The cumulative incidence of and risk factors for morphometric severe vertebral fractures in Japanese men and women: the ROAD study third and fourth surveys.

This population-based cohort study with a 3-year follow-up revealed that the annual incidence rates of vertebral fracture (VF) and severe VF (sVF) were 5.9%/year and 1.7%/year, respectively. The presence of mild VF at the baseline was a significant risk factor for incident sVF in participants without prevalent sVF. INTRODUCTION: This study aimed to estimate the incidence of morphometric vertebral fracture (VF) and severe VF (sVF) in men and women and clarify whether the presence of a mild VF (mVF) increases the risk of incident sVF. METHODS: Data from the population-based cohort study, entitled the Research on Osteoarthritis/Osteoporosis Against Disability (ROAD) study, were analyzed. In total, 1190 participants aged ≥ 40 years (mean age, 65.0 ± 11.2) years completed whole-spine lateral radiography both at the third (2012-2013, baseline) and fourth surveys performed 3 years later (2015-2016, follow-up). VF was defined using Genant's semi-quantitative (SQ) method: VF as SQ ≥ 1, mVF as SQ = 1, and sVF as SQ ≥ 2. Cumulative incidence of VF and sVF was estimated. Multivariate logistic regression analyses were performed to evaluate risk factors for incident sVF. RESULTS: The baseline prevalence of mVF and sVF were 16.8% and 6.0%, respectively. The annual incidence rates of VF and sVF were 5.9%/year and 1.7%/year, respectively. The annual incidence rates of sVF in participants without prevalent VF, with prevalent mVF, and with prevalent sVF were 0.6%/year, 3.8%/year, and 11.7%/year (p < 0.001), respectively. Multivariate logistic regression analyses in participants without prevalent sVF showed that the adjusted odds ratios for incident sVF were 4.12 [95% confident interval 1.85-9.16] and 4.53 [1.49-13.77] if the number of prevalent mVF at the baseline was 1 and ≥ 2, respectively. CONCLUSIONS: The annual incidence rates of VF and sVF were 5.9%/year and 1.7%/year, respectively. The presence of prevalent mVF was an independent risk factor for incident sVF.

Bone strength of the proximal femur in healthy subjects with ossification of the posterior longitudinal ligament.

We compared the bone strength measured via quantitative computed tomography-based finite element method (QCT/FEM) between healthy adults with and without ossification of the posterior longitudinal ligament (OPLL). No statistically significant difference was observed in the bone strength between healthy adults with and without OPLL. Hyperostosis of the posterior longitudinal ligament in OPLL may not be associated with the systemic bone strength. INTRODUCTION: Although patients with OPLL have been reportedly associated with increased level of bone mineral density (BMD) using dual-energy X-ray absorptiometry (DXA), little is known about the bone strength in OPLL subjects. The aim of this study is to investigate the bone strength measured via QCT/FEM in healthy subjects with OPLL using the medical check-up data, including whole-body CT scans. METHODS: We examined 796 participants (529 men and 267 women) who underwent CT scans in a single health center between January 2008 and May 2009. We identified OPLL in whole spine and divided the subjects into two groups: non-OPLL and OPLL groups. We calculated the predicted bone strength (PBS) of the proximal femur using QCT/FEM and examined the bone mineral status of the calcaneus using quantitative ultrasound (QUS). We compared the PBS and the QUS parameters between the non-OPLL and OPLL groups. RESULTS: Seventy-four subjects (9.3%; 57 men and 17 women) were diagnosed with OPLL in the whole spine. The OPLL group was significantly older than the non-OPLL group. No statistically significant difference was observed in the PBS and the QUS parameters between the non-OPLL and OPLL groups in both sexes. Furthermore, no statistically significant difference was noted in the PBS and the QUS parameters between two groups in age- and gender-matched analysis. CONCLUSIONS: Our results suggest that hyperostosis of the posterior longitudinal ligament in OPLL may not be associated with bone strength and bone mineral status at the extremities.

