Nonsynonymous site heteroplasmy in fish mitochondrial DNA

Heteroplasmic nucleotide polymorphisms are rarely observed in wild animal mitochondrial DNA. The occurrence of such site heteroplasmy is expected to be extremely rare at nonsynonymous sites where the number of nucleotide substitutions per site is low due to functional constraints. This report deals with nonsynonymous mitochondrial heteroplasmy from two wild fish species, chum salmon and Japanese flounder. We detected an A/C nonsynonymous heteroplasmic site corresponding to putative amino acids, Ile or Met, in NADH dehydrogenase subunit-5 (ND5) region of chum salmon. The heteroplasmic site was at the 3rd position of 58th codon. As for Japanese flounder we detected a C/T nonsynonymous heteroplasmic site corresponding to putative amino acids, Leu or Pro, in ND4 region. The heteroplasmic site was at the 2nd position of 450th codon. We also verified heteroplasmy at these sites by sequencing cloned fragments.     

Amyloid-beta fibril formation is not necessarily required for microglial activation by the peptides

There is increasing evidence that microglial activation has pathogenic influence on Alzheimer's disease. According to in vitro studies, microglia activated by amyloid-beta (Abeta) peptides have been reported to damage or kill neurons by the release of neurotoxic molecules such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta, nitric oxide or reactive oxygen species. Although the relationship between the aggregational state of Abeta peptides and their neurotoxic activities has been well investigated, little is known about the relationship between the aggregational state of Abeta peptides and their ability to induce microglial activation. In the present study, we thus performed both structural and biochemical studies to clarify the relationship between the aggregational state of Abeta peptides and their ability to activate microglia. Our results have shown that, in the presence of interferon-gamma, the Abeta25-35(M(35)Nle) peptide had almost the same potency of activating microglia and producing TNF-alpha as the Abeta25-35 peptide on both protein and mRNA levels, in spite of the fact that former peptide represented much less amyloid fibril formation than the latter in a thioflavine-T fluorometric assay. These results suggest that Abeta fibril formation is not necessarily required for microglial activation by the peptides.     

Effectiveness of aroma on work efficiency: lavender aroma during recesses prevents deterioration of work performance

The present study investigated whether exposure to aromas during recess periods affects work performance. Subjects comprised 36 healthy male students (mean age, 24.2 +/- 2.2 years) who were randomly divided into three groups: (1) control group, not exposed to aroma during recesses; (2) jasmine group, exposed to jasmine aroma during recesses; and (3) lavender group, exposed to lavender aroma during recesses. All participants completed five work sessions performing a task requiring concentration on a computer monitor, with each session lasting 60 min. Recess periods of 30 min were provided between each session. To clarify the time at which work concentration was lowest, work performance for the control group was analyzed. Concentration was lowest in the afternoon period, where afternoon drowsiness is strongest. Comparison of the three groups for this time period indicated significantly higher concentration levels for the lavender group than for the control group. No such effect was noted for the jasmine group. Although lavender is a sedative-type aroma, use during recess periods after accumulation of fatigue seems to prevent deterioration of performance in subsequent work sessions.     

A possible intermediate step during apoptotic execution

Many proteases are known to be involved in apoptosis. Among them, interleukin-1beta converting enzyme (ICE) and its family proteases, which are called caspases, play critical roles in the execution stage of apoptosis. We previously reported that a proteasome-inhibitor, benzyloxycarbonyl Leu-Leu-leucinal (ZLLLal), induced apoptosis in MOLT-4 cells. In the present study, in order to analyze the detailed mechanism of ZLLLal-induced apoptosis, we examined the effect of a caspase-inhibitor, acetyl(Ac)-Tyr-Val-Ala-Asp-chloromethyl ketone (AcYVADcmk), on ZLLLal-induced apoptosis in the cells. Agarose gel electrophoresis revealed that low concentrations of AcYVADcmk efficiently suppressed apoptotic DNA fragmentation. However, the cells presented morphology different from normal, apoptotic or necrotic cells, although DNA fragmentation was suppressed. The same examination was performed on the cells with anti-Fas antibody-induced apoptosis, and the same results were obtained. Some cells with a similar morphology were found even without the caspase-inhibitor in the early stage of anti-Fas antibody-induced physiological apoptosis. In addition, apoptotic cascade was reactivated by washing out the caspase inhibitor from the DNA degradation-suppressed cells. Therefore, this newly found morphological feature shows the presence of a step prior to caspase activation in the cells, and this is the first report presenting the pre-caspase-activated step in the apoptotic cascade.     

