DNA strand breaks in mammalian tissues induced by methylarsenics

DNA damage induced by administration of dimethylarsinic acid (DMAA) to rats and mice was investigated. At 12 h after administration of DMAA, DNA single-strand breaks were induced markedly in lung. The majority of dimethylarsine, one of the main metabolites, in the expired air was excreted within 6–18 h after administration of DMAA to rats. In vitro experiments using nuclei isolated from lung of mice indicated that DNA strand breaks were caused by dimethylarsine. Furthermore, the strand breaks after exposure to dimethylarsine were reduced in the presence of catalase and/or superoxide dismutase. These results strongly suggest that the strand breaks are induced not by dimethylarsine itself but by active oxygen, e.g., O ₂ ^? and ·OH, produced both by dimethylarsine and molecular oxygen. When DNA was exposed to dimethylarsine, thiobarbituric acid (TBA)-reactive intermediates andcis-thymine glycol were produced. Dimethylarsine appears to induce DNA damage by the mechanism similar to the damage produced by ionizing radiation.     

Isolation of an actin polymerization stimulator from bovine thyroid plasma membranes

An actin polymerization stimulator was purified from bovine thyroid plasma membranes by DNase I affinity column chromatography. Although the molecular weight of the protein was about 42,000 (42K) by sodium dodecyl sulfate polyacrylamide gel electrophoresis, it did not comigrate with actin. In the presence of 30 mM KCl, the 42K protein facilitated formation of actin filaments when analyzed by a centrifugation method, accelerated the initial phase of actin polymerization as measured in an Ostwald viscometer and increased the length of filaments as shown by electron microscopy. The 42K protein also accelerated the initial phase of actin polymerization in the presence of 100 mM KCl and 2 mM MgCl₂ but did not affect the final viscosity. The effect of the 42K protein was diminished by 5 uM cytochalasin B or 1 uM cytochalasin D. This 42K protein may anchor actin filaments onto the thyroid plasma membrane.     

Localization of the ribosome-releasing factor gene in the Escherichia coli chromosome.

The ribosome-releasing factor (RRF) gene was localized at a position between 2 and 6 min on the Escherichia coli chromosome by measuring the gene-dosage-dependent production of RRF in various E. coli F' merozygotes. This position was confirmed and refined by using a nucleotide probe corresponding to a 16-amino-acid sequence in RRF. It was found that the RRF gene was contained in pLC 6-32 of the Clark-Carbon Gene Bank. Restriction enzyme mapping of E. coli genomic DNA with the above probe led us to conclude that the RRF gene is situated in the 4-min region, somewhere downstream (clockwise) of the elongation factor Ts gene, tsf. A pLC 6-32-derived DNA fragment which carries the RRF gene was found to contain a partial sequence of tsf. The exact location of the translational initiation site of the RRF gene was determined to be 1.1 kilobases downstream from the translational termination site of tsf. The RRF gene is designated frr.     

Simulation analysis of excitation conduction in the heart: Propagation of excitation in different tissues

The normal excitation and conduction in the heart are maintained by the coordination between the dynamics of ionic conductance of each cell and the electrical coupling between cells. To examine functional roles of these two factors, we proposed a spatially-discrete model of conduction of excitation in which the individual cells were assumed isopotential. This approximation was reasoned by comparing the apparent space constant with the measured junctional resistance between myocardial cells. We used the four reconstruction models previously reported for five kinds of myocardial cells. Coupling coefficients between adjacent cells were determined quantitatively from the apparent space constants. We first investigated to what extent the pacemaker activity of the sinoatrial node depends on the number and the coupling coefficient of its cells, by using a one-dimensional model system composed of the sinoatrial node cells and the atrial cells. Extensive computer simulation revealed the following two conditions for the pacemaker activity of the sinoatrial node. The number of the sinoatrial node cells and their coupling coefficients must be large enough to provide the atrium with the sufficient electric current flow. The number of the sinoatrial node cells must be large so that the period of the compound system is close to the intrinsic period of the sinoatrial node cell. In this simulation the same sinoatrial node cells produced action potentials of different shapes depending on where they were located in the sinoatrial node. Therefore it seems premature to classify the myocardial cells only from their waveforms obtained by electrical recordings in the compound tissue. Second, we investigated the very slow conduction in the atrioventricular node compared to, for example, the ventricle. This was assumed to be due to the inherent property of the membrane dynamics of the atrioventricular node cell, or to the small value of the coupling coefficient (weak intercellular coupling), or to the electrical load imposed on the atrioventricular node by the Purkinje fibers, because the relatively small atrioventricular node must provide the Purkinje fibers with sufficient electric current flow. Relative contributions of these three factors to the slow conduction were evaluated using the model system composed of only the atrioventricular cells or that composed of the atrioventricular and Purkinje cells. We found that the weak coupling has the strongest effect. In the model system composed of the atrioventricular cells, the propagation failure was not observed even for very small values of the coupling coefficient.(ABSTRACT TRUNCATED AT 400 WORDS)     