Phonon Lifetime Observation in Epitaxial ScN Film with Inelastic X-Ray Scattering Spectroscopy.

Phonon-phonon scattering dominates the thermal properties in nonmetallic materials, and it directly influences device performance in applications. The understanding of the scattering has been progressing using computational approaches, and the direct and systematic observation of phonon modes that include momentum dependences is desirable. We report experimental data on the phonon dispersion curves and lifetimes in an epitaxially grown ScN film using inelastic x-ray scattering measurements. The momentum dependence of the optical phonon lifetimes is estimated from the spectral width, and the highest-energy phonon mode around the zone center is found to possess a short lifetime of 0.21 ps. A comparison with first-principles calculations shows that our observed phonon lifetimes are quantitatively explained by three-body phonon-phonon interactions.

Effect of brefelamide on proliferation of 1321N1 human astrocytoma cells induced by glial cell line-derived neurotrophic factor.

Malignant gliomas are highly resistant to chemotherapy and radiation and more effective options for treatment are urgently needed. We reported previously that the aromatic amide brefelamide, which is isolated from methanolic extracts of the cellular slime molds Dictyostelium giganteum and D. brefeldianum, hinders cellular proliferation in a glioma model utilizing 1321N1 human astrocytoma cells. Herein, we examined the mechanisms underlying the inhibition of 1321N1 cell proliferation by brefelamide. Glial cell line-derived neurotrophic factor (GDNF) was found to enhance the rate of proliferation of serum-free cultured 1321N1 cells, but did not affect proliferation in PC12 cells. Brefelamide pretreatment inhibited GDNF-induced cell proliferation and expression of rearranged during transfection (RET). GDNF enhanced the phosphorylation of extracellular signal-regulated kinase (ERK), AKT, and c-jun-N-terminal kinase (JNK); however, brefelamide pretreatment inhibited these effects. Brefelamide also reduced the expression of GDNF mRNA and GDNF secretion. Together, the findings from this study indicate that brefelamide inhibits the proliferation of 1321N1 cell via several mechanisms including reduced GDNF receptor expression and GDNF secretion, and reduced phosphorylation of ERK, AKT, and JNK.

Inhibition of choroidal fibrovascular membrane formation by new class of RNA interference therapeutic agent targeting periostin.

Age-related macular degeneration (AMD) is a vision-threatening disease characterized by choroidal fibrovascular membrane (FVM) formation, choroidal neovascularization (CNV) and choroidal fibrosis. No safe and effective therapeutic method has been developed for the choroidal fibrosis, although anti-vascular endothelial growth factor therapy can partially shrink the CNV. We recently reported that periostin (POSTN), which is produced by retinal pigment epithelial cells, has an important role in the formation of preretinal FVMs, but its role in choroidal FVMs has not been determined. In this study, we used Postn knockout mice to investigate the role played by POSTN in choroidal FVM formation. In addition, we used a new class of RNA interference (RNAi) agent (NK0144) that targets POSTN and determined its effect on choroidal FVM development. Genetic ablation of Postn had an inhibitory effect not only on CNV formation but also on choroidal fibrosis in a mouse CNV model. NK0144 also had a greater inhibitory effect on both the CNV and choroidal fibrosis than control RNAi with no apparent adverse effects. These findings suggest a causal relationship between POSTN and choroidal FVM formation, and also a potential therapeutic role of intravitreal NK0144 for AMD.

Characterization and evolutionary analysis of tributyltin-binding protein and pufferfish saxitoxin and tetrodotoxin-binding protein genes in toxic and nontoxic pufferfishes.

