Emi Yoshigai

Ritsumeikan University, Kioto, Kyōto, Japan

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Publications (17)32.4 Total impact

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    ABSTRACT: Antipyretic analgesic drugs (including non-steroidal anti-inflammatory drugs) inhibit cyclooxygenase-2 and inducible nitric oxide synthase (iNOS), resulting in decreases of the proinflammatory mediators prostaglandin E2 and nitric oxide (NO), respectively. Both mediators are regulated by nuclear factor-kappa B (NF-κB), a key transcription factor in inflammation. Few reports have compared the efficacy and potency of anti-inflammatory drugs as NO inhibitors. In our study, we examined the effects of four popular antipyretic analgesic drugs on NO production induced in hepatocytes and macrophages. Mouse RAW264.7 macrophages treated with bacterial lipopolysaccharide showed the highest efficacy with regard to NO production; aspirin, loxoprofen, ibuprofen, and acetaminophen dose-dependently suppressed NO induction. Ibuprofen showed the highest potency in suppressing the induced production of NO. In rat hepatocytes, all the drugs inhibited interleukin 1β-induced NO production and ibuprofen and loxoprofen inhibited NO induction effectively. Unexpectedly, the potency of NO suppression of each drug in hepatocytes did not always correlate with that observed in RAW264.7 cells. Microarray analyses of mRNA expression in hepatocytes revealed that the effects of the four antipyretic analgesic drugs modulated the NF-κB signaling pathway in a similar manner to the regulation of the expression of genes associated with inflammation, including the iNOS gene. However, the affected signal-transducing molecules in the NF-κB pathway were different for each drug. Therefore, antipyretic analgesic drugs may decrease NO production by modulating the NF-κB pathway in different ways, which could confer different efficacies and potencies with regard to their anti-inflammatory effects. Copyright © 2014. Published by Elsevier Inc.
    Nitric Oxide 12/2014; · 3.18 Impact Factor
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    ABSTRACT: A new flavanone, shisoflavanone A (1), and several flavonoids were purified from the ethyl acetate-soluble fraction of green perilla leaves (Perilla frutescens Britton var. crispa form viridis), and their structures were identified. Shisoflavanone A was elucidated as 8-hydroxy-6,7-dimethoxyflavanone based on its spectral data. Other constituents of the ethyl acetate-soluble fraction, i.e. 5,8-dihydroxy-7-methoxyflavanone (2), negletein (5,6-dihydroxy-7-methoxyflavone) (3), luteolin (4), apigenin (5), esculetin (6), and protocatechuic acid (7), were identified. This is the first time that constituents 2, 3, and 6 have been found in green perilla. Shisoflavanone A and the other constituents (except 7) significantly inhibited nitric oxide production in interleukin 1β-stimulated rat hepatocytes, which have been used to monitor the anti-inflammatory effects of herbal constituents. The present findings suggest that these constituents, including shisoflavanone A, may be involved in the anti-inflammatory effects of green perilla leaves.
    Bioscience Biotechnology and Biochemistry 09/2014; · 1.27 Impact Factor
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    ABSTRACT: Dextran sodium sulfate (DSS)-induced colitis in rats is widely used as an experimental model for elucidating the etiology of ulcerative colitis (UC) and developing its novel remedy. We investigated the temporal and spatial changes in inflammatory mediators such as tumor necrosis factor (TNF)-α and inducible nitric oxide synthase (iNOS) in the regions of rectum and distal colon and examined whether statins, which were designed to lower plasma cholesterol levels, influenced those mediators. Colitis was induced in rats by oral administration of 5 % DSS for 5 days, followed by 2 % DSS for 10 days. 5 % DSS rats were treated with fluvastatin (20 mg/kg) concomitantly for 5 days. The expression of inflammatory mediators of a sequence of four regions in rectum (R) and distal colon (D0, D1, and D2) was determined by quantitative RT-PCR. The peak of colitic damage, which was confirmed clinically and histopathologically, was found on days 4-6. The expression of TNF-α, iNOS, cytokine-induced neutrophil chemoattractant-1, interleukin (IL)-1β, and IL-6 mRNA increased in R time dependently, showing the peak on days 4-6, and then decreased thereafter. The levels of mRNAs reduced from R to D0, D1, and D2 region dependently. Fluvastatin decreased the expression of these markers in addition to the prevention of DSS-induced damage. Results demonstrated that the expression of inflammatory biomarkers had time and region specificity and was markedly inhibited by fluvastatin. To obtain a precise drug effect for UC, it is important to elucidate the temporal and spatial dependence of inflammatory biomarkers in DSS colitis model.
