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BACKGROUND: Oxidative stress is one of the causative factors in the pathogenesis of neurodegenerative diseases including mild cognitive impairment (MCI) and dementia. We previously reported that molecular hydrogen (H2) acts as a therapeutic and preventive antioxidant. OBJECTIVE: We assess the effects of drinking H2-water (water infused with H2) on oxidative stress model mice and subjects with MCI. METHODS: Transgenic mice expressing a dominant-negative form of aldehyde dehydrogenase 2 were used as a dementia model. The mice with enhanced oxidative stress were allowed to drink H2-water. For a randomized double-blind placebo-controlled clinical study, 73 subjects with MCI drank ~300 mL of H2-water (H2-group) or placebo water (control group) per day, and the Alzheimer's Disease Assessment Scale-cognitive subscale (ADAS-cog) scores were determined after 1 year. RESULTS: In mice, drinking H2-water decreased oxidative stress markers and suppressed the decline of memory impairment and neurodegeneration. Moreover, the mean lifespan in the H2-water group was longer than that of the control group. In MCI subjects, although there was no significant difference between the H2- and control groups in ADAS-cog score after 1 year, carriers of the apolipoprotein E4 (APOE4) genotype in the H2-group were improved significantly on total ADAS-cog score and word recall task score (one of the sub-scores in the ADAS-cog score). CONCLUSION: H2-water may have a potential for suppressing dementia in an oxidative stress model and in the APOE4 carriers with MCI.

DOI： 10.2174/1567205014666171106145017

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BACKGROUND: Ischemia-reperfusion injury is one of the leading causes of tissue damage and dysfunction, in particular, free tissue transfer, traumatically amputated extremity, and prolonged tourniquet application during extremity surgery. In this study, the authors investigated the therapeutic effects of hydrogen gas on skeletal muscle ischemia-reperfusion injury. METHODS: The authors compared the concentration of hydrogen in a muscle on intraperitoneal administration of hydrogen-rich saline and on inhalation of hydrogen gas. Animals were subjected to ischemia-reperfusion. Mice were treated with inhalation of hydrogen gas, and the hind gastrocnemius muscle was collected. Muscle morphology and inflammatory change were evaluated after ischemia-reperfusion. Moreover, a footprint test was performed to assess the functional effect of hydrogen. RESULTS: Hydrogen concentration of tissue was significantly higher, and the elevated level was maintained longer by hydrogen gas inhalation than by intraperitoneal administration of hydrogen-rich saline. Infarct zone and area with loss of tissue structure and marked cellular infiltration were significantly decreased in groups treated by hydrogen gas inhalation during ischemia-reperfusion; however, these effects were not observed by posttreatment of hydrogen. One week after ischemia-reperfusion, mice that had been pretreated with hydrogen gas recovered faster and achieved smoother walking in appearance compared with mice in the other groups as assessed by the footprint test. CONCLUSIONS: Inhalation of hydrogen gas attenuates muscle damage, inhibits inflammatory response, and enhances functional recovery. These findings suggest that the optimal route for hydrogen delivery is continuous inhalation of hydrogen gas, which could be a novel clinical mode of treatment in ischemia-reperfusion injury.

DOI： 10.1097/PRS.0000000000003878

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A number of alternations in mitochondrial DNA (mtDNA) have been reported in different types of cancers, and the role of mtDNA in cancer has been attracting increasing interest. In order to investigate the relationship between mtDNA alternations and chemosensitivity, we constructed cybrid (trans-mitochondrial hybrid) cell lines carrying a HeLa nucleus and the mtDNA of healthy individuals because of the presence of somatic alternations in the mtDNA of many cancer cells. After a treatment with 1.0 mu g/mL cisplatin for 10 days, we isolated 100 cisplatin-resistant clones, 70 of which carried the shorter mtDNA OriB variant (16184-16193 poly-cytosine tract), which was located in the control region of mtDNA. Whole mtDNA sequencing of 10 clones revealed no additional alternations. Re-construction of the HeLa nucleus and mtDNA from cisplatin-resistant cells showed that cisplatin resistance was only acquired by mtDNA alternations in the control region, and not by possible alternation(s) in the nuclear genome.

DOI： 10.1038/srep46240

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Histone acetylation has been linked to cardiac hypertrophy and heart failure. However, the pathological implications of changes in histone methylation and the effects of interventions with histone methyltransferase inhibitors for heart failure have not been fully clarified. Here, we focused on H3K9me3 status in the heart and investigated the effects of the histone H3K9 methyltransferase inhibitor chaetocin on prognoses in Dahl salt-sensitive rats, an animal model of chronic heart failure. Chaetocin prolonged survival and restored mitochondrial dysfunction. ChIP-seq analysis demonstrated that chronic stress to the heart induced H3K9me3 elevation in thousands of repetitive elements, including intronic regions of mitochondria-related genes, such as the gene encoding peroxisome proliferator-activated receptor-gamma coactivator 1 alpha. Furthermore, chaetocin reversed this effect on these repetitive loci. These data suggested that excessive heterochromatinization of repetitive elements of mitochondrial genes in the failing heart may lead to the silencing of genes and impair heart function. Thus, chaetocin may be a potential therapeutic agent for chronic heart failure.

DOI： 10.1038/srep39752

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Language：Japanese   Publisher：The Japanese Society of Physical Fitness and Sports Medicine  

DOI： 10.7600/jspfsm.66.63_2

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Fucoidan, a sulfated polysaccharide extracted from brown seaweeds, possesses many biological activities including anti-inflammatory and antioxidantactivities. We aimed to investigate the protective effects of fucoidan on dyslipidemia and atherosclerosis in apolipoprotein E-deficient mice (ApoE(shl) mice) and to elucidate its molecular targets in the liver by using a transcriptomic approach. For 12 weeks, ApoE(shl) mice were fed a high-fat diet (HFD) supplemented with either 1% or 5% fucoidan. Fucoidan supplementation significantly reduced tissue weight (liver and white adipose tissue), blood lipid, total cholesterol (TC), triglyceride (TG), non-high-density lipoprotein cholesterol (non-HDL-C) and glucose levels in HFD-fed ApoE(shl) mice but increased plasma lipoprotein lipase (LPL) activity and HDL-C levels. Fucoidan also reduced hepatic steatosis levels (liver size, TC and TG levels, and lipid peroxidation) and increased white adipose tissue LPL activity. DNA microarray analysis and quantitative reverse transcription-polymerase chain reaction demonstrated differential expression of genes encoding proteins involved in lipid metabolism, energy homeostasis and insulin sensitivity, by activating Ppara and inactivating Srebf1. Fucoidan supplementation markedly reduced the thickness of the lipid-rich plaque, lipid peroxidation and foaming macrophage accumulation in the aorta in HFD-fed ApoE(shl) mice. Thus, fucoidan supplementation appears to have anti-dyslipidemic and anti-atherosclerotic effects by inducing LPL activity and inhibiting the effects of inflammation and oxidative stress in HFD-fed ApoE(shl) mice. (C) 2016 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.jnutbio.2016.01.011