Understanding the evolutionary mechanisms of toxin accumulation in pufferfishes has been long-standing problem in toxicology and evolutionary biology. Pufferfish saxitoxin and tetrodotoxin-binding protein (PSTBP) is involved in the transport and accumulation of tetrodotoxin and is one of the most intriguing proteins related to the toxicity of pufferfishes. PSTBPs are fusion proteins consisting of two tandem repeated tributyltin-binding protein type 2 (TBT-bp2) domains. In this study, we examined the evolutionary dynamics of TBT-bp2 and PSTBP genes to understand the evolution of toxin accumulation in pufferfishes. Database searches and/or PCR-based cDNA cloning in nine pufferfish species (6 toxic and 3 nontoxic) revealed that all species possessed one or more TBT-bp2 genes, but PSTBP genes were found only in 5 toxic species belonging to genus Takifugu. These toxic Takifugu species possessed two or three copies of PSTBP genes. Phylogenetic analysis of TBT-bp2 and PSTBP genes suggested that PSTBPs evolved in the common ancestor of Takifugu species by repeated duplications and fusions of TBT-bp2 genes. In addition, a detailed comparison of Takifugu TBT-bp2 and PSTBP gene sequences detected a signature of positive selection under the pressure of gene conversion. The complicated evolutionary dynamics of TBT-bp2 and PSTBP genes may reflect the diversity of toxicity in pufferfishes.

A-type antiferro-orbital ordering with I4(1)/a symmetry and geometrical frustration in the spinel vanadate MgV2O4.

We conduct a detailed structural analysis of the S=1 pyrochlore antiferromagnet MgV2O4, which exhibits an antiferromagnetic ordering marginally at TN=40 K, triggered by a structural transition from cubic to tetragonal symmetry at TS=62 K, using high resolution synchrotron x-ray diffraction and convergent beam electron diffraction. We reveal that the tetragonal phase below TS has the symmetry of I4(1)/a and that the distortion pattern of VO6 octahedra is consistent with A-type antiferro-orbital ordering with alternating stacking of layers with yz/xy orbital chains and zx/xy orbital chains along the tetragonal c axis. This implies that an anisotropic coupling of V moments produced by the orbital ordering below TS primarily brings about the antiferromagnetic ordering.

Correlation-enhanced effective mass of two-dimensional electrons in Mg(x)Zn(1-x)O/ZnO heterostructures.

We performed combined magnetotransport and cyclotron resonance experiments on two-dimensional electron systems confined in the Mg(x)Zn(1-x)O/ZnO heterostructures over a wide range of carrier densities, from 1.9 to 12 × 10(11) cm(-2) (3.5

Enhanced removal of sodium salts supported by in-situ catalyst synthesis in a supercritical water oxidation process.

For practical applications of supercritical water oxidation to wastewater treatment, the deposition of inorganic salts in supercritical phase must be controlled to prevent a reactor from clogging. This study investigated enhanced removal of sodium salts with titanium particles, serving as a salt trapper and a catalyst precursor, and sodium recovery by sub-critical water. When Na(2)CO(3) was tested as a model salt, sodium removal efficiency was higher than theoretically maximum efficiency defined by Na(2)CO(3) solubility. The enhanced sodium removal resulted from in-situ synthesis of sodium titanate, which could catalyse acetic acid oxidation. The kinetics of sodium removal was described well by a diffusion mass-transfer model combined with a power law-type rate model of sodium titanate synthesis. Titanium particles showed positive effect on sodium removal in the case of NaOH, Na(2)SO(4) and Na(3)PO(4). However, they had negligible effect for NaCl and negative effect for Na(2)CrO(4), respectively. More than 99% of trapped sodium was recovered by sub-critical water except for Na(2)CrO(4). In contrast, sodium recovery efficiency remained less than 50% in the case of Na(2)CrO(4). Reused titanium particles showed the same performance for enhanced sodium removal. Enhanced salt removal supported by in-situ catalyst synthesis has great potential to enable both salt removal control and catalytic oxidation.

Electron microscopy at a sub-50 pm resolution.

An aberration-corrected electron microscope developed in CREST project has been applied for imaging atoms and clusters buried inside crystals. The resolution of the microscope in scanning transmission electron microscopy (STEM) has experimentally proved to be better than 47 pm by use of a cold-field emission gun at 300 kV. The high resolution has given an advantage for imaging light elements such as lithium atoms discriminating one by one. Moreover, a three-dimensional structure imaging has been demonstrated for dopant clusters by a sub-50 pm STEM, using its high depth resolution.

