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Novel biomarkers of rheumatoid arthritis (RA), in addition to antibodies against cyclic citrullinated peptides, are required. Metabolome analysis is a promising approach to identify metabolite biomarkers for clinical diagnosis. We adopted a comprehensive non-targeted metabolomics approach combining capillary electrophoresis time-of-flight mass spectrometry (TOFMS) and liquid chromatography TOFMS. We constructed metabolomics profiling of 286 plasma samples of a Japanese population [92 RA patients, 13 systemic lupus erythematosus (SLE) patients and 181 healthy controls). RA case–control association tests showed that seven metabolites exhibited significantly increased levels in RA samples compared with controls (P < 1.0 × 10−4; UTP, ethanolamine phosphate, ATP, GDP, ADP, 6-aminohexanoic acid and taurine), whereas one exhibited a decreased level (xanthine). The plasma levels of these eight metabolites were not significantly different between seropositive and seronegative RA patients (P > 0.05; n = 68 and 24, respectively). The four nucleotide levels (UTP, ATP, GDP and ADP) were significantly higher in the non-treatment patients in comparison between patients with and without treatment (P < 0.014; n = 57 and 35, respectively). Furthermore, we found that none of the four nucleotide levels showed significant differences in SLE case–control association tests (P > 0.2; 13 patients with SLE and the 181 shared controls) and psoriatic arthritis (PsA) case–control association tests (P > 0.11; 42 patients with PsA and 38 healthy controls), indicating disease specificity in RA. In conclusion, our large-scale metabolome analysis demonstrated the increased plasma nucleotide levels in RA patients, which could be used as potential clinical biomarkers of RA, especially for seronegative RA. [ABSTRACT FROM AUTHOR]

The basidiomycetous yeast Pseudozyma antarctica GB-4(0) esterase (PaE) is a promising candidate for accelerating degradation of used biodegradable plastics (BPs). To increase safety and reduce costs associated with the use of PaE, we constructed a self-cloning strain with high-PaE productivity. A Lys12 gene (PaLYS12)-deleted lysine auxotroph strain GB4-(0)-L1 was obtained from GB-4(0) by ultraviolet mutagenesis and nystatin enrichment. Subsequently, the PaE gene (PaCLE1) expression cassette consisting of GB-4(0)-derived PaCLE1, under the control of a xylose-inducible xylanase promoter with PaLYS12, was randomly introduced into the GB4-(0)-L1 genome. A PaE high-producing strain, PGB474, was selected from among the transformants by high throughput double-screening based on its ability to degrade emulsified polybutylene succinate-co-adipate. Quantitative PCR revealed that four copies of the PaE gene expression cassette were introduced into the PGB474 genome. PGB474 produced 2.0 g/L of PaE by xylose-fed-batch cultivation using a 3-L jar fermentor for 72 h. Pseudozyma antarctica self-cloning strain that produces a high volume of biodegradable plastic degrading enzyme was constructed and selected by high throughput screening. [ABSTRACT FROM AUTHOR]

The article presents a study which examines the two isolates ACT04T and AC05, isolated from the red ginger gathered from Chiang Mai, Thailand. Based on 16S rRNA gene sequences, the isolates were included within a group of different genera and were formed as an independent cluster along with a certain type of strain. The study suggests that based on the information obtained, the two isolates can be classified into a separate genus, the Ameyameae chiangmaiensis gen. nov., sp. nov.

Phylogenetic relationships of the species belonging to the genus Myxococcus were elucidated based on the sequences of 16S rRNA genes and 16S–23S rRNA gene internal transcribed spacer (ITS) regions. The Myxococcus species were consequently classified into four distinct groups. The type strain of Myxococcus coralloides occupied an independent position (Group 1); it has been recently reclassified as Corallococcus coralloides. Group 2 comprised the type strains of both Myxococcus virescens and Myxococcus xanthus, and some strains assigned to Myxococcus flavescens. The type strain of M. flavescens was contained in Group 3 along with the strains of Myxococcus fulvus. Group 4 included the strains belonging to C. coralloides, M. fulvus, and M. stipitatus. The type strain of M. fulvus that was allocated outside Group 4 in the 16S rRNA gene tree belonged to Group 3 in the ITS tree. These results strongly suggest that the morphological characteristics of Myxococcus species are not consistent with the phylogenetic relationships. The Myxococcus species must therefore be redefined according to the phylogenetic relationships revealed in this study. [ABSTRACT FROM AUTHOR]

