefore showed that the coronal and sagittal mechanical axes of the lower extremity do not always pass through the center of the knee. This has important implications for alignment in surgery of lower extremities such as total knee arthroplasty and osteotomy.

Classification of tea quality is now mainly performed according to the sensory results by professional tea tasters. However, this evaluation method is inconsistent in differentiating their qualities. A combination of a (1)H NMR technique and a multivariate analysis was introduced to the quality evaluation of green tea by means of a metabolomic technique. A broad range of metabolites were detected by (1)H NMR spectrometry. The principal component analysis (PCA) was used to reduce the complexity of the (1)H NMR spectra data set and provided the quality discrimination result. It offered an extensive clue for classification and quality assessment without any prepurification method. A set of green teas from a Japanese tea contest were analyzed by (1)H NMR to classify the quality with respect to that judged by tea tasters and to conceive a quality prediction model. Metabolic profiling and fingerprinting of (1)H NMR spectra of green teas with different quality were studied. PCA showed a separation between the high- and the low-quality green teas. The taste marker compounds contributing to the discrimination of tea quality were identified. Reliable prediction models were obtained by the partial least-squares projection to latent structure (PLS) analysis together with a preprocessing filter of both orthogonal signal correction (OSC) and a combination between OSC and wavelet transform algorithms.

Isoprenoids are synthesized through the condensation of five-carbon intermediates, isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), derived from two distinct biosynthetic routes: cytosolic mevalonate (MVA) and plastidial 2-C-methyl-D: -erythritol 4-phosphate (MEP) pathways. 1-Hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase (IDS; EC 1.17.1.2), which catalyzes the last step of MEP pathway, was cloned as a multicopy gene from gymnosperms Ginkgo biloba (GbIDS1, GbIDS2, and GbIDS2-1) and Pinus taeda (PtIDS1 and PtIDS2), and characterized. Phylogenetic tree constructed with other plant IDSs demonstrated gymnosperm IDSs were distinctively different from angiosperm IDSs. The gymnosperm IDS clade contained two subclades, one composed of GbIDS1 and PtIDS1, and the other composed of GbIDS2, GbIDS2-1, and PtIDS2. G. biloba IDSs, except GbIDS2-1, successfully complemented Escherichia coli DLYT1, a lytB disruptant, confirming the in vivo competency of isozymes. During the 4 weeks study period, although transcript levels of GbIDS1s were similar both in roots and leaves of cultured G. biloba embryo, the transcripts of GbIDS2 predominantly occurred in the embryo roots, where diterpene ginkgolides are biosynthesized. Levels of PtIDS2 transcripts in the diterpenoid resin-producing wood were 4-5 times higher than those in other tissues. Higher levels of GbIDS1 transcripts were induced by light, whereas those of GbIDS2 were increased by methyl jasmonate treatment. These results strongly imply GbIDS2 and PtIDS2 have high correlation with secondary metabolism. In Arabidopsis transient expression system, N-terminal 100 amino acid residues of GbIDS1 delivered fused GFP protein into chloroplast as well as cytosol and nucleus, whereas those of GbIDS2, GbIDS2-1, and two PtIDSs delivered GFP only into chloroplast.

Periploca sepium Bunge (Chinese silk vine) is a woody climbing vine belonging to the family Asclepiadaceae. It originally comes from Northwest China. Periploca resembles the Para-rubber tree, Hevea brasiliensis, regarding a similar body plan to produce a milky exudate containing rubber latex. The Periploca plant was assessed as a rubber-producing plant by rubber structure elucidation and its molecular weight distribution. A rubber fraction purified from the milky exudate was subjected to 1H NMR analysis, and a characteristic signal derived from cis-polyisoprene was observed. In addition, when the molecular weight distribution of rubber components in the exudate was measured (using size-exclusion chromatography), the number-average molecular weight (Mn), weight-average molecular weight (Mw), and polydispersity (Mw/Mn) were estimated to be Mn = 1.3 × 105, Mw = 4.1 × 105, and Mw/Mn = 3.1, respectively. Furthermore, the presence of polyisoprene, with Mn = 4.0 × 104, Mw = 7.6 × 104, and Mw/Mn = 2.5, was also confirmed in plantlets obtained from shoots as a result of tissue culture

Novel tools are needed for efficient analysis and visualization of the massive data sets associated with metabolomics. Here, we describe a batch-learning self-organizing map (BL-SOM) for metabolome informatics that makes the learning process and resulting map independent of the order of data input. This approach was successfully used in analyzing and organizing the metabolome data forArabidopsis thaliana cells cultured under salt stress. Our 6 × 4 matrix presented patterns of metabolite levels at different time periods. A negative correlation was found between the levels of amino acids and metabolites related to glycolysis metabolism in response to this stress. Therefore, BL-SOM could be an excellent tool for clustering and visualizing high dimensional, complex metabolome data in a single map.

We describe the cloning and functional characterization of the fatty acid elongase gene HpELO2, a homologue of the HpELO1 gene required for the production of C24:0 in the yeast Hansenula polymorpha. The open reading frame (ORF) of HpELO2 consists of 1,035 bp, encoding 344 amino acids, sharing about 65% identity with that of Saccharomyces cerevisiae Elo2. Expression of HpELO2 rescued the lethality of the S. cerevisiae elo2Delta elo3Delta double disruptant. An accumulation of C18:0 and a significant increase and decrease in the levels of C24:0 and C26:0, respectively, were observed in the Hpelo2Delta disruptant. These results supported an idea that HpELO2 encodes a fatty acid elongase involved in the elongation of C18:0 to very long-chain fatty acids. The Hpelo1Delta Hpelo2Delta double disruption was nonviable, suggesting that HpELO1 and HpELO2 are the only two genes necessary for the biosynthesis in H. polymorpha. Interestingly, transcription of HpELO2 and HpELO1 were found to be transiently up-regulated by exogenous long-chain fatty acids; however, this up-regulation was not observed with HpELO1 and HpELO2 genes driven by the constitutively expressed promoter of the HpACT gene, suggesting that exogenous fatty acids specifically trigger the transcriptional induction of HpELO1 and HpELO2 through their promoter regions.