Mn and Cu concentrations in mixed saliva of elementary school children in relation to sex,age, and dental caries

To examine the standard Mn and Cu concentrations in mixed saliva from children and the relationship between these levels and dental caries, resting mixed saliva samples obtained from 527 children of an elementary school in Kitakyushu City were collected at 10:00–11:30 a.m. during December 2004. The Mn and Cu concentrations were determined using simultaneous multi-element atomic absorption spectrometry. The standard Mn and Cu levels were 22.0±15.2 and 3.8±4.1 ng/mL, respectively, in the sound teeth group. Mn levels were significantly higher in boys (25.4±17.4 ng/mL) than girls (19.1±12.3 ng/mL) and also higher in upper (25.5±16.4 ng/mL) than lower (19.0±13.5 ng/mL) grades. The Cu level was unaffected by sex and age in the sound teeth group. The Cu level in children with caries experience (5.7±5.3 ng/mL) was significantly higher than that of the sound teeth group. Moreover, the Cu levels in children with untreated caries were significantly higher than that of the sound teeth group, and increased with the number of untreated teeth. No significant difference was found in the Cu concentrations between the group in which all decayed teeth were treated and the sound teeth group. The Mn levels were similar with or without caries and treatment. These findings indicate that the Mn level in mixed saliva depended on sex and age, and suggest the possibility of Cu dissolving into mixed saliva by demineralization due to dental caries.     

Are the effects of α-glucosidase inhibitors on cardiovascular events related to elevated levels of hydrogen gas in the gastrointestinal tract?

The major side-effect of treatment with α-glucosidase inhibitors, flatulence, occurs when undigested carbohydrates are fermented by colonic bacteria, resulting in gas formation. We propose that the cardiovascular benefits of α-glucosidase inhibitors are partly attributable to their ability to neutralise oxidative stress via increased production of H₂ in the gastrointestinal tract. Acarbose, which is an α-glucosidase inhibitor, markedly increased H₂ production, with a weaker effect on methane production. Our hypothesis is based on our recent discovery that H₂ acts as a unique antioxidant, and that when inhaled or taken orally as H₂-dissolved water it ameliorates ischaemia-reperfusion injury and atherosclerosis development.     

Malonylation is a key reaction in the metabolism of xenobiotic phenolic glucosides in Arabidopsis and tobacco

Tobacco cells (Nicotiana tabacum L.) accumulate harmful naphthols in the form of malonylated glucosides ( Taguchi et al., 2005 ). Here, we showed that the malonylation of glucosides is a system to metabolize xenobiotics and is common to higher plants. Moreover, some plantlets including Arabidopsis thaliana excreted some of the incorporated naphthols into the culture media as their glucosides. In order to analyze the function of malonylation in the metabolism of these xenobiotics, we identified a malonyltransferase gene (At5g39050) responsible for the malonylation of these compounds in A. thaliana. The recombinant enzyme had malonyltransferase activity toward several phenolic glucosides including naphthol glucosides. A knockout mutant of At5g39050 (pmat1) exposed to naphthols accumulated only a few malonylglucosides in the cell, and released larger amounts of simple glucosides into the culture medium. In contrast, forced expression of At5g39050 in the pmat1 mutant resulted in increased malonylglucoside accumulation and decreased glucoside excretion to the media. The results provided clear evidence of whether the release of glucosides or the storage of malonylglucosides was determined by the At5g39050 expression level. A similar event in naphthol metabolism was observed in the tobacco mutant with a suppressed malonyltransferase gene (NtMaT1). These results suggested that malonylation could be a key reaction to separate the way of xenobiotics disposition, that is, release from cell surface or storage in vacuoles.     

Identification of a human peripheral blood monocyte subset that differentiates into osteoclasts

Increased bone resorption mediated by osteoclasts causes various diseases such as osteoporosis and bone erosion in rheumatoid arthritis (RA). Osteoclasts are derived from the monocyte/macrophage lineage, but the precise origin remains unclear. In the present study, we show that the purified CD16^- human peripheral blood monocyte subset, but not the CD16⁺ monocyte subset, differentiates into osteoclast by stimulation with receptor activator of NF-κB ligand (RANKL) in combination with macrophage colony-stimulating factor (M-CSF). Integrin-β3 mRNA and the integrin-αvβ3 heterodimer were only expressed on CD16^- monocytes, when they were stimulated with RANKL + M-CSF. Downregulation of β3-subunit expression by small interfering RNA targeting β3 abrogated osteoclastogenesis from the CD16^- monocyte subset. In contrast, the CD16⁺ monocyte subset expressed larger amounts of tumor necrosis factor alpha and IL-6 than the CD16^- subset, which was further enhanced by RANKL stimulation. Examination of RA synovial tissue showed accumulation of both CD16⁺ and CD16^- macrophages. Our results suggest that peripheral blood monocytes consist of two functionally heterogeneous subsets with distinct responses to RANKL. Osteoclasts seem to originate from CD16^- monocytes, and integrin β3 is necessary for osteoclastogenesis. Blockade of accumulation and activation of CD16^- monocytes could therefore be a beneficial approach as an anti-bone resorptive therapy, especially for RA.     