    Digestive Diseases and Sciences 04/2014; · 2.26 Impact Factor
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    ABSTRACT: Flavanol (flavan-3-ol)-rich lychee fruit extract (FRLFE) is a mixture of oligomerized polyphenols primarily derived from lychee fruit and is rich in flavanol monomers, dimers, and trimers. Supplementation with this functional food has been shown to suppress inflammation and tissue damage caused by high-intensity exercise training. However, it is unclear whether FRLFE has in vitro anti-inflammatory effects, such as suppressing the production of the proinflammatory cytokine tumor necrosis factor α (TNF-α) and the proinflammatory mediator nitric oxide (NO), which is synthesized by inducible nitric oxide synthase (iNOS). Here, we analyzed the effects of FRLFE and its constituents on the expression of inflammatory genes in interleukin 1β (IL-1β)-treated rat hepatocytes. FRLFE decreased the mRNA and protein expression of the iNOS gene, leading to the suppression of IL-1β-induced NO production. FRLFE also decreased the levels of the iNOS antisense transcript, which stabilizes iNOS mRNA. By contrast, unprocessed lychee fruit extract, which is rich in flavanol polymers, and flavanol monomers had little effect on NO production. When a construct harboring the iNOS promoter fused to the firefly luciferase gene was used, FRLFE decreased the luciferase activity in the presence of IL-1β, suggesting that FRLFE suppresses the promoter activity of the iNOS gene at the transcriptional level. Electrophoretic mobility shift assays indicated that FRLFE reduced the nuclear transport of a key regulator, nuclear factor κB (NF-κB). Furthermore, FRLFE inhibited the phosphorylation of NF-κB inhibitor α (IκB-α). FRLFE also reduced the mRNA levels of NF-κB target genes encoding cytokines and chemokines, such as TNF-α. Therefore, FRLFE inhibited NF-κB activation and nuclear translocation to suppress the expression of these inflammatory genes. Our results suggest that flavanols may be responsible for the anti-inflammatory and hepatoprotective effects of FRLFE and may be used to treat inflammatory diseases.
    PLoS ONE 04/2014; 9(4):e93818. · 3.53 Impact Factor
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    ABSTRACT: Nobiletin is a polymethoxylated flavone that is abundant in the peels of citrus fruits, such as Citrus unshiu (Satsuma mandarin) and C. sinensis. The dried peels of C. unshiu (chinpi) have been included in several formulae of Japanese Kampo medicines. Nobiletin may suppress the induction of inducible nitric oxide synthase (iNOS), which synthesizes the inflammatory mediator nitric oxide (NO) in hepatocytes. A Citrus unshiu peel (CUP) extract was prepared. Primary cultured rat hepatocytes were treated with the CUP extract or nobiletin in the presence of interleukin 1β (IL-1β), which induces iNOS expression. NO production and iNOS gene expression were analyzed. High-performance liquid chromatography analyses revealed that the nobiletin content in the CUP extract was 0.14%. Nobiletin dose-dependently reduced the NO levels and decreased iNOS expression at the protein, mRNA and antisense transcript levels. Flavone, which does not contain any methoxy groups, also suppressed iNOS induction. Nobiletin reduced the transcriptional activity of iNOS promoter-luciferase constructs and the DNA-binding activity of nuclear factor κB (NF-κB) in the nuclei. The suppression of iNOS induction by nobiletin suggests that nobiletin may be responsible for the anti-inflammatory effects of citrus peels and have a therapeutic potential for liver diseases.