Facilitation of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate receptor transmission in the suprachiasmatic nucleus by aniracetam enhances photic responses of the biological clock in rodents

This study was designed to test whether the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor-facilitating drug, aniracetam, could potentiate photic responses of the biological clock in the suprachiasmatic nucleus (SCN) of rodents. Using the whole-cell patch technique, we first demonstrated that AMPA currents elicited by either local AMPA application or optic chiasm stimulation were augmented by aniracetam in the neurons of the SCN. The AMPA application-elicited increase of intracellular Ca2+ concentration in SCN slices was also enhanced by aniracetam treatment. The systemic injection of aniracetam dose-dependently (10-100 mg/kg) potentiated the phase delay in behavioral rhythm induced by brief light exposure of low intensity (3 lux) but not high intensity (10 or 60 lux) during early subjective night. Under the blockade of NMDA receptors by (+) MK801, aniracetam failed to potentiate a light (3 lux)-induced phase delay in behavioral rhythm. Aniracetam increased the photic induction of c-Fos protein in the SCN that was elicited by low intensity light exposure (3 lux). These results suggest that AMPA receptor-mediated responses facilitated by aniracetam can explain enhanced photic responses of the biological clock in the SCN of rodents.     

Epicubenol and Ferruginol induce DC from human monocytes and differentiate IL-10-producing regulatory T cells in vitro

Epicubenol and 19-hydroxyferruginol (Ferruginol) are sesquiterpenes isolated from the black heartwood of Cryptomeria japonica. Dendritic cells (DC) are specialized antigen-presenting cells that monitor the antigenic environment and activate na?ve T cells. The role of DC is not only to sense danger but also to tolerize the immune system to antigens encountered in the absence of maturation/inflammatory stimuli. In this study, we attempted to investigate the effects of Epicubenol and Ferruginol on the phenotypic and functional maturation of human monocytes-derived DC in vitro. Human monocytes were cultured with GM-CSF and IL-4 for 6 days under standard conditions, followed by another 2 days with Epicubenol or Ferruginol. The expression levels of CD1a, CD83, and HLA-DR as expressed by mean fluorescence intensity (MFI) on Epicubenol-primed DC or Ferruginol-primed DC were enhanced. Allogeneic Epicubenol-primed DC or Ferruginol-primed DC co-cultured with na?ve T cells at 1:5 ratio, secreted IL-10 and TGF-beta, but little IL-4. Moreover, T cells that develop in co-culture of Epicubenol-primed DC or Ferruginol-primed DC and na?ve T cells at 1:5 ratio suppressed the proliferation of autologous T cells at Treg cells: Ttarget cells and this suppression of proliferation was inhibited by anti-IL-10 mAb. The expression of FoxP3 mRNA on T cells that develop in co-culture of Epicubenol-primed DC or Ferruginol-primed DC and na?ve T cells was lower. From these results, Epicubenol and Ferruginol may induce IL-10-producing Treg 1 cells from na?ve T cells by modulating DC function. It seems that Epicubenol and Ferruginol appear to be a target for tolerance after transplantation and in autoimmune diseases.     

Interflavin one-electron transfer in the inducible nitric oxide synthase reductase domain and NADPH-cytochrome P450 reductase

In this study, we have analyzed interflavin electron transfer reactions from FAD to FMN in both the full-length inducible nitric oxide synthase (iNOS) and its reductase domain. Comparison is made with the interflavin electron transfer in NADPH-cytochrome P450 reductase (CPR). For the analysis of interflavin electron transfer and the flavin intermediates observed during catalysis we have used menadione (MD), which can accept an electron from both the FAD and FMN sites of the enzyme. A characteristic absorption peak at 630 and 520 nm can identify each FAD and FMN semiquinone species, which is derived from CPR and iNOS, respectively. The charge transfer complexes of FAD with NADP+ or NADPH were monitored at 750 nm. In the presence of MD, the air-stable neutral (blue) semiquinone form (FAD-FMNH*) was observed as a major intermediate during the catalytic cycle in both the iNOS reductase domain and full-length enzyme, and its formation occurred without any lag phase indicating rapid interflavin electron transfer following the reduction of FAD by NADPH. These data also strongly suggest that the low level reactivity of a neutral (blue) FMN semiquinone radical with electron acceptors enables one-electron transfer in the catalytic cycle of both the FAD-FMN pairs in CPR and iNOS. On the basis of these data, we propose a common model for the catalytic cycle of both CaM-bound iNOS reductase domain and CPR.