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

DOI： doi.org/10.3390/nu14142862

Repository Public URL： http://hdl.handle.net/2324/5208865

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ポリフェノールフィセチンが、育毛促進効果を示した。

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GABAが腸脳相関を活性化する分子メカニズムを明らかにした。

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DOI： https://doi.org/10.22146/ijc.53411

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カルノシンがエクソソームを介して腸脳相関を活性化する分子基盤を明らかにした。

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血中βアラニンが認知症発症のマーカーになりうることを明らかにした。

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

皮膚表皮細胞における長寿遺伝子SIRT1を活性化するポリフェノールが、紫外線により誘導されるDNA損傷を修復した。

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独自に同定した細胞老化因子SMARCD1が、肝臓での脂肪滴合成に関わることを明らかにした。

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DOI： 10.1093/jb/mvw038

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ポリフェノールが、SIRT3の活性化を通じて活性酸素種を抑制することを示した。

DOI： 10.1155/2016/2927131

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

乳酸菌T2102株が腸管上皮細胞でのSIRT1発現の増強を通じて、がんを抑制することを見出した。

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

イミダゾールジペプチド高含有エキスを用いて、中高齢者を対象としたランダム化群間比較二重盲検試験を実施した。その結果、イミダゾールジペプチド摂取群において、言語性記憶の低下抑制効果（記録力の維持）が確認された。

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細胞老化因子として独自に同定したATP6V0A2が、ゴルジ体の生合成の維持を介して細胞老化に関わっていることを明らかにした。

DOI： 10.1038/srep17342

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

DOI： 10.1016/j.jff.2014.12.024

Language：Japanese   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s10616-015-9849-x

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DOI： 10.1093/jb/mvu040

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DOI： 10.1016/j.jff.2014.07.006

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長寿遺伝子FOXO3aが、cーmyc依存的にテロメラーゼを増強することを分子レベルで明らかにした。

DOI： 10.1371/journal.pone.0101864

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DOI： 10.1016/j.imbio.2013.08.011

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DOI： 10.1007/s10616-013-9596-9

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抗体精製のために用いられるProtein Aに変わる、新たな温度感受性のProtein Aの開発に関わる研究。

DOI： 10.1016/j.chroma.2013.07.015

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DOI： 10.1007/s10616-010-9276-y

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Language：English   Publisher：Nutrients  

Background: Urolithin A (Uro-A), a type of polyphenol derived from pomegranate, is known to improve memory function when ingested, in addition to its direct effect on the skin epidermal cells through the activation of longevity gene SIRT1. However, the molI ecular mechanism by which orally ingested Uro-A inhibits cognitive decline via the intestine remains unexplored. Objectives: This study aimed to evaluate the role of Uro-A in improving cognitive function via improved intestinal function and the effect of Uro-A on the inflammation levels and gene expression in hippocampus. Methods: Research to clarify the molecular basis of the functionality of Uro-A was also conducted. Results: The results demonstrated that Uro-A suppressed age-related memory impairment in Aged mice (C57BL/6J Jcl, male, 83 weeks old) by reducing inflammation and altering hippocampal gene expression. Furthermore, exosomes derived from intestinal cells treated with Uro-A and from the serum of Aged mice fed with Uro-A both activated neuronal cells, suggesting that exosomes are promising candidates as mediators of the Uro-A-induced activation of gut–brain interactions. Additionally, neurotrophic factors secreted from intestinal cells may contribute to the Uro-A-induced activation of gut–brain interactions. Conclusions: This study suggests that Uro-A suppresses age-related cognitive decline and that exosomes and other secreted factors may contribute to the activation of the gut–brain interaction. These findings provide new insights into the therapeutic potential of Uro-A for cognitive health.

DOI： 10.3390/nu16193369

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Springer Science and Business Media LLC  

Abstract

In filamentous fungi, microtubules are important for polar growth and morphological maintenance and serve as rails for intracellular trafficking. The molecular mechanisms associated with microtubules have been analyzed. However, little is known about when and where tubulin, a component of microtubules, is biosynthesized in multinuclear and multicellular filamentous fungi. In this study, we visualized microtubules based on the enhanced green fluorescence protein (EGFP)-labeled α-tubulin and β-tubulin mRNA tagged by the EGFP-mediated MS2 system in living yellow Koji mold Aspergillus oryzae cells in order to understand the spatiotemporal production mechanism of tubulin. We found that mRNA of btuA, encoding for β-tubulin, localized at dot-like structures through the apical, middle and basal regions of the hyphal cells. In addition, some btuA mRNA dots showed microtubule-dependent motor protein-like dynamics in the cells. Furthermore, it was found that btuA mRNA dots were decreased in the cytoplasm just before mitosis but increased immediately after mitosis, followed by a gradual decrease. In summary, the localization and abundance of β-tubulin mRNA is spatiotemporally regulated in living A. oryzae hyphal cells.

DOI： 10.1038/s41598-024-64531-5

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Other Link： https://www.nature.com/articles/s41598-024-64531-5

Publishing type：Research paper (scientific journal)   Publisher：IEEE Transactions on Dielectrics and Electrical Insulation  

In this study, the first to investigate the potential of using extremely low-frequency electric fields (ELFEFs) to improve osteoporosis, we examined the effects of ELFEF exposure on bone formation using osteoblast-like MC3T3-E1 cells. We first constructed a system to regulate the intensity of the ELFEF exposure. Next, we used this system to examine the effect of ELFEF exposure on the proliferation of undifferentiated and differentiated MC3T3-E1 cells. ELFEF exposure increased calcium deposition in an intensity-dependent manner. In addition to increasing alkaline phosphatase (ALP) activity and collagen production, ELFEF exposure also stimulated mRNA expression of osteoblast markers Col1a1, Alpa, Sp7, and Dlx5 in an intensity-dependent manner. In contrast, ELFEF exposure did not enhance RANKL expression. These results indicate that ELFEF exposure has potential for application in the prevention and/or improvement of bone fragility.

DOI： 10.1109/TDEI.2024.3374241

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

Fructobacillus is a lactic-acid bacterium recently identified in fructose-rich environments. Fructobacillus is also known to exhibit unusual growth characteristics due to an incomplete gene encoding alcohol/acetaldehyde hydrogenase, which results in an imbalance in the nicotinamide adenine mononucleotide (NAD+)/NADN levels. Recently, the addition of d-fructose to the culture medium of Fructobacillus strains increased the intracellular nicotinamide mononucleotide (NMN) content. In the present study, we evaluated the functionality of Fructobacillus that produces high levels of NMN, using one substrain (Fructobacillus fructosus OS-1010). Therefore, in this study, we examined its functionality in the interaction between intestinal cells and muscle cells. The results showed that supernatant derived from intestinal epithelial cells (Caco-2 cells) treated with F. fructosus OS-1010 activated muscle cells (C2C12 cells). Further analysis revealed that Caco-2 cells treated with F. fructosus OS-1010 secreted exosomes known as extracellular vesicles, which activated the muscle cells. Furthermore, pathway analysis of the target genes of miRNA in exosomes revealed that pathways involved in muscle cell activation, including insulin signaling and cardiac muscle regulation, neurotrophic factors, longevity, and anti-aging, can be activated by exosomes. In other words, F. fructosus OS-1010 could activate various cells such as the skin and muscle cells, by secreting functional exosomes from the intestinal tract.