    Biochemical and Biophysical Research Communications 08/2013; · 2.28 Impact Factor
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    ABSTRACT: http://functionalfoodscenter.net/files/69624283.pdf Background: Active hexose correlated compound (AHCC) is the extract from cultured mycelia of Lentinula edodes, a species of Basidiomycetes mushroom. AHCC contains various polysaccharides, including partially acylated alpha-1,4-glucan, which is one of its major constituents. The application of AHCC has been markedly increased in complementary and alternative medicine as a functional food because AHCC improved the prognosis of postoperative hepatocellular carcinoma patients. AHCC has anti-inflammatory and antioxidant effects, such as the suppression of nitric oxide production in hepatocytes. AHCC might affect resistance to environmental stress, which is assumed to play a pivotal role in the longevity of many organisms. Objective: To investigate the effect of AHCC on longevity, we measured the lifespan of the nematode Caenorhabditis elegans, a model animal that is widely used to assess longevity. We also examined the effect of AHCC on resistance to heat stress, i.e., thermotolerance. Methods: The lifespan of C. elegans animals grown on media in the absence or presence of AHCC at 20°C was evaluated. Thermotolerance assays were performed at 35°C, the restrictive temperature of the animals. The effects of AHCC on lifespan and thermotolerance were analyzed with longevity mutants. Expression levels of stress-related genes, including heat shock genes, were measured by strand-specific reverse transcription-polymerase chain reaction after heat shock. Results: Wild-type C. elegans animals exhibited a longer mean lifespan by up to 10% in the presence of AHCC in the growth media than animals in the absence of AHCC. Furthermore, AHCC markedly increased thermotolerance at 35°C. Epistasis analyses showed that lifespan extension by AHCC at least partly required two longevity-promoting transcription factors: DAF-16 (C. elegans homolog of FOXO) and HSF-1 (C. elegans homolog of heat shock transcription factor 1). After heat shock, AHCC activated the transcription of the heat shock genes, which are the targets of HSF-1. Similarly, the expression of hsf-1 mRNA was elevated following AHCC treatment. Recently, natural antisense transcripts were shown to regulate mRNA stability at the posttranscriptional level. In nematodes, AHCC increased the natural antisense transcript of the hsf-1 gene. Conclusion: AHCC conferred lifespan extension and thermotolerance to C. elegans. Our analyses suggest that the beneficial effects of AHCC on longevity are involved in the activation of at least two transcription factors, DAF-16 and HSF-1, most likely through an antisense transcript-mediated mechanism.
    Functional Foods in Health and Disease. 06/2013; 3(6):166-182.
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    ABSTRACT: AIM: Tumor necrosis factor α (TNF) is a pleiotropic cytokine involved in various inflammatory diseases. The only production of TNF in the liver is thought to be from hepatic macrophages known as Kupffer cells, predominantly in response to bacterial lipopolysaccharide (LPS). METHODS: Primary cultured rat hepatocytes were used to analyze TNF expression in response to the proinflammatory cytokine, interleukin 1β (IL-1β). Livers of rats subjected to LPS-induced endotoxemia were analyzed. RESULTS: Immunocytochemistry and enzyme-linked immunosorbent assays demonstrated that IL-1β-treated rat hepatocytes secreted TNF, and RNA analyses indicated that TNF mRNA was induced specifically by IL-1β. Northern blot analysis showed that not only mRNA, but also a natural antisense transcript (asRNA), was transcribed from the rat Tnf gene in IL-1β-treated hepatocytes. TNF was detected in the hepatocytes of LPS-treated rats. Both TNF mRNA and asRNA were expressed in the hepatocytes of LPS-treated rats, human hepatocellular carcinoma, and human monocyte/macrophage cells. To disrupt the interaction between TNF asRNA and TNF mRNA, sense oligonucleotides corresponding to TNF mRNA were introduced into rat hepatocytes resulting in significantly increased levels of TNF mRNA. One of these sense oligonucleotides increased a half-life of TNF mRNA, suggesting that the TNF asRNA may reduce the stability of TNF mRNA. CONCLUSIONS: IL-1β-stimulated rat hepatocytes are a newly identified source of TNF in the liver. TNF mRNA and asRNA are expressed in rats and humans, and the TNF asRNA reduces the stability of the TNF mRNA. Hepatocytes and TNF asRNA may be therapeutic targets to regulate levels of TNF mRNA.