Mapping quantitative trait loci for root development under hypoxia conditions in soybean (<Emphasis Type="Italic">Glycine max</Emphasis> L. Merr.)

Key message

Greatest potential, QTLs for hypoxia and waterlogging tolerance in soybean roots were detected using a new phenotypic evaluation method.

Abstract

Waterlogging is a major environmental stress limiting soybean yield in wet parts of the world. Root development is an important indicator of hypoxia tolerance in soybean. However, little is known about the genetic control of root development under hypoxia. This study was conducted to identify quantitative trait loci (QTLs) responsible for root development under hypoxia. Recombinant inbred lines (RILs) developed from a cross between a hypoxia-sensitive cultivar, Tachinagaha, and a tolerant landrace, Iyodaizu, were used. Seedlings were subjected to hypoxia, and root development was evaluated with the value change in root traits between after and before treatments. We found 230 polymorphic markers spanning 2519.2 cM distributed on all 20 chromosomes (Chrs.). Using these, we found 11 QTLs for root length (RL), root length development (RLD), root surface area (RSA), root surface area development (RSAD), root diameter (RD), and change in average root diameter (CARD) on Chrs. 11, 12, 13 and 14, and 7 QTLs for hypoxia tolerance of these root traits. These included QTLs for RLD and RSAD between markers Satt052 and Satt302 on Chr. 12, which are important markers of hypoxia tolerance in soybean; those QTLs were stable between 2 years. To validate the QTLs, we developed a near-isogenic line with the QTL region derived from Iyodaizu. The line performed well under both hypoxia and waterlogging, suggesting that the region contains one or more genes with large effects on root development. These findings may be useful for fine mapping and positional cloning of gene responsible for root development under hypoxia.

     

Plasma anti-Müllerian hormone as a biomarker for bovine granulosa-theca cell tumors: Comparison with immunoreactive inhibin and ovarian steroid concentrations

Granulosa-theca cell tumors (GTCTs) are the most frequently reported ovarian tumors in cattle. Clinically, GTCTs could be confused with other ovarian abnormalities; therefore, the only definitive diagnosis for such tumors is histopathology of a biopsy from the affected ovary. However, this is an invasive technique and unsuitable for farm conditions. As a result, the key aim of this study was to evaluate the diagnostic value of anti-Müllerian hormone (AMH), a glycoprotein hormone that is synthesized exclusively by ovarian granulosa cells, as a sensitive noninvasive biomarker for diagnosing GTCTs in cattle. To achieve this aim, we conducted two experiments. In experiment 1, four clinically healthy Japanese Black cows had blood samples taken daily over one estrous cycle to characterize their AMH profiles throughout the estrous cycle. Additionally, single blood samples were collected from 21 cyclic cows to clarify the physiological range of AMH. In experiment 2, cows with histologically confirmed GTCT (GTCT group, n = 9) and cows affected with cystic ovarian disease (COD group, n = 8) had one blood sample taken before extraction of the tumorous ovary or therapeutic treatment for the COD. Blood samples (n = 105) from cyclic cows (n = 25) in experiment 1 were assigned as a physiologically cyclic group (PC group). Plasma AMH, immunoreactive inhibin (ir-INH), estradiol-17β (E₂), testosterone (T), and progesterone (P₄) concentrations were assayed in all samples. In experiment 1, the mean plasma AMH concentration was 0.09 ± 0.003 ng/mL and did not show substantial fluctuation throughout the estrous cycle. In experiment 2, plasma AMH, ir-INH, and E₂ concentrations were significantly elevated in the GTCT group in comparison with the PC group; among these parameters, only the AMH concentrations were significantly higher in the GTCT group than in the COD group. The area under the receiver operating characteristic curve of AMH for diagnosis of GTCT was 0.99 and was significantly higher than that of ir-INH (P < 0.001) and E₂ (P < 0.01). Moreover, the AMH at a cutoff point of ≥0.36 ng/mL had the highest diagnostic accuracy (99.2%), sensitivity (100%), and specificity (99.1%) compared with the other tested parameters. In conclusion, plasma AMH concentration is probably a more reliable and sensitive biomarker for bovine GTCTs than the concentrations of ir-INH or ovarian steroids.     