Regional insertional mutagenesis of genes on Arabidopsis thaliana chromosome V using the Ac/Ds transposon in combination with a cDNA scanning method

For regional insertional mutagenesis of Arabidopsis thaliana genes, we combined a cDNA scanning method (Hayashida et al. Gene 1995; 165:155-161) and an Ac/Ds transposon designed for local mutagenesis, and evaluated this approach with two overlapping yeast artificial chromosome (YAC) clones, CIC7E11 and CIC8B11, on A. thaliana chromosome 5. We applied a previously developed novel cDNA selection method using DNA latex particles (cDNA scanning method) to the two YAC clones and constructed two sub-libraries in which cDNAs for genes on each YAC DNA were concentrated. From each sub-library we isolated cDNAs for genes on each YAC DNA, partially sequenced them, and produced expressed sequence tags (ESTs). In total, 113 non-redundant groups of cDNAs were obtained. Forty-four per cent of these EST clones were novel, and 34% had significant homology to functional proteins from various organisms. In parallel, we transposed Ds from a donor Ds-GUS-T-DNA line, Ds4391-20, already mapped to the CIC7E11/8B11 region. We obtained Ds-transposed lines and recovered their Ds-flanking genomic DNAs by thermal asymmetric interlaced (TAIL) polymerase chain reaction (PCR). Dot-blot analysis indicated that 20% of the lines contained transposed Ds in the CIC7E11/8B11 region, suggesting that this Ac/Ds transposon system is effective for regional insertional mutagenesis. To isolate Ds insertion mutants in the genes identified from the CIC7E11/8B11 region, we carried out PCR screening from 800 Ds-containing lines using Ds-specific and gene-specific primers that were designed from the 113 cDNA sequences identified by the cDNA scanning method. We found that 49 lines contain Ds insertion mutations, and that five lines contain Ds mutations in genes that are mapped to the sequenced CIC7E11/8B11 genomic DNA region. These results indicate that combining the cDNA scanning method and the Ac/Ds transposon gives a powerful tool for regional insertional mutagenesis not only in Arabidopsis but also in other plants or crops whose genomes are not sequenced.     

Molecular cloning and characterization of three cDNAs encoding putative mitogen-activated protein kinase kinases (MAPKKs) in Arabidopsis thaliana.

We isolated three Arabidopsis thaliana cDNA clones (ATMKK3, ATMKK4 and ATMKK5) encoding protein kinases with extensive homology to the mitogen-activated protein kinase kinases (MAPKKs) of various organisms in the catalytic domain. ATMKK3 shows high homology (85% identity) to NPK2, a tobacco MAPKK homologue. ATMKK4 and 5 are closely related to each other (84% identity). Phylogenetic analysis showed that the plant MAPKKs constitute at least three subgroups. The recombinant ATMKK3 and ATMKK4 were expressed as a fusion protein with glutathione S-transferase (GST) in Escherichia coli. Affinity purified GST-ATMKK3 and GST-ATMKK4 proteins contained phosphorylation activity, which shows that both the ATMKK3 and ATMKK4 genes encode functional protein kinases. Northern blot analysis revealed that the ATMKK3 gene expressed in all the organs. The levels of ATMKK4 and 5 mRNAs were relatively higher in steins and leaves than in flowers and roots. We determined the map positions of the ATMKK3, 4 and 5 genes on Arabidopsis chromosomes by RFLP mapping using P1 genomic clones.     

Mechanism of antibody-mediated protection against herpes simplex virus infection in athymic nude mice: requirement of Fc portion of antibody

Experiments were performed to investigate the resistance of the host due to antibody-mediated mechanisms to herpes simplex virus (HSV) infection. Transfer of hyperimmune anti-HSV mouse serum inhibited the development of skin lesions and prolonged the survival of lethally HSV-infected nude mice. Relatively high concentrations of antibody were required to achieve this protection. Antisera prepared in heterologous animals were also effective, while administration of anti-cowpox virus serum or interferon provided no protection. This type of protection is therefore due to specific antibody and cannot be attributed to interferon. In order to delineate the requirement for antibody in antibody-mediated protection, human gamma globulin preparations were transferred to lethally HSV-infected nude mice. Transfer of intact human gamma globulin (GG) was effective in controlling infection. S-sulfonation of GG did not diminish the protective ability. However, purified F(ab')2 did not have any protective action even when it was administered frequently to maintain serum neutralizing antibody titer. GG was effective in C5-deficient mice lethally infected with HSV. These results indicate that in vivo antibody-mediated protection to HSV infection requires the Fc region of the intact IgG molecule and suggest that antibody-dependent cell-mediated cytotoxicity may be operative in vivo.