    Hepatology Research 05/2013; · 2.22 Impact Factor
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    ABSTRACT: Natural antisense transcripts (asRNAs) are frequently transcribed from mammalian genes. Recently, we found that non-coding asRNAs are transcribed from the 3' untranslated region (3'UTR) of the rat and mouse genes encoding inducible nitric oxide synthase (iNOS), which catalyzes the production of the inflammatory mediator nitric oxide. The iNOS asRNA stabilizes iNOS mRNA by interacting with the mRNA 3'UTR. Furthermore, single-stranded 'sense' oligonucleotides corresponding to the iNOS mRNA sequence were found to reduce iNOS mRNA levels by interfering with mRNA-asRNA interactions in rat hepatocytes. This method was named natural antisense transcript-targeted regulation (NATRE) technology. In this study, we detected human iNOS asRNA expressed in hepatocarcinoma and colon carcinoma tissues. The human iNOS asRNA harbored a sequence complementary to an evolutionarily conserved region of the iNOS mRNA 3'UTR. When introduced into hepatocytes, iNOS sense oligonucleotides that were modified by substitution with partial phosphorothioate bonds and locked nucleic acids or 2'-O-methyl nucleic acids greatly reduced levels of iNOS mRNA and iNOS protein. Moreover, sense oligonucleotides and short interfering RNAs decreased iNOS mRNA to comparable levels. These results suggest that NATRE technology using iNOS sense oligonucleotides could potentially be used to treat human inflammatory diseases and cancers by reducing iNOS mRNA levels.
    Nitric Oxide 01/2013; · 3.27 Impact Factor
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    ABSTRACT: Antisense transcription is a widespread phenomenon in the mammalian genome and is believed to play a role in regulating gene expression. However, the exact functional significance of antisense transcription is largely unknown. Here, we show that natural antisense (AS) RNA is an important modulator of interferon-α1 (IFN-α1) mRNA levels. A ~4-kb, spliced IFN-α1 AS RNA targets a single-stranded region within a conserved secondary structure element of the IFN-α1 mRNA, an element which was previously reported to function as the nuclear export element. Following infection of human Namalwa lymphocytes with Sendai virus or infection of guinea pig 104C1 fetal fibroblasts with influenza virus A/PR/8/34, expression of IFN-α1 AS RNA becomes elevated. This elevated expression results in increased IFN-α1 mRNA stability because of the cytoplasmic (but not nuclear) interaction of the AS RNA with the mRNA at the single-stranded region. This results in increased IFN-α protein production. The silencing of IFN-α1 AS RNA by sense oligonucleotides or over-expression of antisense oligoribonucleotides, which were both designed from the target region, confirmed the critical role of the AS RNA in the post-transcriptional regulation of IFN-α1 mRNA levels. This AS RNA stabilization effect is caused by the prevention of the microRNA (miRNA)-induced destabilization of IFN-α1 mRNA due to masking of the miR-1270 binding site. This discovery not only reveals a regulatory pathway for controlling IFN-α1 gene expression during the host innate immune response against virus infection but also suggests a reason for the large number of overlapping complementary transcripts with previously unknown function.
    Cellular and Molecular Life Sciences CMLS 12/2012; · 5.86 Impact Factor
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    ABSTRACT: Gomishi is the dried fruit of Schisandra chinensis Baillon (Fructus Schisandrae chinensis, FSC) and has been used in Japanese Kampo medicine to treat inflammatory and liver diseases. However, it is unclear which constituent of FSC is primarily responsible for its pharmacological effects. FSC was extracted with methanol, fractionated by hydrophobicity, and further purified. We measured the effects of each fraction or constituent thereof on the induction of the inflammatory mediator nitric oxide (NO), which was induced by interleukin 1β in primary cultured rat hepatocytes. The hydrophobic fraction markedly suppressed NO induction and reduced the expression of inducible nitric oxide synthase (iNOS) in interleukin 1β-treated hepatocytes. Gomisin N and γ-schizandrin, two major constituents of the hydrophobic fraction, significantly reduced NO production and the levels of the iNOS protein, mRNA, and antisense transcript. Gomisin N and γ-schizandrin also decreased the transcription of interleukin 1β and inflammatory chemokines. The overexpression of the p65 subunit of nuclear factor κB or CCAAT/enhancer-binding protein β increased the promoter activity of the iNOS gene in the firefly luciferase assay, whereas gomisin N decreased the promoter activity. The anti-inflammatory activity of FSC and its constituents were analysed, and we demonstrated that gomisin N and γ-schizandrin are involved in the hepatoprotective effect of the FSC extract, which has therapeutic potential for liver disease.