DOI： 10.1007/s10616-023-00610-1

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

The number of patients with allergies to pollen and food is increasing worldwide. In Japan, the prevalence of cedar pollinosis, a type I allergy, is nearly 30% and accounts of hay fever are rising. A potential natural remedy for these allergic diseases may be Hita Tenryo Water™ (referred to simply as Hita Tenryo water), water that is pumped from deep underground in the Hita region of Oita, Japan, which has been the subject of various research reports. Here, we investigated the potential of using Hita Tenryo water to suppress the onset of cedar pollinosis in a mouse model and explored the immunological mechanism of the suppression. Test model mice were given Hita Tenryo water ad libitum to drink and received intraperitoneal administration of (i) tap water (Hw1), (ii) 25% Hita Tenryo water (Hw2) or (iii) 100% Hita Tenryo (Hw3). There were no significant differences in body weight change, feed intake, or water intake among the groups during the experimental period. We examined nose rubbing and sneezing as allergic symptoms. The frequency of rubbing and sneezing tended to decrease in the Hw1 and Hw2 group, and significantly decreased in the Hw3 group compared to control. Total IgE levels in serum were also significantly reduced in Hita Tenryo water intraperitoneal administration groups. In vitro examination of the rate of release of β-hexosaminidase from BL-2H3 cells showed that there were no significantly differences between Hita Tenryo water-treated and control cells. In addition, measurement of Th2-related cytokine levels in concanavalin A-stimulated peripheral blood mononuclear cells revealed a significant decrease in IL-4, IL-6, and IL-10 levels in medium (p < 0.01). In contrast, production of IFN-γ significantly increased (p < 0.01). These results indicate that Hita Tenryo water may alleviate and/or suppress allergic symptoms.

DOI： 10.1016/j.heliyon.2024.e26915

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Bioscience, Biotechnology and Biochemistry  

Scalp bacteria on the human scalp and scalp hair comprise distinct community structures for sites and individuals. To evaluate their effect on human keratinocyte cellular activity, including that of the hair follicular keratinocytes, the expression of several longevity genes was examined using HaCaT cells. A screening system that uses enhanced green fluorescent protein ( EGFP ) fluorescence was established to identify scalp bacteria that enhance silent mating type information regulation 2 homolog-1 ( SIRT1 ) promoter activity in transformed HaCaT cells ( SIRT1p-EGFP ) . The results of quantitative polymerase chain reaction revealed that several predominant scalp bacteria enhanced ( Cutibacterium acnes and Pseudomonas lini ) and repressed ( Staphylococcus epidermidis ) the expressions of SIRT1 and telomerase reverse transcriptase ( TERT ) genes in HaCaT cells. These results suggest that the predominant scalp bacteria are related to the health of the scalp and hair, including repair of the damaged scalp and hair growth, by regulating gene expression in keratinocytes. Graphical Abstract : Schematic sketch of SIRT1 and TERT gene expression in hair follicle keratinocytes by scalp bacteria.

DOI： 10.1093/bbb/zbad122

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

Hericium erinaceus, a mushroom species commonly known as Yamabushitake in Japan, is known to have a stimulatory effect on neurotrophic factors, such as brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF). Hericenone C, a meroterpenoid with palmitic acid as the fatty acid side chain, is reported to be one such stimulant. However, according to the structure of the compound, the fatty acid side chain seems highly susceptible to lipase decomposition, under in vivo metabolic conditions. To study this phenomenon, hericenone C from the ethanol extract of the fruiting body was subjected to lipase enzyme treatment and observed for changes in the chemical structure. The compound formed after the lipase enzyme digestion was isolated and identified using LC-QTOF-MS combined with 1H-NMR analysis. It was found to be a derivative of hericenone C without its fatty acid side chain and was named deacylhericenone. Interestingly, a comparative investigation of the neuroprotective properties of hericenone C and deacylhericenone showed that the BDNF mRNA expression in human astrocytoma cells (1321N1) and the protection against H2O2-induced oxidative stress was considerably higher in the case of deacylhericenone. These findings suggest that the stronger bioactive form of the hericenone C compound is in fact deacylhericenone.

DOI： 10.3390/molecules28114549

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

Carnosine is known to improve brain function. The molecular basis for the carnosine-mediated interaction between intestinal cells and neuronal cells is that carnosine acts on intestinal cells and stimulates exosome secretion, which can induce neurite outgrowth in neuronal cells. This study aimed to infer the carnosine-mediated interaction between muscle cells and neuronal cells. The results revealed that carnosine induces muscle cell differentiation, as well as the secretion of exosomes and myokines that can act on neuronal cells. Carnosine acts not only on intestinal cells but also on muscle cells, stimulating the secretion of secretory factors including exosomes that induce neurite outgrowth in neuronal cells, as well as myokines known to be involved in neuronal cell activation. As the miRNAs in exosomes secreted from intestinal cells and muscle cells upon carnosine treatment are different, it could be assumed that carnosine acts on each cell to interact with neuronal cell through separate factors and mechanisms.

DOI： 10.3390/nu15061479

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Language：Japanese   Publisher：フレグランスジャーナル社  

Language：English   Publisher：MDPI (Multidisciplinary Digital Publishing Institute)  

Regulatory T cells (Tregs) and CD4+/CD25+ T cells play an important role in the suppression of excessive immune responses, homeostasis of immune function, and oral tolerance. In this study, we screened for food-derived polyphenols that induce Tregs in response to retinaldehyde dehydrogenase (RALDH2) activation using macrophage-like THP-1 cells. THP-1 cells were transfected with an EGFP reporter vector whose expression is regulated under the control of mouse Raldh2 promoter and named THP-1 (Raldh2p-EGFP) cells. The THP-1 (Raldh2p-EGFP) cells were treated with 33 polyphenols after inducing their differentiation into macrophage-like cells using phorbol 12-myristate 13-acetate. Of the 33 polyphenols, five (kaempferol, quercetin, morin, luteolin and fisetin) activated Raldh2 promoter activity, and both quercetin and luteolin activated the endogenous Raldh2 mRNA expression and enzymatic activity. Furthermore, these two polyphenols increased transforming growth factor beta 1 and forkhead box P3 mRNA expression, suggesting that they have Treg-inducing ability. Finally, we verified that these polyphenols could induce Tregs in vivo and consequently induce IgA production. Oral administration of quercetin and luteolin increased IgA production in feces of mice. Therefore, quercetin and luteolin can induce Tregs via RALDH2 activation and consequently increase IgA production, suggesting that they can enhance intestinal barrier function.