The article reports on results of a study of Tanticharoenia sakaeratensis, an osmotolerant acetic acid bacterium in the alpha-proteobacteria. A description of the study design and method is presented. The study concluded that three strains from Tanticharoenia sakaeratensis had DNA base compositions. It adds that the three strains did not oxidize acetate or lactate.

Abstract: Objective: The use of pig hearts can solve the problem of shortage of donor hearts for transplantation. However, targeting rejection by single genetic modification was proven to be ineffective, highlighting the requirement for complex genetic modifications and more effective methods for transgenic animal production. We evaluated here whether hearts of hDAF transgenic pigs generated using our technique sperm-mediated gene transfer (SMGT) will be protected from structural damage, metabolic changes, and mechanical dysfunction during perfusion with human blood. Methods: Hearts from control (C, n =6) or transgenic (T, n =5) pigs were perfused ex vivo for 4 h with fresh human blood using the ex vivo working mode system allowing monitoring of the function, metabolism, and structure. Results: Cardiac output (mean±SEM) was maintained in T constant throughout the experiment, at 3.58±0.36 and 3.83±0.14 l/min after 30 min and 4 h, respectively, while cardiac output decreased to 1.95±0.35 l/min in C after 30 min of perfusion (p <0.01 vs. T). The maximum increase in coronary perfusion pressure was reduced in T to 154±16% as compared to C (237±10%, p <0.001). Myocardial ATP after 4 h was 21.1±1.1 nmol/mg dry wt (similar to initial) in T, while it decreased in C to 17.2±1.4 (p <0.05). Deposition of complement factors C3 and C5b9 was present in C but not in T after perfusion. Conclusion: We have shown that hearts from hDAF transgenic pigs produced by SMGT are protected during perfusion with human blood and are metabolically stable and maintain mechanical function above the threshold level for life support. [Copyright &y& Elsevier]

Abstract: Objective: We hypothesized that modification of the infusion route may improve the efficiency of superoxide dismutase (SOD)-induced cardioprotection against reperfusion injury. The routes for SOD delivery previously examined were intravenous, via the left atrium, or by a combination of these, all of which can deliver SOD into the ischemic myocardium only after reperfusion. In contrast, retrograde intracoronary infusion may be able to deliver SOD before reperfusion. We investigated the feasibility and efficiency of the retrograde intracoronary infusion of SOD to attenuate reperfusion injury. Methods and results: Lewis rats underwent 30-min left coronary artery occlusion followed by reperfusion for 24 h. Just before reperfusion, CuZn–SOD was administered intravenously (15,000 U/kg, V-SOD group) or by retrograde intracoronary infusion (1500 U/kg, R-SOD group) through a catheter inserted into left cardiac vein via left superior vena cava as we have previously reported. This method has been shown to perfuse the whole left ventricular free walls. Controls for each group were injected with phosphate buffer saline only via the same routes (V-PBS and R-PBS group). The R-SOD group demonstrated significantly preserved left ventricular ejection fraction (LVEF; 71.3±1.7% vs. 60.8±2.3%, p =0.028), reduced infarct size (23.3±2.3% vs. 42.4±3.5%, p <0.001), and attenuated polymorphonuclear leukocyte (PMNL) infiltration (11.8±0.4 vs. 14.8±0.2 103/mm2, p <0.001) compared to the V-SOD group. The V-SOD group demonstrated significantly improved reflow (64.3±2.1% vs. 53.4±2.4%, p =0.017) and attenuated PMNL infiltration (14.8±0.2 vs. 16.8±0.7 103/mm2, p =0.018) compared to the V-PBS group. Conclusion: Retrograde intracoronary infusion is a promising, clinically applicable method to enhance the efficacy of SOD-induced myocardial protection against ischemia–reperfusion injury. [Copyright &y& Elsevier]