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We previously showed that H2 acts as a novel antioxidant to protect cells against oxidative stress. Subsequently, numerous studies have indicated the potential applications of H2 in therapeutic and preventive medicine. Moreover, H2 regulates various signal transduction pathways and the expression of many genes. However, the primary targets of H2 in the signal transduction pathways are unknown. Here, we attempted to determine how H2 regulates gene expression. In a pure chemical system, H2 gas (approximately 1%, v/v) suppressed the autoxidation of linoleic acid that proceeds by a free radical chain reaction, and pure 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphocholine (PAPC), one of the major phospholipids, was autoxidized in the presence or absence of H2. H2 modified the chemical production of the autoxidized phospholipid species in the cell-free system. Exposure of cultured cells to the H2-dependently autoxidized phospholipid species reduced Ca(2+) signal transduction and mediated the expression of various genes as revealed by comprehensive microarray analysis. In the cultured cells, H2 suppressed free radical chain reaction-dependent peroxidation and recovered the increased cellular Ca(2+), resulting in the regulation of Ca(2+)-dependent gene expression. Thus, H2 might regulate gene expression via the Ca(2+) signal transduction pathway by modifying the free radical-dependent generation of oxidized phospholipid mediators.

DOI： 10.1038/srep18971

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We previously reported that molecular hydrogen (H2) acts as a novel antioxidant to exhibit multiple functions. Moreover, long-term drinking of H2-water (water infused with H2) enhanced energy expenditure to improve obesity and diabetes in db/db mice accompanied by the increased expression of fibroblast growth factor 21 (FGF21) by an unknown mechanism. H2 was ingested by drinking of H2-water or by oral administration of an H2-producing material, MgH2. The comprehensive gene expression profile in the liver of db/db mice was analyzed by DNA microarray. The molecular mechanisms underlying the gene expression profile was investigated using cultured HepG2 cells. Moreover, the effects on lifespan of drinking H2-water were examined using wild-type mice that were fed a fatty diet. Pathway analyses based on comprehensive gene expression revealed the increased expression of various genes involved in fatty acid and steroid metabolism. As a transcription pathway, the PPARα signaling pathway was identified to upregulate their genes by ingesting H2. As an early event, the gene expression of PGC-1α was transiently increased, followed by increased expression of FGF21. The expression of PGC-1α might be regulated indirectly through sequential regulation by H2, 4-hydroxy-2-nonenal, and Akt/FoxO1 signaling, as suggested in cultured cell experiments. In wild-type mice fed the fatty diet, H2-water improved the level of plasma triglycerides and extended their average of lifespan. H2 induces expression of the PGC-1α gene, followed by stimulation of the PPARα pathway that regulates FGF21, and the fatty acid and steroid metabolism.

DOI： 10.1038/npjamd.2016.8

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY-BLACKWELL  

BackgroundOxidative and nitrative processes have an important role in the pathogenesis of glaucomatous neurodegeneration. Oxidative stress occurs when cellular production of reactive oxygen species outweighs the protective capacity of antioxidant defences. Reactive oxygen species are generated as by-products of cellular metabolism, primarily in the mitochondria. Herein, we present a novel investigation of the effects of molecular hydrogen (H-2) on retinal cells exposed to oxidative stress.
MethodsWe cultured adult rat retinal tissues in an organotypic culture system with a nitric oxide donor, S-nitroso-N-acetylpenicillamine, in the presence or absence of H-2. Loss of mitochondrial membrane potential and apoptosis of retinal cells were analysed using a MitoTMRE detection kit and TdT-mediated dUTP nick end labeling (TUNEL) assay, respectively. Tyrosine nitration levels and oxidative stress damage in the retina were evaluated using immunohistochemical staining. Retinal damage was quantified by measuring the numbers of cells in the ganglion cell and inner nuclear layers and the thickness of the retina.
ResultsH(2) suppressed loss of mitochondrial membrane potential and apoptosis in retinal cells. Moreover, H-2 decreased the tyrosine nitration level and suppressed oxidative stress damage in retinal cells. S-nitroso-N-acetylpenicillamine treatment decreased the cell numbers in the ganglion cell layer and inner nuclear layer, but the presence of H-2 inhibited this reduction. These findings suggest that H-2 has a neuroprotective effect against retinal cell oxidative damage, presumably by scavenging peroxynitrite.
ConclusionsH(2) reduces cellular peroxynitrite, a highly toxic reactive nitrogen species. Thus, H-2 may be an effective and novel clinical tool for treating glaucoma and other oxidative stress-related diseases.

DOI： 10.1111/ceo.12525

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Stem cell transplantation therapy is currently in clinical trials for the treatment of ischemic stroke, and several beneficial aspects have been reported. Similarly, in Alzheimer's disease (AD), stem cell therapy is expected to provide an efficient therapeutic approach. Indeed, the intracerebral transplantation of stem cells reduced amyloid-β (Aβ) deposition and rescued memory deficits in AD model mice. Here, we show that intravenous transplantation of bone marrow-derived mononuclear cells (BMMCs) improves cognitive function in two different AD mouse models, DAL and APP mice, and prevents neurodegeneration. GFP-positive BMMCs were isolated from tibiae and femurs of 4-week-old mice and then transplanted intravenously into DAL and APP mice. Transplantation of BMMCs suppressed neuronal loss and restored memory impairment of DAL mice to almost the same level as in wild-type mice. Transplantation of BMMCs to APP mice reduced Aβ deposition in the brain. APP mice treated with BMMCs performed significantly better on behavioral tests than vehicle-injected mice. Moreover, the effects were observed even with transplantation after the onset of cognitive impairment in DAL mice. Together, our results indicate that intravenous transplantation of BMMCs has preventive effects against the cognitive decline in AD model mice and suggest a potential therapeutic effect of BMMC transplantation therapy.