CiNii Research

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

We examined the association of serum s-adenosylmethionine (SAM), s-adenosylhomocysteine (SAH) (methionine metabolites), and their ratio on the risk of dementia and death in a community-dwelling population of older Japanese individuals. 1371 residents of Hisayama, Japan, aged 65 years or older and without dementia, were followed for a median of 10.2 years (2007-2017). We divided serum SAM, SAH, and SAM/SAH ratio into quartiles. Cox proportional hazards models were used to estimate the hazard ratios (HRs) and their 95% confidence intervals (CIs) of serum SAM, SAH, and SAM/SAH ratio levels on the risk of a composite outcome of all-cause dementia or death, and each outcome. During the follow-up, 635 participants developed all-cause dementia and/or died, of which 379 participants developed dementia and 394 deaths occurred. The multivariable-adjusted HRs of the composite outcome decreased significantly with increasing serum SAM levels (P for trend = 0.01), while they increased significantly with higher serum SAH levels (P for trend = 0.03). Higher serum SAM/SAH ratio levels were significantly associated with a lower risk of the composite outcome (P for trend = 0.002), as well as with lower risk of each outcome. Our findings suggest that the balance of methionine metabolites may closely associate with the risk of dementia and death.

DOI： 10.1038/s41598-022-16242-y

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

Regulatory T cells (Tregs) and CD4+/CD25+ T cells play an important role in the suppression of excessive immune responses, homeostasis of immune function, and oral tolerance. In this study, we screened for food-derived polyphenols that induce Tregs in response to retinaldehyde dehydrogenase (RALDH2) activation using macrophage-like THP-1 cells. THP-1 cells were transfected with an EGFP reporter vector whose expression is regulated under the control of mouse Raldh2 promoter and named THP-1 (Raldh2p-EGFP) cells. The THP-1 (Raldh2p-EGFP) cells were treated with 33 polyphenols after inducing their differentiation into macrophage-like cells using phorbol 12-myristate 13-acetate. Of the 33 polyphenols, five (kaempferol, quercetin, morin, luteolin and fisetin) activated Raldh2 promoter activity, and both quercetin and luteolin activated the endogenous Raldh2 mRNA expression and enzymatic activity. Furthermore, these two polyphenols increased transforming growth factor beta 1 and forkhead box P3 mRNA expression, suggesting that they have Treg-inducing ability. Finally, we verified that these polyphenols could induce Tregs in vivo and consequently induce IgA production. Oral administration of quercetin and luteolin increased IgA production in feces of mice. Therefore, quercetin and luteolin can induce Tregs via RALDH2 activation and consequently increase IgA production, suggesting that they can enhance intestinal barrier function.

DOI： 10.3390/nu14142862

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Language：Japanese   Publisher：(公社)日本精神神経学会  

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

Cistus L. is a genus of dicotyledonous perennial herbaceous plants. Cistus species have been commonly used in folk medicine in the Mediterranean region. In the present study, the biological activities of essential oils derived from Cistus species (Cistus laurifolius, C. monspeliensis, C. creticus, and C. salviifolius) were evaluated. Essential oils derived from C. laurifolius and C. monspeliensis were found to augment the expression of SIRT1, an anti-aging gene, in the normal culture of HaCaT cells. Furthermore, these essential oils increased the number and size of mitochondria and augmented their activity. These effects were thought to be caused by the up- and downregulated expression of MITOL and Drp1 in HaCaT cells, respectively, in response to the essential oil treatment. In addition, these essential oils were found to attenuate ultraviolet-B-induced mitochondrial damage and cellular senescence in HaCaT cells. These findings indicate that essential oils derived from C. laurifolius and C. monspeliensis may inhibit skin aging through mitochondrial regulation via SIRT1 activation.

DOI： 10.3390/molecules27072053

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

This study aimed to investigate the changes in B cell functional decline and antigen sensitization with aging using two Epstein Barr virus (EBV)-immortalized human B cell lines, one from a 22-year-old man (EBV-B young) and the other from a 65-year-old man (EBV-B old). The activity of senescence-associated β-galactosidase, a marker of cellular senescence, was enhanced in the EBV-B old cells compared with EBV-B young cells. Moreover, the levels of p16, p21, IL-6, TNF-α, and TGF-β1, which are senescence-associated secretary phenotypes, were also increased in EBV-B old cells. In vitro immunization of EBV-B cells with β-lactoglobulin further showed that EBV-B old cells had a reduced cell population of naïve B cells than that of EBV-B young cells. Furthermore, HLA-DR expression, which is important for antigen presentation, was decreased in the EBV-B old cells. Comparative microarray analysis between EBV-B young and old cells also showed decreased expression of antibody genes, such as those of the heavy chain and light chain (κ chain). These results suggest that cellular senescence and decreased gene expression are responsible, at least in part, for the decline in B cell function and antigen sensitization capacity with aging, which ultimately impairs the function of the acquired immune system.

DOI： 10.1007/s10616-021-00513-z

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GABAにより分泌誘導された腸管由来エクソソームが脳細胞を活性化することを明らかにした。

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

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

ポリフェノールフィセチンが、皮膚表皮由来のエクソソーム分泌を誘導し、毛包幹細胞を活性化させ、育毛を誘導することを明らかにした。

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DOI： 10.1155/2014/19305

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DOI： 10.1016/j.bbrc.2013.10.007

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DOI： 10.1016/j.bbrc.2005.06.109

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DOI： 10.1007/s10616-005-3761-8

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DOI： 10.1007/s10616-005-3759-2

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DOI： 10.1007/s10585-005-3973-5

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DOI： 10.1271/bbb.68.2279

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DOI： 10.1271/bbb.68.1591

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DOI： 10.1023/B:CYTO.0000039901.92984.7a

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An aqueous extract of Kefir, fermented milk originally produced in the Caucasus mountains, suppressed morphological changes of human melanoma HMV-1 and SK-MEL cells and human normal fibroblast TIG-1 cells caused by UVC-irradiation, suggesting that UV damage can be suppressed by the Kefir extract. The addition of the Kefir extract after UVC-irradiation of HVM-1 cells resulted in a remarkable decrease in intracellular reactive oxygen species (ROS) which had been increased by UVC irradiation. The Kefir extract also stimulated unscheduled DNA synthesis and suppressed UVC-induced apoptosis of HMV-1 cells. A colony formation assay revealed that the Kefir extract rescued HMV-1 cells from cell death caused by UVC irradiation. The Kefir extract, as well as methyl methanethiosulfonate which is known to enhance the nucleotide excision repair (NER) activity, exhibited strong thymine dimer repair-enhancing activity. Epigalocatechin exhibited a weak NER activity but vitamins A, C, and E and catechin showed no NER activity. The thymine dimer repair-enhancing factors in the Kefir extract were heatstable and assumed to be molecules with a molecular weight of less than 5000. The treatment of HMV-1 cells with the Kefir extract during or before UVC-irradiation also prevented the generation of ROS and thymine dimmer, and suppressed the apoptosis of HMV-1 cells, suggesting that application of Kefir can prevent UV damage.