Objective: Heat shock protein 72 (HSP72) is involved in the myocardial self-preservation system under several conditions such as ischemia–reperfusion injury or late preconditioning. However, its mechanism is not fully understood. Ecto-5′-nucleotidase is a key enzyme for synthesizing adenosine and plays an important role in ischemic preconditioning. In this study, we tested the hypothesis that ecto-5′-nucleotidase plays a role in the cardioprotection of HSP72. Methods: Rat hearts (H group, n=6) were transfected with HSP72 gene by an intracoronary infusion of hemagglutinating virus of Japan (HVJ)–liposome complex. Control hearts (C group, n=6) were transfected with the β-galactosidase gene. Following 30 min of normothermic ischemia, grafts were reperfused using Langendorff apparatus. Results: The activity of ecto-5′-nucleotidase was significantly higher in H group than C group both before and after ischemia–reperfusion (H vs. C; 0.51±0.05 vs. 0.29±0.06, and 1.41±0.15 vs. 0.85±0.11 nmol/mg protein/min, P<0.05). H group also showed significant better functional recoveries than C group (P<0.05), as well as less creatine phosphokinase leakage (4.4±2.8 vs. 14.2±3.4 mU/min, P<0.05) and higher adenosine release (247.5±35.1 vs. 54.3±1.7 pmol/min, P<0.05). Administration of α,β-methylene adenosine diphosphate (AMP-CP), an inhibitor of ecto-5′-nucleotidase, significantly diminished the tolerance to ischemia–reperfusion injury in H group (P<0.05). Conclusion: These results demonstrated that ecto-5′-nucleotidase activated by an overexpression of HSP72 attenuated ischemia–reperfusion injury in the rat myocardium. They suggest that ecto-5′-nucleotidase plays a role in the cardioprotective effects of HSP72 in rat hearts. [ABSTRACT FROM PUBLISHER]

The cDNAs encoding putrescine N-methyltransferase (PMT), which catalyzes the S-adenosylmethionine-dependent N-methylation of putrescine at the first committed step in the biosynthetic pathways of tropane alkaloids, were isolated from Atropa belladonna and Hyoscyamus niger. These PMTs, however, lacked the N-terminal tandem repeat arrays previously found in Nicotiana PMTs. AbPMT1 RNA was much more abundant in the root of A. belladonna than was AbPMT2 RNA. The 5′-flanking region of the AbPMT1 gene was fused to the β-glucuronidase (GUS) reporter gene and transferred to A. belladonna. Histochemical analysis showed that GUS is expressed specifically in root pericycle cells and that the 0.3-kb 5′-upstream region was sufficient for pericycle-specific expression. Treatment of A. belladonna roots with methyl jasmonate did not up-regulate the expression of GUS or endogenous AbPMT genes. The regulation of tropane alkaloid biosynthesis is discussed and compared with that of nicotine biosynthesis. [ABSTRACT FROM AUTHOR]

Four cDNAs for spermidine synthase (SPDS), which converts the diamine putrescine to the higher polyamine spermidine using decarboxylated S-adenosylmethionine as the co-factor, were isolated from Nicotiana sylvestris, Hyoscyamus niger, and Arabidopsis thaliana. When the N. sylvestris SPDS cDNA was expressed in a SPDS-deficient E. coli mutant, the recombinant protein showed high SPDS activity, but did not have any spermine synthase activity. The plant SPDSs have molecular masses of about 34 kDa, possess the co-factor binding motifs which have been proposed for S-adenosylmethionine, and are more homologous in amino acid sequence to tobacco putrescine N-methyltransferase (PMT) than to SPDSs from mammals and E. coli. The SPDS gene is expressed in root, stem, and leaf in N. sylvestris, whereas the PMT gene is expressed only in root. The potential evolution of plant SPDS and PMT, and their evolutionary relationships with animal SPDS are discussed. [ABSTRACT FROM AUTHOR]