Myoglobin subfractions: abnormality in duchenne type of progressive muscular dystrophy.

Human metmyoglobin was separated electrophoretically into four subfractions: Mb(1), Mb(2), Mb(3), and Mb(4), which divide into at least two biochemically independent groups: Mb(1) and Mb(2), and Mb(3), and Mb(4). In normal subjects, Mb(1) constituted the predominant component; Mb(2), Mb(3), and Mb(4) were the minor components in this descending order. In the Duchenne type of progressive muscular dystrophy, on the contrary, a remarkable decrease in Mb(1) and a concomitant increase in Mb(3) were observed. This unique abnormality in the relative distribution of myoglobin subfractions was recognized only in the Duchenne type and not in other types of progressive muscular dystrophy or in other myopathies.

Measurement method of aberration from Ronchigram by autocorrelation function.

Aberrations up to the fifth-order were successfully measured using an autocorrelation function of the segmental areas of a Ronchigram. The method applied to aberration measurement in a newly developed 300kV microscope that is equipped with a spherical aberration corrector for probe-forming systems. The experimental Ronchigram agreed well with the simulated Ronchigram that was calculated by using the measured aberrations. The Ronchigram had an infinite magnification area with a half-angle of 50mrad, corresponding to the convergence angle of a uniform phase.

[Surgical treatment of transposition of great arteries: the coronary buttons transfer using medially based trapdoor flaps].

The arterial switch operation (ASO) has become the primary surgical approach used for correction of transposition of the great arteries. All the prerequisites for a successful ASO were recognized in time and dealt with, which allowed general acceptation of the technique. We report on our technique for the procedure and the result to date. From January 1991 to January 2008, a total of 100 patients underwent ASO at our unit using medially-based trapdoor flap method. The neo-pulmonary artery (PA) was reconstructed using a single rectangular pericardial patch. The initial patient having intramural coronary artery died due to ischemic event after Aubert procedure. Three patients had re-right ventricular out flow tract repair (RVOTR) in a long-term follow-up period. There was no significant aortic insufficiency, no ischemic event and no lethal arrhythmia.

Specific cis-acting sequence for PHO8 expression interacts with PHO4 protein, a positive regulatory factor, in Saccharomyces cerevisiae.

The PHO8 gene of Saccharomyces cerevisiae encodes repressible alkaline phosphatase (rALPase; EC 3.1.3.1). The rALPase activity of the cells is two to three times higher in medium containing a low concentration of Pi than in high-Pi medium due to transcription of PHO8. The Pi signals are conveyed to PHO8 by binding of PHO4 protein, a positive regulatory factor, to a promoter region of PHO8 (PHO8p) under the influence of the PHO regulatory circuit. Deletion analysis of PHO8p DNA revealed two separate regulatory regions required for derepression of rALPase located at nucleotide positions -704 to -661 (distal region) and -548 to -502 (proximal region) and an inhibitory region located at -421 to -289 relative to the translation initiation codon. Gel retardation experiments showed that a beta-galactosidase-PHO4 fusion protein binds to a 132-bp PHO8p fragment bearing the proximal region but not to a 226-bp PHO8 DNA bearing the distal region. The fusion protein also binds to a synthetic oligonucleotide having the same 12-bp nucleotide sequence as the PHO8p DNA from positions -536 to -525. The 132-bp PHO8p fragment, connected at position -281 of the 5' upstream region of a HIS5'-'lacZ fused gene, could sense Pi signals in vivo, but a 20-bp synthetic oligonucleotide having the same sequence from -544 to -525 of the PHO8p DNA could not. Linker insertions in the PHO8p DNA indicated that the 5-bp sequence 5'-CACGT-3' from positions -535 to -531 is essential for binding the beta-galactosidase-PHO4 fusion protein and for derepression of rALPase.

Functional domains of a positive regulatory protein, PHO4, for transcriptional control of the phosphatase regulon in Saccharomyces cerevisiae.