DOI： 10.1023/A:1023984304610

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DOI： 10.1023/A:1023918206060

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DOI： 10.1023/A:1023926407877

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DOI： 10.1023/A:1023936421448

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DOI： 10.1023/A:1008179919857

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DOI： 10.1023/A:1008129020728

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DOI： 10.1023/A:1008179302906

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

DOI： 10.1006/bbrc.1999.0129

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

DOI： 10.1143/JJAP.38.L1102

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

DOI： 10.1023/A:1008048928053

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

DOI： 10.1023/A:1008016020785

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

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

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

DOI： 10.1271/bbb.63.1624

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

DOI： 10.1016/S0167-4838(99)00099-0

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

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

Human HB4C5 hybridoma cells produce a lung cancer-specific IgM human monoclonal antibody (mAb). HB4C5 human mAb cross-reacts with Candida cytochrome c (Cyt c) and carboxypeptidase (Cpase). Concanavalin A (ConA)-resistant variants of HB4C5 cells loss the original light chain followed by expression of various new light chains at a high incidence (light chain shifting) (Tachibana et al., 1996). HTD8 cells, one of the ConA-resistant variant subclones of HB4C5 cells, undergo the active light chain shifting and produce various sublines, each of which stably secretes new mAb consisting of a new light chain and a HB4C5 heavy chain. The new mAb exhibits altered antigen binding ability from that of the original antibody. We could expect that HTD8 cells can be used as 'a light chain stem cell line' to improve antigen binding ability and specificity of established human mAbs. A BD9D12 IgG human mAb recognizes lung cancer cells and cross-reacts with cytokeratin 8. Introduction of the heavy chain gene of BD9D12 mAb into HTD8 cells resulted in establishment of various sublines which secreted various kinds of hybrid antibodies consisting of different light chains derived from HTD8 subclones which underwent light chain shifting and a common IgG heavy chain derived from BD9D12. These hybrid antibodies exhibited different or improved reactivities to Cyt, Cpase, cytokeratin 8 and various cancer cells from those of parental mAb, demonstrating that light chain shifting can be applied to improve the affinity and specificity of human mAb.

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

High level and inducible production of human interleukin 6 (hIL-6) was achieved using a novel expression system in Chinese hamster ovary (CHO) cells. In this system, the transcription of hIL-6 gene under the control of PhCMV*-1 promoter composed of tetracycline operator sequences and a minimal promoter is activated by a chimeric transactivator (tTA) composed of tetracycline repressor and transactivating domain of VP16 protein of herpes simplex virus. The transcription of tTA gene, which is also under the control of PhCMV*-1 promoter, is activated by itself via a positive feedback cycle. The expression of both genes is further enhanced by potentiating the VP16 transactivating domain of tTA transactivator with pX protein of hepatitis B virus. In the presence of tetracycline, the tTA transactivators can not bind to PhCMV*-1 promoter, therefore, the expression of hIL-6 and tTA gene is suppressed, and the pX will not activate basal transcription. In the absence of tetracycline, tTA transactivators bind to PhCMV*-1 promoter and activate efficient transcription of hIL-6 and tTA gene, and the transcription is further enhanced by pX via VP16 transactivating domain. Using this strategy, we isolated a clone (UX1) producing hIL-6 at a rate about 1425 ng/10⁶ cells/day. Furthermore, the hIL-6 production is stringently regulated by tetracycline. This results suggested a novel strategy to establish highly efficient, inducible and cell type independent recombinant protein production system by using an artificial promoter to recruit transactivators and coactivators which can synergistically activate transcription.

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

An autocatalytic inducible mammalian expression system was established. This system is composed of two sets of vectors: one carries a selectable marker gene and the other carries a bicistronic expression unit consisting of a target gene, an internal ribosomal entry site, and a tetracycline- controlled transactivator gene. Both the selectable marker and the bicistronic unit are controlled by the tetracycline-responsive promoter (PhCMV(*)-1). When the two vectors are cotransfected into host cells, only a single selection round in the absence of tetracycline is necessary to generate clones expressing the target gene. The expression level is high and easily regulated by tetracycline. Combination with a gene amplification system may allow further enhance the foreign gene expression. Because the PhCMV-1 promoter is relatively independent of cellular regulation signals, this system is expected to work in a wide variety of cell types.

DOI： 10.1006/abio.1997.2325

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

Active oxygen species or free radicals are considered to cause extensive oxidative damage to biological macromolecules, which brings about a variety of diseases as well as aging. The ideal scavenger for active oxygen should be 'active hydrogen'. 'Active hydrogen' can be produced in reduced water near the cathode during electrolysis of water. Reduced water exhibits high pH, low dissolved oxygen (DO), extremely high dissolved molecular hydrogen (DH), and extremely negative redox potential (RP) values. Strongly electrolyzed-reduced water, as well as ascorbic acid, (+)-catechin and tannic acid, completely scavenged O
₂
̇
^-
produced by the hypoxanthine-xanthine oxidase (HX-XOD) system in sodium phosphate buffer (pH 7.0). The superoxide dismutase (SOD)-like activity of reduced water is stable at 4°C for over a month and was not lost even after neutralization, repeated freezing and melting, deflation with sonication, vigorous mixing, boiling, repeated filtration, or closed autoclaving, but was lost by opened autoclaving or by closed autoclaving in the presence of tungsten trioxide which efficiently adsorbs active atomic hydrogen. Water bubbled with hydrogen gas exhibited low DO, extremely high DH and extremely low RP values, as does reduced water, but it has no SOD-like activity. These results suggest that the SOD-like activity of reduced water is not due to the dissolved molecular hydrogen but due to the dissolved atomic hydrogen (active hydrogen). Although SOD accumulated H
₂
O
₂
when added to the HX-XOD system, reduced water decreased the amount of H
₂
O
₂
produced by XOD. Reduced water, as well as catalase ascorbic acid, could directly scavenge H
₂
O
₂
. Reduced water suppresses single-strand breakage of DNA by active oxygen species produced by the Cu(II)-catalyzed oxidation of ascorbic acid in a dose-dependent manner, suggesting that reduced water can scavenge not only O
₂
̇
^-
and H
₂
O
₂
, but also
¹
O
₂
and
^.
OH.