DOI： 10.1016/j.brainres.2015.02.011

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Oxidative stress is known to play a prominent role in the onset and early stage progression of Alzheimer's disease (AD). For example, protein oxidation and lipid peroxidation levels are increased in patients with mild cognitive impairment. Here, we created a double-transgenic mouse model of AD to explore the pathological and behavioral effects of oxidative stress. Double transgenic (APP/DAL) mice were constructed by crossing Tg2576 (APP) mice, which express a mutant form of human amyloid precursor protein (APP), with DAL mice expressing a dominant-negative mutant of mitochondrial aldehyde dehydrogenase 2 (ALDH2), in which oxidative stress is enhanced. Y-maze and object recognition tests were performed at 3 and 6 months of age to evaluate learning and memory. The accumulation of amyloid plaques, deposition of phosphorylated-tau protein, and number of astrocytes in the brain were assessed histopathologically at 3, 6, 9, and 12-15 months of age. The life span of APP/DAL mice was significantly shorter than that of APP or DAL mice. In addition, they showed accelerated amyloid deposition, tau phosphorylation, and gliosis. Furthermore, these mice showed impaired performance on Y-maze and object recognition tests at 3 months of age. These data suggest that oxidative stress accelerates cognitive dysfunction and pathological insults in the brain. APP/DAL mice could be a useful model for exploring new approaches to AD treatment.

DOI： 10.1016/j.neulet.2014.12.033

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：LIPPINCOTT WILLIAMS & WILKINS  

Background-We have previously shown that hydrogen (H-2) inhalation, begun at the start of hyperoxic cardiopulmonary resuscitation, significantly improves brain and cardiac function in a rat model of cardiac arrest. Here, we examine the effectiveness of this therapeutic approach when H-2 inhalation is begun on the return of spontaneous circulation (ROSC) under normoxic conditions, either alone or in combination with targeted temperature management (TTM).
Methods and Results-Rats were subjected to 6 minutes of ventricular fibrillation cardiac arrest followed by cardiopulmonary resuscitation. Five minutes after achieving ROSC, post-cardiac arrest rats were randomized into 4 groups: mechanically ventilated with 26% O-2 and normothermia (control); mechanically ventilated with 26% O-2, 1.3% H-2, and normothermia (H-2); mechanically ventilated with 26% O-2 and TTM (TTM); and mechanically ventilated with 26% O-2, 1.3% H-2, and TTM (TTM+H-2). Animal survival rate at 7 days after ROSC was 38.4% in the control group, 71.4% in the H-2 and TTM groups, and 85.7% in the TTM+H-2 group. Combined therapy of TTM and H-2 inhalation was superior to TTM alone in terms of neurological deficit scores at 24, 48, and 72 hours after ROSC, and motor activity at 7 days after ROSC. Neuronal degeneration and microglial activation in a vulnerable brain region was suppressed by both TTM alone and H-2 inhalation alone, with the combined therapy of TTM and H-2 inhalation being most effective.
Conclusions-H-2 inhalation was beneficial when begun after ROSC, even when delivered in the absence of hyperoxia. Combined TTM and H-2 inhalation was more effective than TTM alone.

DOI： 10.1161/CIRCULATIONAHA.114.011848

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Oxidative stress is involved in many age-associated diseases, as well as in the aging process itself. The development of interventions to reduce oxidative stress is hampered by the absence of sensitive detection methods that can be used in live animals. We generated transgenic mice expressing ratiometric redox-sensitive green fluorescent protein (roGFP) in the cytosol or mitochondria of several tissues, including skin epidermal keratinocytes. Crossbreeding into hairless albino mice allowed noninvasive optical measurement of skin oxidative state. Topical application of hydrogen peroxide emulsion shifted the keratinocyte redox state toward oxidation within minutes and could be observed in real time by fluorescence ratio imaging. Exposing skin to 365 nm UVA radiation oxidized roGFP localized in keratinocyte mitochondria, but not when roGFP was localized in the cytosol. This suggests that significant amounts of the endogenous photosensitizers that mediate UVA induced oxidative stress are located in the mitochondria. UVR is the major environmental cause of skin aging and UVA mediated oxidative stress has been associated with the development of wrinkles in humans. Direct measurements of redox state in defined cell compartments of live animals should be a powerful and convenient tool for evaluating treatments that aim to modulate oxidative stress.

DOI： 10.1038/jid.2013.428

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：CELL PRESS  

Accumulating evidence has suggested a role for p53 activation in various age-associated conditions. Here, we identified a crucial role of endothelial p53 activation in the regulation of glucose homeostasis. Endothelial expression of p53 was markedly upregulated when mice were fed a high-calorie diet. Disruption of endothelial p53 activation improved dietary inactivation of endothelial nitric oxide synthase that upregulated the expression of peroxisome proliferator-activated receptor-gamma coactivator-1 alpha in skeletal muscle, thereby increasing mitochondrial biogenesis and oxygen consumption. Mice with endothelial cell-specific p53 deficiency fed a high-calorie diet showed improvement of insulin sensitivity and less fat accumulation, compared with control littermates. Conversely, upregulation of endothelial p53 caused metabolic abnormalities. These results indicate that inhibition of endothelial p53 could be a novel therapeutic target to block the vicious cycle of cardiovascular and metabolic abnormalities associated with obesity.

DOI： 10.1016/j.celrep.2014.04.046

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There is increasing evidence that nutrient-sensing machinery is critically involved in the regulation of aging. The insulin/insulin-like growth factor-1 signaling pathway is the best-characterized pathway with an influence on longevity in a variety of organisms, ranging from yeast to rodents. Reduced expression of the receptor for this pathway has been reported to prolong the lifespan
however, the underlying mechanisms are largely unknown. Here we show that haploinsufficiency of Akt1 leads to an increase of the lifespan in mice. Akt1+/- mice had a lower body weight than their littermates with less fat mass and normal glucose metabolism. Ribosomal biogenesis and the mitochondrial DNA content were significantly reduced in these mice, along with a decrease of oxidative stress. Consistent with the results obtained in mice, inhibition of Akt-1 promoted longevity in nematodes (Caenorhabditis elegans), whereas activation of Akt-1 shortened the lifespan. Inhibition of Akt-1 led to a decrease of ribosomal gene expression and the mitochondrial DNA content in both human cells and nematodes. Moreover, deletion of ribosomal gene expression resulted in a decrease of the mitochondrial DNA content and normalized the lifespan shortened by Akt-1 activation in nematodes. These results suggest that an increase of mitochondrial amount and energy expenditure associated with enhanced protein synthesis accelerates both aging and the onset of age-associated diseases. © 2013 Nojima et al.