    Nitric Oxide 10/2012; · 3.27 Impact Factor
  • Clinical Nutrition Supplements 09/2012; 7(1):65-66.
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    ABSTRACT: Background: Flos Lonicerae japonicae (FLJ; Kinginka) is the dried flowers and buds of the Japanese honeysuckle Lonicera japonica Thunberg. FLJ has been used as a Japanese Kampo medicine to treat infectious and inflammatory diseases. However, it is not clear which constituent of FLJ is responsible for its pharmacological effects. Methods: FLJ was extracted with methanol and fractionated by hydrophobicity. We measured the effects of each fraction on the induction of the inflammatory mediator nitric oxide (NO), which was induced by interleukin 1β in primary cultured rat hepatocytes. To estimate cytotoxicity, the activity of lactate dehydrogenase released from the hepatocytes was measured. The expression of inducible nitric oxide synthase (iNOS) was analyzed by Western blot analysis and reverse transcription-polymerase chain reaction. Results: The methanol extract was fractionated into hydrophobic (11.1%), butanol-soluble (16.4%), and water-soluble fractions (72.5%). These three fractions dose-dependently suppressed the induction of NO and reduced the level of iNOS protein in interleukin 1β-stimulated hepatocytes. Chlorogenic acid, a major constituent of the water-soluble fraction, significantly reduced the levels of NO production, iNOS protein, and iNOS mRNA. Chlorogenic acid also decreased the levels of mRNAs encoding cytokines and chemokines that are involved in inflammatory disease. Caffeic acid, which is formed by the hydrolysis of chlorogenic acid, markedly reduced the induction of NO, although it did not exist at a detectable level in the water-soluble fraction. In contrast, other constituents of the water-soluble fraction, such as inositol fructose, glucose, and sucrose, did not affect the induction of NO. Conclusions: The anti-inflammatory effects of the FLJ extract and its constituents were analyzed by measuring the induction of NO and iNOS in hepatocytes. We demonstrated that chlorogenic acid, one of the main constituents of FLJ, is involved in the anti-inflammatory effect of the FLJ extract, suggesting its therapeutic potential.
    HOAJ Biology. 04/2012; 1(2).
  • Cytokine 10/2011; 56(1):5-5. · 2.87 Impact Factor
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    ABSTRACT: Flavanol-rich lychee fruit extract (FRLFE) is a processed lychee fruit extract that is higher in flavanols (monomers, dimers and trimers) than its unprocessed counterpart. FRLFE exerts antioxidant activities in vitro and is expected to protect against inflammation and tissue damage. However, the physiological effects of FRLFE intake have not been explored in vivo. The aim of this study was to examine the effects of FRLFE supplementation on inflammation and tissue damage in young athletes during intense physical training. Twenty healthy male long-distance runners at a university were randomly assigned to receive FRLFE or placebo in a double-blind manner. Blood and serum parameters associated with inflammation, tissue damage and oxidative stress were evaluated before (pre-training), during (mid-training) and after (post-training) a 2-month training period. Some parameters, including the white blood cell count, were significantly modified by FRLFE supplementation. Compared with the placebo group, the change in the serum interleukin-6 level between pre- and mid-training were significantly lower in the FRLFE group, while the change in the transforming growth factor-β level between pre- and post-training was significantly greater in the FRLFE group. These findings suggest that FRLFE supplementation may suppress inflammation or tissue damage caused by high-intensity exercise training.
    Phytotherapy Research 02/2011; 25(10):1486-93. · 2.40 Impact Factor
  • Clinical Nutrition Supplements 01/2011; 6(1):202-202.
  • Clinical Nutrition Supplements 01/2011; 6(1):136-136.
  • Clinical Nutrition Supplements 01/2010; 5(2):198-199.