DOI： 10.1006/bbrc.1997.6622

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

We have succeeded in immortalizing human lymphocytes derived from the peripheral blood of a healthy donor and of an atopic patient, and from the lymph node of a cancer patient by oncogene transfection (Alam et al., 1996). All immortalized human lymphocytes were shown to be CD3⁺ and CD19^-, indicating that these immortalized human lymphocytes were all T cells. We established 317, 154 and 692 individual immortalized human T cell lines derived from the healthy donor, the atopic patient and the cancer patient, respectively. The ratios of CD4⁺ and CD8⁺ subpopulations within the set containing immortalized T cells derived from the healthy donor were shown to be varied depending on the combinations of transfected oncogenes used. However, CD8⁺ cells were found to be the dominant subpopulation of immortalized T cells derived from the atopic patient and the cancer patient. These immortalized T cells showed different proliferative responses in the presence of exogenous human IL-2 depending on their origin, and was consistent with the surface expression of the IL-2 receptor. Furthermore, the cytokine secretion patterns of these immortalized T cells stimulated with mitogen were investigated. The results showed that the immortalized T cells from the healthy donor is able to secrete various kinds of cytokines such as IL-2, IL-10, γ-IFN and GM-CSF. However, immortalized T cells from the cancer patient was shown to only secrete IL-2 and GM-CSF. These results suggest that depending on the origin, the immortalized T cells came from different subsets or from cells in different activated states. Mixed lymphocytes reactions demonstrated that these immortalized T cells are able to proliferate in the presence of allogenic or xenogenic stimulator cells, suggesting that they maintain the ability to recognize specific antigens on the stimulator cells and can proliferate even after the immortalization. Furthermore, immortalized T cells derived from the healthy donor and the cancer patient strongly responded to K562 cells, suggesting that MHC-nonrestricted killer T cells were also immortalized.

DOI： 10.1023/a:1007928021139

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

We purified recombinant bovine β-lactoglobulin (rβ-LG) from the culture supernatant of transformed yeast and investigated whether rβ-LG maintained the functional ability and antigenicity of native β-LG. Immunostaining following gel electrophoresis and reversed-phase high-performance liquid chromatography confirmed that rβ-LG was purified homogeneously. rβ-LG showed almost the same retinol-binding ability as native β-LG purified from bovine milk. However, affinities of two anti-β-LG monoclonal antibodies (mAbs) to rβ-LG were different from those to native β-LG, although three other mAbs bound these two proteins equally. Since our panel of five mAbs has been previously shown to be able to detect structural changes occurring in β-LG, this variance in antigenicity can be attributed to conformational differences between rβ-LG and native β-LG. Then, we studied which step in the production and purification procedure was responsible for altering the antigenicity of rβ-LG. Bovine milk native β-LG was added to several steps in this procedure and purified in the same manner as rβ-LG. The results suggested that incubation in the yeast culture had adverse effects on maintaining the antigenicity of this recombinant protein. We conclude from these results that even if no difference between the native and recombinant proteins can be detected by functional analysis, some subtle conformational change which can be distinguished by mAbs may be incorporated into the recombinant protein during its production and ultimately cause a different immune reaction in vivo.

DOI： 10.1023/a:1007977709348

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

A new disease of epidemic proportions caused by fish viruses within the Iridoviridae family inflicts serious damage on red sea breams (Pagrus major) and striped jack (Caranx delicatissimus) populations grown in aquacultures in Japan. A partial segment of the fish iridoviral DNA was directly amplified using the polymerase chain reaction (PCR) with synthetic primers designed from well conserved nucleotide sequences between the frog virus 3 (Ranavirus) and the silkworm iridescent virus type 6. The deduced amino acid sequence from the nucleotide sequence of the PCR fragment demonstrates a high correlation with a partial sequence from the frog virus 3. Using the PCR method with specific primers, we could detect three of four different known types of fish iridoviruses in diseased fishes. To construct more reliable detection methods specific for this viral family, DNA fragments which can specifically hybridize with all of the four known iridoviridae viral DNAs were screened from the genomic library of one iridoviridae strain. The hybridization assay, using a specific fragment which contains regions which are highly homologous with a characterized partial sequence from the frog virus 3, proved to be a reliable diagnostic tool for fish iridoviral diseases.

DOI： 10.1023/a:1007966325278

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

The signal transduction from insulin to its receptors and Ras has been extensively studied, while little has been reported beyond these steps. We found that the expression of human interleukin 6 gene under the control of immediate early gene promoter of human cytomegalovirus was enhanced by insulin stimulation in Chinese hamster ovary cells. The induction effect of insulin was not significantly affected by inhibitors or activators of conventional protein kinase C, cAMP dependent protein kinase and phosphoinositide -3 kinase, however, pre-incubation of the cells with D609, a specific inhibitors of phosphatidylcholine-specific phospholipase C completely abolished the induction effect. These results clearly demonstrate that phosphatidylcholine-specific phospholipase C is a key molecule mediating insulin-induced enhancement of hIL-6 expression from the human cytomegalovirus promoter in Chinese hamster ovary cells and strongly suggest that it plays an important role in the insulin signaling pathways.

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

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

In today's 'modern' society, no one can escape from the stresses of daily life. Stress stimulates the secretion of stress hormones (e.g. cortisol or noradrenaline) which generally suppress the immune response system, thus rendering the body vulnerable to infectious diseases and cancer. Therefore finding anti-stress food components, which diminish and/or inhibit the stress related suppression of the immune response system would be helpful in maintaining and promoting the health of the human population. Here we established a screening system for anti-stress substances using the cultured human cell line MG-63. The production of interferon-β (IFN-β) by MG-63 cells super-induced by Poly (I): Poly (C) was shown to decrease in a dose dependent manner upon the addition of 0.01-10 μg/ml of cortisol or noradrenaline (NA). 1,2-Diacylglycerol (DG) was demonstrated to abrogate this suppression. Lipid from the fermented milk, kefir, also inhibited the influence of cortisol. Kefiran, a polysaccharide secreted from L. kafiranofasiens GKL-28 diminished the cortisol or NA influenced IFN-β production. But phosphatidylcholine had no significant effect in this system. These results suggest that DG, lipids from kefir and kefiran may be equated as anti-stress food component.

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

A subset of type 2, but not type 1, CD4 T cell clones expresses IL-3R and can be stimulated by IL-3. Expression of IL-3R on these type 2 T cell clones is induced by TCR stimulation, and subsequent stimulation by IL-3 augmented the proliferation of and IL-4 production by these cells. This augmented response is inhibited by anti-IL-4 mAb, suggesting the involvement of IL-4 in this response. In place of TCR stimulation, treatment of these type 2 CD4 T cell clones with PMA rendered them responsive to further stimulation of proliferation by IL-3, indicating the cooperation between the IL-3R-elicited signals and PKC-mediated signals in stimulating proliferation. Although the augmentation of the TCR-mediated proliferative response by IL-3 was mainly due to the increased production of IL-4, we also demonstrated the presence of IL-4-independent mechanism mediating the response to IL-3. In situ, we found that splenic T cells could be induced to respond to IL-3 by TCR stimulation. Thus, IL-3 can stimulate a specific population of T cells and influence the immune response.