DOI： 10.1371/journal.pone.0069178

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Language：Japanese   Publisher：(一社)日本抗加齢医学会  

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The formation, maintenance and reorganization of synapses are critical for brain development and the responses of neuronal circuits to environmental challenges. Here we describe a novel role for peroxisome proliferator-activated receptor gamma co-activator 1 alpha, a master regulator of mitochondrial biogenesis, in the formation and maintenance of dendritic spines in hippocampal neurons. In cultured hippocampal neurons, proliferator-activated receptor gamma coactivator 1 alpha overexpression increases dendritic spines and enhances the molecular differentiation of synapses, whereas knockdown of proliferator-activated receptor gamma coactivator 1 alpha inhibits spinogenesis and synaptogenesis. Proliferator-activated receptor gamma co-activator 1 alpha knockdown also reduces the density of dendritic spines in hippocampal dentate granule neurons in vivo. We further show that brain-derived neurotrophic factor stimulates proliferator-activated receptor gamma co-activator-1 alpha-dependent mitochondrial biogenesis by activating extracellular signal-regulated kinases and cyclic AMP response element-binding protein. Proliferator-activated receptor gamma co-activator-1 alpha knockdown inhibits brain-derived neurotrophic factor-induced dendritic spine formation without affecting expression and activation of the brain-derived neurotrophic factor receptor tyrosine receptor kinase B. Our findings suggest that proliferator-activated receptor gamma co-activator-1 alpha and mitochondrial biogenesis have important roles in the formation and maintenance of hippocampal dendritic spines and synapses.

DOI： 10.1038/ncomms2238

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Background-All clinical and biological manifestations related to postcardiac arrest (CA) syndrome are attributed to ischemia-reperfusion injury in various organs including brain and heart. Molecular hydrogen (H-2) has potential as a novel antioxidant. This study tested the hypothesis that inhalation of H-2 gas starting at the beginning of cardiopulmonary resuscitation (CPR) could improve the outcome of CA.
Methods and Results-Ventricular fibrillation was induced by transcutaneous electrical epicardial stimulation in rats. After 5 minutes of the subsequent CA, rats were randomly assigned to 1 of 4 experimental groups at the beginning of CPR: mechanical ventilation (MV) with 2% N-2 and 98% O-2 under normothermia (37 degrees C), the control group; MV with 2% H-2 and 98% O-2 under normothermia; MV with 2% N-2 and 98% O-2 under therapeutic hypothermia (TH), 33 degrees C; and MV with 2% H-2 and 98% O-2 under TH. Mixed gas inhalation and TH continued until 2 hours after the return of spontaneous circulation (ROSC). H-2 gas inhalation yielded better improvement in survival and neurological deficit score (NDS) after ROSC to an extent comparable to TH. H-2 gas inhalation, but not TH, prevented a rise in left ventricular end-diastolic pressure and increase in serum IL-6 level after ROSC. The salutary impact of H-2 gas was at least partially attributed to the radical-scavenging effects of H-2 gas, because both 8-OHdG- and 4-HNE-positive cardiomyocytes were markedly suppressed by H-2 gas inhalation after ROSC.
Conclusions-Inhalation of H-2 gas is a favorable strategy to mitigate mortality and functional outcome of post-CA syndrome in a rat model, either alone or in combination with TH.

DOI： 10.1161/JAHA.112.003459

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Objective Post-transcriptional taurine modification at the first anticodon ("wobble") nucleotide is deficient in A3243G-mutant mitochondrial (mt) tRNA(Leu(UUR)) of patients with myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS). Wobble nucleotide modifications in tRNAs have recently been identified to be important in the accurate and efficient deciphering of codons. We herein examined whether taurine can alleviate mitochondrial dysfunction in patient-derived pathogenic cells and prevent clinical symptoms in MELAS patients.
Methods and Results The addition of taurine to the culture media ameliorated the reduced oxygen consumption, decreased the mitochondrial membrane potential, and increased the oxidative stress in MELAS patient-derived cells. Moreover, high dose oral administration of taurine (0.25 g/kg/day) completely prevented stroke-like episodes in two MELAS patients for more than nine years.
Conclusion Taurine supplementation may be a novel potential treatment option for preventing the stroke-like episodes associated with MELAS.

DOI： 10.2169/internalmedicine.51.7529

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC  

Kruppel-like factor 15 (KLF15), a member of the Kruppel-like factor family of transcription factors, has been found to play diverse roles in adipocytes in vitro. However, little is known of the function of KLF15 in adipocytes in vivo. We have now found that the expression of KLF15 in adipose tissue is down-regulated in obese mice, and we therefore generated adipose tissue-specific KLF15 transgenic (aP2-KLF15Tg) mice to investigate the possible contribution of KLF15 to various pathological conditions associated with obesity in vivo. The aP2-KLF15 Tg mice manifest insulin resistance and are resistant to the development of obesity induced by maintenance on a high fat diet. However, they also exhibit improved glucose tolerance as a result of enhanced insulin secretion. Furthermore, this enhancement of insulin secretion was shown to result from down-regulation of the expression of stearoyl-CoA desaturase 1 (SCD1) in white adipose tissue and a consequent reduced level of oxidative stress. This is supported by the findings that restoration of SCD1 expression in white adipose tissue of aP2-KLF15 Tg mice exhibited increased oxidative stress in white adipose tissue and reduced insulin secretion with hyperglycemia. Our data thus provide an example of cross-talk between white adipose tissue and pancreatic beta cells mediated through modulation of oxidative stress.

DOI： 10.1074/jbc.M111.242651

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Terasaki Y, Ohsawa I, Terasaki M, Takahashi M, Kunugi S, Dedong K, Urushiyama H, Amenomori S, Kaneko-Togashi M, Kuwahara N, Ishikawa A, Kamimura N, Ohta S, Fukuda Y. Hydrogen therapy attenuates irradiation-induced lung damage by reducing oxidative stress. Am J Physiol Lung Cell Mol Physiol 301: L415-L426, 2011. First published July 15, 2011; doi: 10.1152/ajplung.00008.2011.-Molecular hydrogen (H-2) is an efficient antioxidant that diffuses rapidly across cell membranes, reduces reactive oxygen species (ROS), such as hydroxyl radicals and peroxynitrite, and suppresses oxidative stress-induced injury in several organs. ROS have been implicated in radiation-induced damage to lungs. Because prompt elimination of irradiation-induced ROS should protect lung tissue from damaging effects of irradiation, we investigated the possibility that H-2 could serve as a radioprotector in the lung. Cells of the human lung epithelial cell line A549 received 10 Gy irradiation with or without H-2 treatment via H-2-rich PBS or medium. We studied the possible radioprotective effects of H-2 by analyzing ROS and cell damage. Also, C57BL/6J female mice received 15 Gy irradiation to the thorax. Treatment groups inhaled 3% H-2 gas and drank H-2-enriched water. We evaluated acute and late-irradiation lung damage after H-2 treatment. H-2 reduced the amount of irradiation-induced ROS in A549 cells, as shown by electron spin resonance and fluorescent indicator signals. H-2 also reduced cell damage, measured as levels of oxidative stress and apoptotic markers, and improved cell viability. Within 1 wk after whole thorax irradiation, immunohistochemistry and immunoblotting showed that H-2 treatment reduced oxidative stress and apoptosis, measures of acute damage, in the lungs of mice. At 5 mo after irradiation, chest computed tomography, Ashcroft scores, and type III collagen deposition demonstrated that H-2 treatment reduced lung fibrosis (late damage). This study thus demonstrated that H-2 treatment is valuable for protection against irradiation lung damage with no known toxicity.