The PHO4 gene encodes a positive regulatory factor involved in regulating transcription of various genes in the phosphatase regulon of Saccharomyces cerevisiae. Besides its own coding region, the 1.8-kilobase PHO4 transcript contains a coding region for a mitochondrial protein which does not appear to be translated. Four functional domains were found in the PHO4 protein, which consists of 312 amino acid (aa) residues as deduced from the open reading frame of PHO4. A gel retardation assay with beta-galactosidase::PHO4 fused protein revealed that the 85-aa C terminus is the domain responsible for binding to the promoter DNA of PHO5, a gene under the control of PHO4. This region has similarities with the amphipathic helix-loop-helix motif of c-myc protein. Determination of the nucleotide sequences of four PHO4c mutant alleles and insertion and deletion analyses of PHO4 DNA indicated that a region from aa 163 to 202 is involved in interaction with a negative regulatory factor PHO80. Complementation of a pho4 null allele with the modified PHO4 DNAs suggested that the N-terminal region (1 to 109 aa), which is rich in acidic aa, is the transcriptional activation domain. The deleterious effects of various PHO4 mutations on the constitutive transcription of PHO5 in PHO4c mutant cells suggested that the region from aa 203 to 227 is involved in oligomerization of the PHO4 protein.

Function of the ste signal transduction pathway for mating pheromones sustains MAT alpha 1 transcription in Saccharomyces cerevisiae.

Sterile mutants of Saccharomyces cerevisiae were isolated from alpha * cells having the a/alpha aar1-6 genotype (exhibiting alpha mating ability and weak a mating ability as a result of a defect in a1-alpha 2 repression). Among these sterile mutants, we found two ste5 mutants together with putative ste7, ste11, and ste12 mutants of the signal transduction pathway of mating pheromones. The amino acid sequence of the Ste5p protein predicted from the nucleotide sequence of a cloned STE5 DNA has a domain rich in acidic amino acids close to its C terminus, a cysteine-rich sequence, resembling part of a zinc finger structure, in its N-terminal half, and a possible target site of cyclic AMP-dependent protein kinase at its C terminus. Northern (RNA) blot analysis revealed that STE5 transcription is under a1-alpha 2-Aar1p repression. The MAT alpha 1 cistron has a single copy of the pheromone response element in its 5' upstream region, and its basal level of transcription was reduced in these ste mutant cells. However, expression of the MAT alpha 1 cistron was not enhanced appreciably by pheromone signals. One of the ste5 mutant alleles conferred a sterile phenotype to a/alpha aar1-6 cells but a mating ability to MATa cells.

AAR2, a gene for splicing pre-mRNA of the MATa1 cistron in cell type control of Saccharomyces cerevisiae.

We have isolated a class of mutants, aar2, showing the alpha mating type due to a defect in a1-alpha 2 repression but with alpha 2 repression activity from a nonmater strain of Saccharomyces cerevisiae expressing both a and alpha mating-type information in duplicate. Cells of the aar2 mutant and the aar2 disruptant also show a growth defect. A DNA fragment complementing the aar2 mutation contains an open reading frame consisting of 355 amino acid codons. Northern hybridization showed that cells of the aar2 mutant and disruptant contained alpha 1 and alpha 2 transcripts of the MAT alpha gene (or HML alpha in sir3 cells), but their a1 transcript of MATa (or HMRa in sir3 cells) migrated more slowly than that of the wild-type cells on gel electrophoresis and gave a diffused band. Primer extension analysis showed that the aar2 mutant and disruptant have a defect in splicing two short introns of the a1 pre-mRNA but not in splicing pre-mRNA of ACT1. The alpha mating type, but not the slow-growing phenotype, of the aar2 mutant was suppressed by introduction of an intronless MATa1 DNA. Thus, the AAR2 gene is involved in splicing pre-mRNA of the a1 cistron and other genes that are important for cell growth. The AAR2 locus was mapped on chromosome II beside the SSA3 locus, with a 276-bp space, but was not allelic to either PRP5 or PRP6, which are both located on chromosome II and function in splicing pre-mRNA of ACT1.