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

A ras oncogene-amplified recombinant BHK-21 cell line (ras-rBHK-IgG) has been established, and was shown to hyperproduce the recombinant IgG chimeric human monoclonal antibody (hMAb) AE6F4, which recognizes lung cancer cells. We found that the ras-rBHK-IgG cells could be easily cultured in a protein-free ERDF medium supplemented with iron(III) nitrate, hydroxyethyliminodiacetic acid, and non-protein synthetic attachment factor as well as in a serum-free ERDF medium supplemented with insulin, transferrin, ethanolamine, and sodium selenite. The productivity of recombinant hMAb from the cells cultured in dishes at high cell densities was higher in protein-free medium than in serum-containing medium.True high density culture of the ras-rBHK-IgG cells was done in protein-free medium using the Tecnomouse, which is a novel hollow fiber bioreactor system. After culture for 30 days in protein-free culture, a total amount of about 14 mg of the recombinant hMAb AE6F4 was obtained, and was shown to be reactive against lung cancer cells in tissues.

DOI： 10.1271/bbb.60.811

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

DOI： 10.1007/BF02634252

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

We previously determined the amino acid sequence to the epitope (ATLFKTR) of cytochrome c from Candida krusei, which is cross-reactive to the lung cancer-specific human monoclonal antibody HB4C5. Here we report that an antigen messenger RNA, which codes for a structure similar to the cytochrome c epitope, is expressed in the human lung adenocarcinoma A549. Sequencing analysis has revealed that this messenger RNA encodes a novel 190 amino acid polypeptide of 21-kDa containing an amino acid sequence (ALLFFT) similar to the cytochrome c epitope, although the total messenger RNA sequence is apparently different from the cytochrome c messenger RNA. Western analysis indicated that an antibody-recognizable 21-kDa antigen which has the same molecular weight as the predicted polypeptide is expressed in the A549 adenocarcinoma. The in vitro translated product of the antigen messenger RNA and synthesized ALLFFT peptide were both shown to be reactive with the monoclonal antibody, indicating that this protein contains the epitope which enables A549 cells to specifically react with the antibody. The antigen mRNA was not expressed in non-transformed fibroblasts, suggesting that the antigen mRNA expression was associated with cellular transformation. Also in part of the antigen nucleotide sequence, there was a segment that had about 90% homology to the long terminal repeat sequence (no. 297–475) of the human endogenous retrovirus HERV-K10, which was related to the mouse mammary tumor virus.

DOI： 10.1007/BF02634095

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

The human monoclonal antibody AE6F4 specifically reacts with human lung cancer tissues but does not with normal tissues. This monoclonal antibody recognizes a cytosolic 31 kDa antigen in the cancer cells. In a previous study, we elucidated that the 31 kDa antigen belonged to a family of proteins collectively designated as 14-3-3 proteins, which were known as protein kinase-dependent activators of tyrosine/trytophan hydroxylases, or protein kinase C inhibitor proteins. Here we report molecular cloning of the 31 kDa antigen from the human lung adenocarcinoma cell line, A549. Sequencing analysis indicates that the cloned cDNA is identical to that of previously reported human placental cytosolic phospholipase A₂ (cPLA₂), which is also a member of the 14-3-3 protein family. Western analysis demonstrated that a 31 kDa recombinant cPLA₂ expressed in monkey COS cells was recognized by the AE6F4 monoclonal antibody. Binding of the monoclonal antibody to the recombinant cPLA₂ was abolished when treated with sodium periodate, suggesting that not only are carbohydrate chains associated with the cPLA₂, but they also play a crucial role in antigen recognition by the monoclonal antibody.

DOI： 10.1007/BF00749397

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

Residue 19 of tryptophan in bovine β-lactoglobulin (β-LG) is the only invariant residue throughout the lipocalin superfamily having two characteristic features: binding ability for small hydrophobic molecules and the unique β-barrel three-dimensional structure. In this study, we investigated whether this strictly conserved Trp-19 of β-LG would be indispensable for its structure and function such as maintaining the molecular structure and biological activity of β-LG. Spectroscopic and enzymatic oxidation experiments on retinol bound to W19Y, in which Tyr was substituted for Trp-19, showed that Trp-19 was not critical for this binding, but was important for stably maintaining the environment surrounding retinol and the bound retinol. An analysis, using four anti-β-LG monoclonal antibodies as probes, revealed a structural change in region 20-29, but not in the reverse region of Trp-19. A guanidine hydrochloride-induced unfolding study showed that the conformational stability of W19Y was greatly reduced by 6.9 kcal/mol compared to that of wild-type β-LG. These facts indicated that Trp-19 is one of the important residues in correctly maintaining the local structure of β-LG and stably retaining its overall structure, thereby conserving the bound retinol molecule.

DOI： 10.1016/0167-4838(94)90051-5

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

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

We investigated whether any local moieties within a protein molecule could completely refold from the denatured state to regain the native conformation. Bovine β-lactoglobulin (β-LG) was denatured in the presence of guanidine hydrochloride (GdnHCl) as the denaturant. Renaturation of the denatured β- LG was attempted by dialyzing to remove GdnHCl. The renatured molecules regained the same retinol binding activity as that of native β-LG, and physicochemical studies also indicated that refolding of the denatured β-LG had been almost completely successful. Local structural differences between the native and renatured β-LG molecules were evaluated by using our panel of four anti-β-LG monoclonal antibodies (anti-β-LG mAbs). The structures of the epitope regions in native β-LG recognized by two of these mAbs were the same as those in renatured β-LG. However, it is notable that the binding properties of the other two mAbs to native β-LG indicated a wide structural difference in the epitope regions between the native and renatured β-LG. These regions unable to completely refold were the same as those that unfolded preferentially to the α-helix region, shown in the previous report (Kaminogawa, S., Shimizu, M., Ametani, A., Hattori, M., Ando, O., Hachimura, S., Nakamura, Y., Totsuka, M., and Yamauchi, K. (1989) Biochim. Biophys. Acta 998, 50-56). Complete refolding was never attained by several renaturation conditions such as quicker or slower removal of the denaturant, nor by additional oxidation treatment after reducing the disulfide bonds. These results suggest that some specific moiety(ies) in a protein molecule cannot return to the native conformation from a denatured state, even if the other moieties refold completely, and that such a conformational difference between renatured and native forms has no affect on the biological function of ligand binding.

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

A bovine β-Mactoglobulin (β-LG) was expressed in Saccharomyces cerevisiae carrying bovine pre-β-LG cDNA and secreted into its growth medium. The expression plasmid was constructed by inserting the whole coding region of the cDNA encoding pre-β-LG between the promoter and terminator of the yeast glyceraldehyde 3-phosphate dehydrogenase gene of pYG100, a yeast expression vector. In the supernatant of the yeast growth medium, β-LG with a native conformation was detected by sandwich ELISA, and its amount was estimated to be 1.1 mg/1. A Western-immunoblotting analysis revealed that β-LG secrected in the growth medium had the same mobility as that of authentic bovine β-LG. The N-terminal sequence was also identical with that of authentic mature bovine β-LG.