DOI： 10.1152/ajplung.00008.2011

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：LIPPINCOTT WILLIAMS & WILKINS  

Rationale: Caloric restriction (CR) confers cardioprotection against ischemia/reperfusion injury. However, the exact mechanism(s) underlying CR-induced cardioprotection remain(s) unknown. Recent evidence indicates that Sirtuins, NAD(+)-dependent deacetylases, regulate various favorable aspects of the CR response. Thus, we hypothesized that deacetylation of specific mitochondrial proteins during CR preserves mitochondrial function and attenuates production of reactive oxygen species during ischemia/reperfusion.
Objective: The objectives of the present study were (1) to investigate the effect of CR on mitochondrial function and mitochondrial proteome and (2) to investigate what molecular mechanisms mediate CR-induced cardioprotection.
Methods and Results: Male 26-week-old Fischer344 rats were randomly divided into ad libitum-fed and CR (40% reduction) groups for 6 months. No change was observed in basal mitochondrial function, but CR preserved postischemic mitochondrial respiration and attenuated postischemic mitochondrial H2O2 production. CR decreased the level of acetylated mitochondrial proteins that were associated with enhanced Sirtuin activity in the mitochondrial fraction. We confirmed a significant decrease in the acetylated forms of NDUFS1 and cytochrome bc1 complex Rieske subunit in the CR heart. Low-dose Resveratrol treatment mimicked the effect of CR on deacetylating them and attenuated reactive oxygen species production during anoxia/reoxygenation in cultured cardiomyocytes without changing the expression levels of manganese superoxide dismutase. Treatment with nicotinamide completely abrogated the effect of low-dose Resveratrol.
Conclusions: These results strongly suggest that CR primes mitochondria for stress resistance by deacetylating specific mitochondrial proteins of the electron transport chain. Targeted deacetylation of NDUFS1 and/or Rieske subunit might have potential as a novel therapeutic approach for cardioprotection against ischemia/reperfusion. (Circ Res. 2011;109:396-406.)

DOI： 10.1161/CIRCRESAHA.111.243097

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Recent extensive studies have revealed that molecular hydrogen (H-2) has great potential for improving oxidative stress-related diseases by inhaling H-2 gas, injecting saline with dissolved H-2, or drinking water with dissolved H-2 (H-2-water); however, little is known about the dynamic movement of H-2 in a body. First, we show that hepatic glycogen accumulates H-2 after oral administration of H-2-water, explaining why consumption of even a small amount of H-2 over a short span time efficiently improves various disease models. This finding was supported by an in vitro experiment in which glycogen solution maintained H-2. Next, we examined the benefit of ad libitum drinking H-2-water to type 2 diabetes using db/db obesity model mice lacking the functional leptin receptor. Drinking H-2-water reduced hepatic oxidative stress, and significantly alleviated fatty liver in db/db mice as well as high fat-diet-induced fatty liver in wild-type mice. Long-term drinking H-2-water significantly controlled fat and body weights, despite no increase in consumption of diet and water. Moreover, drinking H-2-water decreased levels of plasma glucose, insulin, and triglyceride, the effect of which on hyperglycemia was similar to diet restriction. To examine how drinking H-2-water improves obesity and metabolic parameters at the molecular level, we examined gene-expression profiles, and found enhanced expression of a hepatic hormone, fibroblast growth factor 21 (FGF21), which functions to enhance fatty acid and glucose expenditure. Indeed, H-2 stimulated energy metabolism as measured by oxygen consumption. The present results suggest the potential benefit of H-2 in improving obesity, diabetes, and metabolic syndrome.

DOI： 10.1038/oby.2011.6

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Background: Molecular hydrogen (H&lt
inf&gt
2&lt
/inf&gt
) functions as an extensive protector against oxidative stress, inflammation and allergic reaction in various biological models and clinical tests
however, its essential mechanisms remain unknown. H&lt
inf&gt
2&lt
/inf&gt
directly reacts with the strong reactive nitrogen species peroxynitrite (ONOO&lt
sup&gt
-&lt
/sup&gt
) as well as hydroxyl radicals (OH), but not with nitric oxide radical (NO). We hypothesized that one of the H &lt
inf&gt
2&lt
/inf&gt
functions is caused by reducing cellular ONOO&lt
sup&gt
-&lt
/sup&gt
, which is generated by the rapid reaction of NO with superoxides (O&lt
inf&gt
2&lt
/inf&gt
&lt
sup&gt
-&lt
/sup&gt
). To verify this hypothesis, we examined whether H&lt
inf&gt
2&lt
/inf&gt
could restore cytotoxicity and transcriptional alterations induced by ONOO &lt
sup&gt
-&lt
/sup&gt
derived from NO in chondrocytes. Methods. We treated cultured chondrocytes from porcine hindlimb cartilage or from rat meniscus fibrecartilage with a donor of NO, S-nitroso-N-acetylpenicillamine (SNAP) in the presence or absence of H&lt
inf&gt
2&lt
/inf&gt
. Chondrocyte viability was determined using a LIVE/DEAD Viability/Cytotoxicity Kit. Gene expressions of the matrix proteins of cartilage and the matrix metalloproteinases were analyzed by reverse transcriptase-coupled real-time PCR method. Results: SNAP treatment increased the levels of nitrated proteins. H&lt
inf&gt
2&lt
/inf&gt
decreased the levels of the nitrated proteins, and suppressed chondrocyte death. It is known that the matrix proteins of cartilage (including aggrecan and type II collagen) and matrix metalloproteinases (such as MMP3 and MMP13) are down- and up-regulated by ONOO&lt
sup&gt
-&lt
/sup&gt
, respectively. H&lt
inf&gt
2&lt
/inf&gt
restoratively increased the gene expressions of aggrecan and type II collagen in the presence of H&lt
inf&gt
2&lt
/inf&gt
. Conversely, the gene expressions of MMP3 and MMP13 were restoratively down-regulated with H&lt
inf&gt
2&lt
/inf&gt
. Thus, H&lt
inf&gt
2&lt
/inf&gt
acted to restore transcriptional alterations induced by ONOO&lt
sup&gt
-&lt
/sup&gt
. Conclusions: These results imply that one of the functions of H&lt
inf&gt
2&lt
/inf&gt
exhibits cytoprotective effects and transcriptional alterations through reducing ONOO&lt
sup&gt
-&lt
/sup&gt
. Moreover, novel pharmacological strategies aimed at selective removal of ONOO&lt
sup&gt
-&lt
/sup&gt
may represent a powerful method for preventive and therapeutic use of H&lt
inf&gt
2&lt
/inf&gt
for joint diseases. © 2011 Hanaoka et al
licensee BioMed Central Ltd.