A Convenient Method for the Quantitative Determination of Pipecolic Acid

The quantitative determination of pipecolic acid was examined.

The reaction of 3% ninhydrin solution in n-butanol, saturated with citrate buffer (pH 4.2), with pipecolic acid in boiling water for 3 min yielded the colored products showing λ_max at 570 mμ, but with proline hardly yielded those products. By the colorimetry proposed, it is possible to determine the amount of pipecolic acid in the sample containing proline no more than 50 times the amount of the pipecolic acid, directly from the calibration curve using pipecolic acid.

The method for removal of amino acids from the sample containing pipecolic acid and proline was examined and discussed.     

Inhibition of lipid peroxidation by a novel compound (CV-2619) in brain mitochondria and mode of action of the inhibition

Lipid peroxidation in rat brain mitochondria was induced by NADH in the presence of ADP and FeCl3. CV-2619 inhibited the lipid peroxidation in a concentration-dependent manner; the concentration giving 50% inhibition (IC50) was 84 microM. In addition, the inhibitory effect of CV-2619 was strongly enhanced by adding substrates of mitochondrial respiration; when succinate, glutamate, or succinate plus glutamate was added, the IC50 of CV-2619 was changed to 1.1, 10, or 0.5 microM, respectively. Metabolites of CV-2619 also inhibited the lipid peroxidation. The inhibitory effect of CV-2619 on mitochondrial lipid peroxidation disappeared when TTFA, an inhibitor of complex II in mitochondrial respiratory chain, was added. The results indicate that in mitochondria CV-2619 is changed to its reduced form which inhibits lipid peroxidation.     

Formaldehyde Fixation Contributes to Detoxification for Growth of a Nonmethylotroph, Burkholderia cepacia TM1, on Vanillic Acid

During bacterial degradation of methoxylated lignin monomers, such as vanillin and vanillic acid, formaldehyde is released through the reaction catalyzed by vanillic acid demethylase. When Burkholderia cepacia TM1 was grown on vanillin or vanillic acid as the sole carbon source, the enzymes 3-hexulose-6-phosphate synthase (HPS) and 6-phospho-3-hexuloisomerase (PHI) were induced. These enzymes were also expressed during growth on Luria-Bertani medium containing formaldehyde. To understand the roles of these enzymes, the hps and phi genes from a methylotrophic bacterium, Methylomonas aminofaciens 77a, were introduced into B. cepacia TM1. The transformant strain constitutively expressed the genes for HPS and PHI, and these activities were two- or threefold higher than the activities in the wild strain. Incorporation of [¹⁴C]formaldehyde into the cell constituents was increased by overexpression of the genes. Furthermore, the degradation of vanillic acid and the growth yield were significantly improved at a high concentration of vanillic acid (60 mM) in the transformant strain. These results suggest that HPS and PHI play significant roles in the detoxification and assimilation of formaldehyde. This is the first report that enhancement of the HPS/PHI pathway could improve the degradation of vanillic acid in nonmethylotrophic bacteria.