DOI： 10.1271/bbb1961.54.3111

Language：Japanese   Book type：Scholarly book

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Venue：Web開催   Country：Japan  

Event date： 2020.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：Web開催   Country：Japan  

Event date： 2020.11

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：Web開催   Country：Japan  

Event date： 2020.11

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：Web開催   Country：Japan  

Event date： 2020.11

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：Web開催   Country：Japan  

Event date： 2020.11

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：Web開催   Country：Japan  

Event date： 2020.9

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：Web開催   Country：Japan  

Event date： 2020.3

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：九州大学   Country：Japan  

Event date： 2020.3

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：九州大学   Country：Japan  

Event date： 2020.3

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：九州大学   Country：Japan  

Event date： 2020.1

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2020.1

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：産総研つくば中央共用講堂   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京農業大学   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：マリンメッセ（福岡）   Country：Japan  

Event date： 2019.11 - 2019.12

Language：English   Presentation type：Oral presentation (general)  

Venue：Tunisia   Country：Japan  

Event date： 2019.11 - 2019.12

Language：English   Presentation type：Oral presentation (general)  

Venue：Tunisia   Country：Japan  

Event date： 2019.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：タワーホール船堀（東京）   Country：Japan  

Event date： 2019.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：タワーホール船堀（東京）   Country：Japan  

Event date： 2019.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：琉球大学（沖縄）   Country：Japan  

Event date： 2019.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：琉球大学（沖縄）   Country：Japan  

Event date： 2019.7

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：テックデザイン（東京）   Country：Japan  

Event date： 2019.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学農学部3号館141会議室   Country：Japan  

Event date： 2019.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2019.4

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：ＮＣＢリサーチ＆コンサルティング　セミナー室（福岡）   Country：Japan  

Event date： 2019.3

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2019.3

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2019.3

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2019.1

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：シーエムシー出版セミナールーム（東京）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ（神奈川）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：東京ビッグサイト（東京）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：星陵会館（東京）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：星陵会館（東京）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：筑波国際会議場（つくば市）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：筑波国際会議場（つくば市）   Country：Japan  

Event date： 2018.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：筑波国際会議場（つくば市）   Country：Japan  

Event date： 2018.8

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東北大学（仙台）   Country：Japan  

Event date： 2018.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：大阪国際会議場   Country：Japan  

最近、テロメラーゼ触媒サブユニット（TERT）の上皮細胞における発現誘導が育毛を促進することが報告された。これまでの研究で、ヒト表皮ケラチノサイト由来HaCaT細胞において、hTERTプロモーターを活性化する食品成分の同定に成功している。そこで本研究では、これらhTERTプロモーター増強食品の、内在性hTERT増強効果及びHaCaT細胞活性化効果を検証し、その分子基盤を明らかにするとともに、その育毛効果と表現型をマウスを用いたin vivo試験により検証した。

Event date： 2018.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：大阪国際会議場（大阪）   Country：Japan  

Event date： 2018.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：岡山県立大学   Country：Japan  

Event date： 2018.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：岡山県立大学（岡山）   Country：Japan  

Event date： 2018.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：岡山県立大学   Country：Japan  

Event date： 2018.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：名城大学   Country：Japan  

Event date： 2018.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：名城大学   Country：Japan  

Event date： 2018.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：名城大学   Country：Japan  

Event date： 2018.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：名城大学   Country：Japan  

【背景・目的】
近年、育毛を促進する因子としてテロメラーゼ逆転写酵素（TERT）が注目されている。本研究では、ヒト表皮ケラチノサイト由来HaCaT細胞を用いて、ヒトTERT遺伝子を活性化する食品成分を網羅的に探索するためのスクリーニング系を新たに構築し、有効な食品成分の探索・同定を行い、その機能性を評価することを目的とした。
【材料及び方法】
TERT遺伝子プロモーターが活性化されると蛍光タンパク質EGFPが蛍光を発するベクターを作製し、ヒト表皮角化細胞であるHaCaT細胞に導入し、安定株を樹立した。当該細胞株を用いてスクリーニングを行った。各種食品成分を添加し培養した後、イメージングサイトメーターを用いてEGFPの蛍光強度の変化を定量し、各種食品成分のヒトTERT遺伝子プロモーター活性化能を評価した。選定したヒトTERT遺伝子プロモーター増強食品成分について、HaCaT細胞の内在性ヒトTERT遺伝子発現に対する効果及び、HaCaT細胞の増殖に与える影響を評価した。更に、毛髪の合成や成長促進に関わる遺伝子や、毛髪の退行を誘導する遺伝子の発現に対する効果の検証及び、毛周期・毛髪の成長を促進するWntシグナル経路の因子であり、ヒトTERTプロモーターの制御にも関わることが知られているβカテニンの活性に対する効果の検証を行った。
【結果・考察】
イメージングサイトメーターによる解析、リアルタイムPCRによる内在性hTERT発現検証及び、細胞増殖に対する効果から、3種類のポリフェノールサンプルが育毛効果を有する食品成分として有望であると考えられた。また、これらのポリフェノールは、毛髪合成や成長促進に関わる成長因子の発現制御あるいは毛周期や毛髪の成長を促進するWnt経路の活性化を通じて、ヒトTERTの活性化、延いては育毛効果を発揮しているものと考えられた。（737/850）

Event date： 2017.12

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：神戸   Country：Japan  

Event date： 2017.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：神戸   Country：Japan  

Event date： 2017.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：神戸   Country：Japan  

Event date： 2017.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：神戸   Country：Japan  

Event date： 2017.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：神戸   Country：Japan  

Event date： 2017.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：東京大学   Country：Japan  

【背景・目的】
　近年、脳と腸が密接に関わり影響を及ぼし合う「脳腸相関」という概念が注目されている。当研究室ではこれまでの研究で、食肉成分であるカルノシンが脳機能改善効果を示し、脳腸相関活性化能を有することを明らかにしている。そこで我々は、カルノシンによる脳腸相関活性化の分子基盤として腸管細胞由来エクソソームを想定し、その寄与を明らかにすることを目的として研究を行った。
【材料・方法】
　腸管モデル細胞として、ヒト結腸癌由来細胞株であるCaco-2細胞を、神経細胞モデルとしてヒト神経芽細胞腫SH-SY5Y細胞を用いた。カルノシン処理をしたCaco-2細胞の培養上清から、ExoquickあるいはMagCaptureを用いてエクソソームを単離した。エクソソーム内のmiRNAの発現変動及びエクソソームを添加したSH-SY5Y細胞におけるmRNAの発現変動は、マイクロアレイを用いた網羅的解析により評価した。神経突起長は、Neuro-Chrom Pan Neuronal Markerにより染色後、IN Cell Analyzer 1000により定量化し、評価した。
【結果・考察】
カルノシン処理をしたCaco-2細胞由来のエクソソームにおいて発現変動を示すmiRNAのターゲット遺伝子と当該エクソソームを添加したSH-SY5Y細胞において発現変動を示すmRNAを統合解析し、両者で共通に変動するターゲット遺伝子の探索を行った。さらにIngenuity Pathway Analysisにより、神経関連遺伝子を中心としたネットワークの解析を行い、カルノシンの機能性を規定するmiRNA-ターゲット遺伝子を同定した。発現解析の結果から、上記miRNAがカルノシン処理したCaco-2細胞から分泌されるエクソソームに含有され、神経細胞に運ばれるものと予想された。さらに、mimic及びinhibitorを用いた解析から、今回同定したmiRNAとそれによるターゲット遺伝子の発現制御を通じて、神経細胞の神経突起伸長が誘導されることが明らかとなり、カルノシンによるエクソソームを介した神経細胞活性化の分子基盤を明らかにすることができた。