DOI： 10.1186/2045-9912-1-18

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELSEVIER SCIENCE INC  

It has been reported that dietary energy restriction, including intermittent fasting (IF), can protect heart and brain cells against injury and improve functional outcome in animal models of myocardial infarction (MI) and stroke. Here we report that IF improves glycemic control and protects the myocardium against ischemia-induced cell damage and inflammation in rats. Echocardiographic analysis of heart structural and functional variables revealed that IF attenuates the growth-related increase in posterior ventricular wall thickness, end systolic and diastolic volumes, and reduces the ejection fraction. The size of the ischemic infarct 24 h following permanent ligation of a coronary artery was significantly smaller, and markers of inflammation (infiltration of leukocytes in the area at risk and plasma IL-6 levels) were less, in IF rats compared to rats on the control diet. IF resulted in increased levels of circulating adiponectin prior to and after MI. Because recent studies have shown that adiponectin can protect the heart against ischemic injury, our findings suggest a potential role for adiponectin as a mediator of the cardioprotective effect of IF. Published by Elsevier Inc.

DOI： 10.1016/j.jnutbio.2009.01.020

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：SPRINGER  

Cisplatin is a widely used anti-cancer drug in the treatment of a wide range of tumors; however, its application is limited by nephrotoxicity, which is affected by oxidative stress. We have reported that molecular hydrogen (H(2)) acts as an efficient antioxidant (Ohsawa et al. in Nat Med 13:688-694, 2007). Here we show that hydrogen efficiently mitigates the side effects of cisplatin by reducing oxidative stress.
Mice were administered cisplatin followed by inhaling hydrogen gas (1% H(2) in air). Furthermore, instead of inhaling hydrogen gas, we examined whether drinking water containing hydrogen (hydrogen water; 0.8 mM H(2) in water) is applicable by examining oxidative stress, mortality, and body-weight loss. Nephrotoxicity was assessed by morphological changes, serum creatinine and blood urea nitrogen (BUN) levels.
Inhalation of hydrogen gas improved mortality and body-weight loss caused by cisplatin, and alleviated nephrotoxicity. Hydrogen was detected in blood when hydrogen water was placed in the stomach of a rat. Consuming hydrogen water ad libitum also reduced oxidative stress, mortality, and body-weight loss induced by cisplatin in mice. Hydrogen water improved metamorphosis accompanying decreased apoptosis in the kidney, and nephrotoxicity as assessed by serum creatinine and BUN levels. Despite its protective effects against cisplatin-induced toxicity, hydrogen did not impair anti-tumor activity of cisplatin against cancer cell lines in vitro and tumor-bearing mice in vivo.
Hydrogen has potential for improving the quality of life of patients during chemotherapy by efficiently mitigating the side effects of cisplatin.

DOI： 10.1007/s00280-008-0924-2

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

We have reported that hydrogen (H(2)) acts as an efficient antioxidant by gaseous rapid diffusion. When water saturated with hydrogen ( hydrogen water) was placed into the stomach of a rat, hydrogen was detected at several mu M level in blood. Because hydrogen gas is unsuitable for continuous consumption, we investigated using mice whether drinking hydrogen water ad libitum, instead of inhaling hydrogen gas, prevents cognitive impairment by reducing oxidative stress. Chronic physical restraint stress to mice enhanced levels of oxidative stress markers, malondialdehyde and 4-hydroxy-2-nonenal, in the brain, and impaired learning and memory, as judged by three different methods: passive avoidance learning, object recognition task, and the Morris water maze. Consumption of hydrogen water ad libitum throughout the whole period suppressed the increase in the oxidative stress markers and prevented cognitive impairment, as judged by all three methods, whereas hydrogen water did not improve cognitive ability when no stress was provided. Neural proliferation in the dentate gyrus of the hippocampus was suppressed by restraint stress, as observed by 5-bromo-2'-deoxyuridine incorporation and Ki-67 immunostaining, proliferation markers. The consumption of hydrogen water ameliorated the reduced proliferation although the mechanistic link between the hydrogen-dependent changes in neurogenesis and cognitive impairments remains unclear. Thus, continuous consumption of hydrogen water reduces oxidative stress in the brain, and prevents the stress-induced decline in learning and memory caused by chronic physical restraint. Hydrogen water may be applicable for preventive use in cognitive or other neuronal disorders.

DOI： 10.1038/npp.2008.95

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：PERGAMON-ELSEVIER SCIENCE LTD  

Many anticancer drugs attack rapidly dividing cells, including not only malignant cells but also hair follicle cells, and induce alopecia. Chemotherapy-induced alopecia (CIA) is an emotionally distressing side effect of cancer chemotherapy. There is currently no useful preventive therapy for CIA. We have previously constructed anti-death rFNK protein from rat Bcl-x(L) by site-directed mutagenesis to strengthen cytoprotective activity. When fused to the protein transduction domain (PTD) of HIV/Tat, the fusion protein PTD (TAT)-rFNK successfully entered cells from the outside in vitro and in vivo to exhibit anti-death activity against apoptosis and necrosis. Here, we show that topical application of FNK protected against CIA in a newborn rat model. The protective activity against hair-loss was observed in 30-1000 nM TAT-rFNK administrative groups in a dose-dependent manner. Furthermore, a human version of FNK (hFNK) fused to other PTD peptides exhibited a protective ability. These results suggest that PTD-FNK possesses protective activity against CIA and is not restricted to a sequence of PTD peptides or species of FNK. Thus, PTD-FNK represents potential to develop a useful method for preventing CIA in cancer patients. (c) 2007 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.lfs.2007.11.011

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：OXFORD UNIV PRESS  

To elucidate the transcriptional regulation in eukaryotic genome network, it is important to understand coevolution of transcription factors, transcriptional coactivators, and TATA-box-binding protein (TBP). In this study, coevolution of transcriptional coactivator multiprotein-bridging factor 1 and its interacting target TBP was first evaluated experimentally by examining if compensatory amino acid changes took place at interacting sites of both proteins. The experiments were conducted by identifying interaction sites and comparing the amino acids at these sites among different organisms. Here, we provide evidence for compensatory changes of transcription coactivator and its interacting target, presenting the 1st report that transcription coactivator may have undergone coevolution with TBP.