AAR1/TUP1 protein, with a structure similar to that of the beta subunit of G proteins, is required for a1-alpha 2 and alpha 2 repression in cell type control of Saccharomyces cerevisiae.

We have cloned a DNA fragment complementing the aar1 mutation defective in the a1-alpha 2 repression of the alpha 1 cistron and haploid-specific genes in Saccharomyces cerevisiae. Nucleotide sequence and mapping data indicated that the AAR1 gene is identical with TUP1, which is allelic to the SFL2, FLK1, CYC9, UMR7, AMM1, and AER2 genes, whose mutations are known to confer a variety of phenotypes, such as thymidine uptake, flocculation, insensitivity to glucose repression, a defect in UV-induced mutagenesis, and a defect in ARS plasmid maintenance. The TUP1/AER2 protein is known to have significant similarity with the beta subunits of G proteins in the C-terminal half, in two glutamine-rich domains in the N-terminal half, and in a central region rich in serine and threonine residues. Disruption of the chromosomal AAR1 gene in alpha and a/alpha cells conferred the nonmating phenotype, and the a/alpha diploids could not sporulate. The AAR1/TUP1 gene is transcribed into a 2.5-kb mRNA independently of the mating-type information of the cell. These observations and mRNA analysis of cell-type-specific genes indicated that the AAR1/TUP1 protein is also indispensable for a1-alpha 2 repression of RME1 and for alpha 2 repression of a-specific genes.

Gene conversion associated with site-specific recombination in yeast plasmid pSR1.

A circular DNA plasmid, pSR1, isolated from Zygosaccharomyces rouxii has a pair of inverted repeats consisting of completely homologous 959-base pair (bp) sequences. Intramolecular recombination occurs frequently at the inverted repeats in cells of Saccharomyces cerevisiae, as well as in Z. rouxii, and is catalyzed by a protein encoded by the R gene of its own genome. The recombination is, however, independent of the RAD52 gene of the host genome. A site for initiation of the intramolecular recombination in the S. cerevisiae host was delimited into, at most, a 58-bp region in the inverted repeats by using mutant plasmids created by linker insertion. The 58-bp region contains a pair with 14-bp dyad symmetry separated by a 3-bp spacer sequence. The recombination initiated at this site was accompanied by a high frequency of gene conversion (3 to 50% of the plasmid clones examined). Heterogeneity created by the linker insertion or by a deletion (at most 153 bp so far tested) at any place on the inverted repeats was converted to a homologous combination by the gene conversion, even in the rad52-1 mutant host. A mechanism implying branch migration coupled with DNA replication is discussed.

Putative GTP-binding protein, Gtr1, associated with the function of the Pho84 inorganic phosphate transporter in Saccharomyces cerevisiae.

We have found an open reading frame which is 1.1 kb upstream of PHO84 (which encodes a Pi transporter) and is transcribed from the opposite strand. In Saccharomyces cerevisiae, this gene is distal to the TUB3 locus on the left arm of chromosome XIII and is named GTR1. GTR1 encodes a protein consisting of 310 amino acid residues containing, in its N-terminal region, the characteristic tripartite consensus elements for binding GTP conserved in GTP-binding proteins, except for histidine in place of a widely conserved aspargine residue in element III. Disruption of the GTR1 gene resulted in slow growth at 30 degrees C and no growth at 15 degrees C; other phenotypes resembled those of pho84 mutants and included constitutive synthesis of repressible acid phosphatase, reduced Pi transport activity, and resistance to arsenate. The latter phenotypes were shown to be due to a defect in Pi uptake, and the Gtr1 protein was found to be functionally associated with the Pho84 Pi transporter. Recombination between chromosome V (at the URA3 locus) and chromosome XIII (in the GTR1-PHO84-TUB3 region) by using a plasmid-encoded site-specific recombination system indicated that the order of these genes was telomere-TUB3-PHO84-GTR1-CENXIII.