Event date： 2017.11

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：東京大学   Country：Japan  

Event date： 2017.11

Language：English   Presentation type：Oral presentation (general)  

Venue：Kyushu Univ.   Country：Japan  

Event date： 2017.10

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：長崎県立大学シーボルト校   Country：Japan  

Event date： 2017.10

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：長崎県立大学シーボルト校   Country：Japan  

Event date： 2017.10

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：長崎県立大学シーボルト校   Country：Japan  

Event date： 2017.9

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：大阪府立大学   Country：Japan  

Event date： 2017.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京国際フォーラム   Country：Japan  

【背景・目的】
　近年、脳と腸が密接に関わり影響を及ぼし合う「脳腸相関」という概念が注目されている。当研究室ではこれまでの研究で、食肉成分であるカルノシンが認知機能改善効果を示し、脳腸相関活性化能を有することを明らかにしている。そこで我々は、カルノシンによる脳腸相関活性化の分子基盤として腸管細胞由来エクソソームを想定し、その寄与を明らかにすることを目的として研究を行った。
【材料・方法】
　腸管モデル細胞として、ヒト結腸癌由来細胞株であるCaco-2細胞を用いた。カルノシン処理をしたCaco-2細胞の培養上清から、ExoquickあるいはMagCaptureを用いてエクソソームを単離し、エクソソーム内のmiRNAの発現変動をマイクロアレイを用いた網羅的解析により行った。Caco-2培養上清及びエクソソームの機能性は、ヒト神経芽細胞腫SH-SY5Y細胞を用いて検証した。
【結果・考察】
カルノシン処理をしたCaco-2細胞由来のエクソソームmiRNAの発現変動を解析した結果、カルノシン処理により発現変動を示す10種類のmiRNAを同定した。これらmiRNAのターゲット遺伝子をTargetScanによりについて予測したところ、神経管の形成・発達、神経伝達物質の分泌・輸送、ニューロンの発達・分化、軸索の誘導といった、神経細胞の分化・成熟に関わる遺伝子が多く含まれていることが明らかとなった。現在、カルノシン処理した腸管由来エクソソームのSH-SY5Y細胞への取り込み、及びSH-SY5Y細胞における神経突起伸長効果・分化誘導効果について，検証を行っている。

Event date： 2017.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京国際フォーラム   Country：Japan  

Event date： 2017.5

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：沖縄コンベンションセンター   Country：Japan  

Event date： 2017.5

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：沖縄コンベンションセンター   Country：Japan  

Event date： 2017.5

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2017.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：武庫川女子大学   Country：Japan  

Event date： 2017.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：武庫川女子大学   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都女子大学   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都女子大学   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都女子大学   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都女子大学   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都女子大学   Country：Japan  

Event date： 2017.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：京都女子大学   Country：Japan  

Event date： 2016.11 - 2016.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2016.11 - 2016.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2016.11 - 2016.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2016.11 - 2016.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2016.11 - 2016.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2016.11

Language：English   Presentation type：Oral presentation (general)  

Venue：Kobe   Country：Japan  

Event date： 2016.11

Language：English   Presentation type：Oral presentation (general)  

Venue：Kobe   Country：Japan  

Event date： 2016.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学   Country：Japan  

Event date： 2016.11

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学   Country：Japan  

Event date： 2016.7

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州国際会議場   Country：Japan  

Event date： 2016.7

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州国際会議場   Country：Japan  

Event date： 2016.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2016.6

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：横浜パシフィコ   Country：Japan  

Event date： 2015.3

Presentation type：Oral presentation (general)  

Venue：岡山   Country：Japan  

Event date： 2015.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：岡山   Country：Japan  

Event date： 2015.3

Presentation type：Oral presentation (general)  

Venue：岡山   Country：Japan  

Event date： 2015.3

Presentation type：Oral presentation (general)  

Venue：岡山   Country：Japan  

Event date： 2015.3 - 2015.2

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：大分県   Country：Japan  

Event date： 2013.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：仙台   Country：Japan  

Event date： 2013.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：仙台   Country：Japan  

Event date： 2013.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：仙台   Country：Japan  

Event date： 2013.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：仙台   Country：Japan  

Event date： 2013.3

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：仙台   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.12

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：福岡   Country：Japan  

Event date： 2012.11

Language：English   Presentation type：Oral presentation (general)  

Venue：名古屋   Country：Japan  

Event date： 2012.11

Language：English   Presentation type：Oral presentation (general)  

Venue：名古屋   Country：Japan  

Event date： 2012.10

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.10

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.10

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.10

Language：English   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.10

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.10

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.10

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2012.6

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2012.6

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2012.6

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2012.6

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2012.6

Language：Japanese   Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2012.5

Presentation type：Oral presentation (general)  

Venue：仙台   Country：Japan  

Event date： 2012.3

Presentation type：Oral presentation (general)  

Venue：京都   Country：Japan  

Event date： 2012.3

Presentation type：Oral presentation (general)  

Venue：京都   Country：Japan  

Event date： 2012.3

Presentation type：Oral presentation (general)  

Venue：京都   Country：Japan  

Event date： 2012.3

Presentation type：Oral presentation (general)  

Venue：京都   Country：Japan  

Event date： 2012.3

Presentation type：Oral presentation (general)  

Venue：京都   Country：Japan  

Event date： 2011.12

Presentation type：Symposium, workshop panel (public)  

Venue：横浜   Country：Japan  

Event date： 2011.12

Presentation type：Oral presentation (general)  

Venue：横浜   Country：Japan  

Event date： 2011.12

Presentation type：Oral presentation (general)  

Venue：横浜   Country：Japan  

Event date： 2011.12

Presentation type：Oral presentation (general)  

Venue：横浜   Country：Japan  

Event date： 2011.11

Presentation type：Symposium, workshop panel (public)  

Venue：九州大学   Country：Japan  

Event date： 2011.11

Presentation type：Symposium, workshop panel (public)  

Venue：名古屋   Country：Japan  

Event date： 2011.10

Presentation type：Oral presentation (general)  

Venue：東京   Country：Japan  

Event date： 2011.10

Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2011.7

Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2011.7

Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2011.7

Presentation type：Symposium, workshop panel (public)  

Venue：東京   Country：Japan  

Event date： 2011.7

Presentation type：Oral presentation (general)  

Venue：北九州   Country：Japan  

Event date： 2011.7

Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2011.7

Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan  

Event date： 2011.7

Presentation type：Symposium, workshop panel (public)  

Venue：北九州   Country：Japan