DOI： 10.1093/molbev/msm073

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：COMPANY OF BIOLOGISTS LTD  

To investigate the regulatory system in mitochondrial biogenesis involving crosstalk between the mitochondria and nucleus, we found a factor named MIDAS (mitochondrial DNA absence sensitive factor) whose expression was enhanced by the absence of mitochondrial DNA (mtDNA). In patients with mitochondrial diseases, MIDAS expression was increased only in dysfunctional muscle fibers. A majority of MIDAS localized to mitochondria with a small fraction in the Golgi apparatus in HeLa cells. To investigate the function of MIDAS, we stably transfected HeLa cells with an expression vector carrying MIDAS cDNA or siRNA. Cells expressing the MIDAS protein and the siRNA constitutively showed an increase and decrease in the total mass of mitochondria, respectively, accompanying the regulation of a mitochondria-specific phospholipid, cardiolipin. In contrast, amounts of the mitochondrial DNA, RNA and proteins did not depend upon MMAS. Thus, MIDAS is involved in the regulation of mitochondrial lipids, leading to increases of total mitochondrial mass in response to mitochondrial dysfunction.

DOI： 10.1242/jcs.02645

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER CHEMICAL SOC  

DNA polymerases delta and epsilon (pol delta and epsilon) are the two major replicative polymerases in the budding yeast Saccharomyces cerevisiae. The fidelity of pol delta is influenced by its 3'-5' proofreading exonuclease activity, which corrects misinsertion errors, and by enzyme cofactors. PCNA is a pol delta cofactor, called the sliding clamp, which increases the processivity of pol delta holoenzyme. This study measures the fidelity of 3'-5' exonuclease-proficient and -deficient pol delta holoenzyme using a synthetic 30mer primer/ 100mer template in the presence and absence of PCNA. Although PCNA increases pol delta processivity, the presence of PCNA decreased pol delta fidelity 2-7-fold. In particular, wild-type pol delta demonstrated the following nucleotide substitution efficiencies for mismatches in the absence of PCNA: G(.)G, 0.728 x 10(-4); T(.)G, 1.82 x 10(-4); A(.)G, <0.01 x 10(-4). In the presence of PCNA these values increased as follows: G(.)G, 1.30 x 10(-4); T(.)G, 2.62 x 10(-4); A(.)G, 0.074 x 10(-4). A similar but smaller effect was observed for exonuclease-deficient pol delta (i.e., 2-4-fold increase in nucleotide substitution efficiencies in the presence of PCNA). Thus, the fidelity of wild-type pol delta in the presence of PCNA is more than 2 orders of magnitude lower than the fidelity of wild-type pol is holoenzyme and is comparable to the fidelity of exonuclease-deficient pol epsilon holoenzyme.

DOI： 10.1021/bi0348359

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC  

Saccharomyces cerevisiae POL2 encodes the catalytic subunit of DNA polymerase epsilon. This study investigates the cellular functions performed by the polymerase domain of Pol2p and its role in DNA metabolism. The pol2-16 mutation has a deletion in the catalytic domain of DNA polymerase a that eliminates its polymerase and exonuclease activities. It is a viable mutant, which displays temperature sensitivity for growth and a defect in elongation step of chromosomal DNA replication even at permissive temperatures. This mutation is synthetic lethal in combination with temperature-sensitive mutants or the 3'- to 5'-exonuclease-deficient mutant of DNA polymerase delta in a haploid cell. These results suggest that the catalytic activity of DNA polymerase a participates in the same pathway as DNA polymerase delta, and this is consistent with the observation that DNA polymerases delta and epsilon colocalize in some punctate foci on yeast chromatids during S phase. The pol2-16 mutant senesces more rapidly than wild type strain and also has shorter telomeres. These results indicate that the DNA polymerase domain of Po12p is required for rapid, efficient, and highly accurate chromosomal DNA replication in yeast.

DOI： 10.1074/jbc.M111573200

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATL ACAD SCIENCES  

We report that POL5 encodes the fifth essential DNA polymerase in Saccharomyces cerevisiae. Pol5p was identified and purified from yeast cell extracts and is an aphidicolin-sensitive DNA polymerase that is stimulated by yeast proliferating cell nuclear antigen (PCNA). Thus, we named Pol5p DNA polymerase phi. Temperature-sensitive pol5-1similar to-3 mutants did not arrest at G(2)/M at the restrictive temperature. Furthermore, the polymerase active-site mutant POL5dn gene complements the lethality of Deltapol5. These results suggest that the polymerase activity of Pol5p is not required for the in vivo function of Pol5p. rRNA synthesis was severely inhibited at the restrictive temperature in the temperature-sensitive pol5-3 mutant cells, suggesting that an essential function of Pol5p is rRNA synthesis. Pol5p is localized exclusively to the nucleolus and binds near or at the enhancer region of rRNA-encoding DNA repeating units.

DOI： 10.1073/pnas.142277999

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY  

Background: DNA polymerase II (PolII), the homologue of mammalian DNA polymerase epsilon, is essential for chromosomal DNA replication in the budding yeast Saccharomyces cerevisiae and also participates in S-phase checkpoint control. An important issue is whether chromosomal DNA replication in other eukaryotes, including the fission yeast Schizosaccharomyces pombe-in which the characteristics of replication origins are poorly defined-also requires DNA polymerase E. It has been shown that DNA polymerase epsilon is not required for the in vitro replication of SV40 DNA by human cell extracts.
Results: We have cloned and sequenced S. pombe pol2(+), which is identical to the cell-cycle gene cdc20(+), encoding the catalytic polypeptide of DNA polymerase epsilon (Pol epsilon). The predicted amino acid sequence of Pol epsilon is highly homologous to that of S. cerevisiae PolII and human Pol epsilon. Consistent with this, the Pol epsilon polypeptide was recognized by polyclonal antibodies against S. cerevisiae PolII holoenzyme (PolII(star)). The terminal morphology of cells containing the disrupted pol2 gene was similar to that of DNA replication mutant cells and cdc20 mutant cells. Furthermore, the Pol epsilon activity from temperature-sensitive S. pombe cdc20 mutant cells was temperature-sensitive, and chromosomal DNA replication in the mutant cells was inhibited at the restrictive temperatures.
Conclusion: These data strongly suggest that Pol epsilon is required for normal chromosomal DNA replication in S. pombe, as is PolII in S. cerevisiae. Thus, eukaryotic chromosomal DNA is replicated differently from that of viral SV40 DNA.

DOI： 10.1046/j.1365-2443.1998.00169.x

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：OXFORD UNIV PRESS  

DNA polymerase III (delta) of Saccharomyces cerevisiae is purified as a complex of at least two polypeptides with molecular masses of 125 and 55 kDa as judged by SDS-PAGE. In this paper we determine partial amino acid sequences of the 125 and 55 kDa polypeptides and find that they match parts of the amino acid sequences predicted from the nucleotide sequence of the CDC2 and HYS2 genes respectively, We also show by Western blotting that Hys2 protein co-purifies with DNA polymerase III activity as well as Cdc2 polypeptide. The complex form of DNA polymerase III activity could not be detected in thermosensitive hys2 mutant cell extracts, although another form of DNA polymerase III was found, This form of DNA polymerase III, which could also be detected in wild-type extracts, was not associated with Hys2 protein and was not stimulated by addition of proliferating cell nuclear antigen (PCNA), replication factor A (RF-A) or replication factor C (RF-C), The temperature-sensitive growth phenotype of hys2-1 and hys2-2 mutations could be suppressed by the CDC2 gene on a multicopy plasmid, These data suggest that the 55 kDa polypeptide encoded by the HYS2 gene is one of the subunits of DNA polymerase III complex in S.cerevisiae and is required for highly processive DNA synthesis catalyzed by DNA polymerase III in the presence of PCNA, RF-A and RF-C.

DOI： 10.1093/nar/26.2.477

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Language：English   Publisher：ACADEMIC PRESS INC  

DOI： 10.1016/s0065-230x(08)60859-3

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY  

In the yeast cell cycle 'start' requires sets of the Cdc10/ SWI family of transcriptional factors which activate the MCB cis elements contained in genes essential for S phase progression. Fission yeast possess two such overlapping systems, Res1-Cdc10 and Res2-Cdc10, both of which act to start the mitotic and meiotic cycles, We have recently isolated rep2(+) as a multicopy suppressor of a temperature-sensitive cdc10 mutant which encodes a zinc finger protein. Here we show that the Rep2 zinc finger protein is an essential component of the active Res2-Cdc10 transcriptional regulator complex and likely to play a role in the control of cell cycle 'start'. Our data suggest that Rep2 is a transcriptional activator subunit which interacts with the MCB binding subunit complex formed by Res2 and Cdc10.

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：OXFORD UNIV PRESS UNITED KINGDOM  

We cloned by phenotypic complementation a novel Saccharomyces cerevisiae's multicopy suppressor of the Schizosaccharomyces pombe cdc10-129 mutant which we call HAC1, an acronym of 'homologous to ATF/CREB 1'. It encodes a bZIP (basic-leucine zipper) protein of 230 amino acids with close homology to the mammalian ATF/CREB transcription factor and gel-retardation assays showed that it binds specifically to the CRE motif. HAC1 is not essential for viability. However, the had disruptant becomes caffeine sensitive, which is suppressed by multicopy expression of the yeast PDE2 (Phosphodiesterase 2) gene. Although the mRNA level of HAC1 is almost constitutive throughout the cell cycle, it fluctuates during meiosis. The upstream region of the HAC1 gene contains a T4C site, a URS (upstream repression sequence) and a TR (T-rich) box-like sequence, which reside upstream of many meiotic genes. These results suggest that HAC1 may also be one of the meiotic genes.

DOI： 10.1093/nar/22.24.5279

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC  

This is the first report on a unique vitronectin molecule, yolk vitronectin, which is similar to its blood homologue in cell spreading activity but different in molecular size, bound carbohydrate, and heparin and collagen binding activity. Yolk vitronectin was purified 2,500-fold from chick egg yolk by a combination of hydroxylapatite, DEAE-cellulose, and anti-vitronectin-Sepharose column chromatographies. In SDS-polyacrylamide gel electrophoresis under reducing conditions, yolk vitronectin was separated into 54- and 45-kDa bands, which are 16 and 25 kDa smaller, respectively, than the 70-kDa major band of chick blood vitronectin. The 54-kDa band shares the same NH2-terminal sequence as chick blood vitronectin. In contrast, the NH2-terminal sequence of the 45-kDa band is somewhat homologous with the internal sequences of mammalian vitronectins beginning at the 50th amino acid from the NH2 terminus. The bound carbohydrate of the 54- and 45-kDa species of yolk vitronectin is similar to, but distinct from, that of blood vitronectin. Unlike blood vitronectin, yolk vitronectin cannot bind to either heparin or collagen.

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELSEVIER SCIENCE BV  

We isolated vitronectins from the plasma or sera of 14 animal species including mouse and rat by heparin affinity chromatography. They cross-reacted with anti-vitronectin antibody and their amino terminal sequences showed strong homology. They also promoted spreading of BHK cells and were bound to heparin and collagen in the same way. Therefore, these properties appear to be essential for vitronectin function. However, the apparent molecular weights of these vitronectins varied considerably from 59 to 78 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). In addition, the number of bands also varied from 1 to 3. To search for the uniformity of vitronectin polypeptide, vitronectins were deglycosylated and examined by Ferguson plot analysis. The size of the polypeptide portion of vitronectins was estimated to range from 40 to 57 kDa which was 19-26 kDa smaller than original values. Supposing a possible cleavage site at 5-13 kDa far from the carboxyl terminus, all vitronectin polypeptides were speculated to be synthesized de novo in the size range of 50-57 kDa. Proteins reacting with anti-vitronectin antibody were also detected on the immunoblot of 13 more species including Drosophila and Physarum. Almost all of these vitronectin-like proteins showed marked species-specific variations in their apparent molecular weights from 51 to 96 kDa in SDS-PAGE.

DOI： 10.1016/0167-4838(92)90417-c

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Language：English   Publishing type：Rapid communication, short report, research note, etc. (scientific journal)   Publisher：LIPPINCOTT WILLIAMS & WILKINS  

DOI： 10.1161/CIRCULATIONAHA.115.016785

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：LIPPINCOTT WILLIAMS & WILKINS  

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Language：Japanese   Publisher：(一財)代謝異常治療研究基金  

血管内皮細胞p53の抑制は食餌性肥満における代謝異常を改善するか検討した。野生型マウスに高脂肪高蔗糖食を投与して食餌性肥満モデルを作製し、大動脈・肺/骨格筋の血管内皮細胞を評価した。これらのマウスを用いて標本切片の染色・蛋白抽出やRNA抽出による分子発現確認を行った。次に血管内皮細胞p53の役割を評価するために血管内皮細胞特異的p53ノックアウトマウスを作製した。高カロリー食負荷によって血管内皮細胞でのGlut1発現がp53活性化に伴って減少し、この変化がp53ノックアウトによって消失するとともに骨格筋への糖取り込みが亢進した。p53はPTEN並びにGlut1の発現を調節することで糖代謝に関与していることが示唆された。

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Language：Japanese   Publishing type：Meeting report  

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：LIPPINCOTT WILLIAMS & WILKINS  

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Language：Japanese   Publisher：(一社)日本肥満学会  

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