Updated on 2021/12/29

写真a

 
OHARA Naoya
 
Organization
Medicine, Dentistry and Pharmaceutical Sciences Professor
Position
Professor
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Degree

  • 博士(歯学) ( 長崎大学 )

Research Interests

  • pathogenic bacteria

  • 病原細菌

Research Areas

  • Life Science / Oral biological science

  • Life Science / Bacteriology

Research History

  • 岡山大学学術研究院医歯薬学域口腔微生物学分野 教授

    2021.4

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  • 岡山大学大学院医歯薬学総合研究科口腔微生物学分野 教授

    2009.7 - 2021.3

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  • 長崎大学大学院医歯薬学総合研究科口腔微生物学分野 助教授

    2003.4 - 2007.1

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Papers

  • Porphyromonas gingivalis gingipains induce cyclooxygenase-2 expression and prostaglandin E2 production via ERK1/2-activated AP-1 (c-Jun/c-Fos) and IKK/NF-κB p65 cascades Reviewed

    Masaaki Nakayama, Mariko Naito, Kazuhiro Omori, Shintaro Ono, Koji Nakayama, Naoya Ohara

    Journal of Immunology   in press   2022

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  • GRIM-19 is a target of mycobacterial Zn2+ metalloprotease 1 and indispensable for NLRP3 inflammasome activation Reviewed

    Tomomi Kurane, Tetsuro Matsunaga, Tomoaki Ida, Kazuko Sawada, Akira Nishimura, Masayuki Fukui, Masayuki Umemura, Masaaki Nakayama, Naoya Ohara, Sohkichi Matsumoto, Takaaki Akaike, Goro Matsuzak, Giichi Takaesu

    FASEB Journal   in press   2021.12

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  • Attempt of thyX gene silencing and construction of a thyX deleted clone in a Mycobacterium bovis BCG. Reviewed International journal

    Yuki Arimura, Yusuke Minato, Takayuki Wada, Masaaki Nakayama, Ayako Ryumon, Nao Hirata, Chie Nakajima, Yasuhiko Suzuki, Manabu Ato, Kazuo Kobayashi, Naoko Ohara, Seiji Iida, Naoya Ohara

    Microbiology and immunology   2021.9

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    Mycobacterium tuberculosis, the causative agent of tuberculosis, possess flavin-dependent thymidylate synthase, ThyX. Since thyX is absent in humans and was shown to be essential for M. tuberculosis normal growth, ThyX is thought to be an attractive novel TB drug target. In this study, we assessed thyX essentiality in Mycobacterium bovis BCG strains using CRISPR interference based gene silencing and found that thyX is not essential in an M. bovis BCG Tokyo derivative strain. We successfully constructed a thyX deletion mutant strain from that strain, which reinforces the non-essentiality of thyX under a certain genetic background. This article is protected by copyright. All rights reserved.

    DOI: 10.1111/1348-0421.12944

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  • Comparative Study of the Susceptibility to Oxidative Stress between Two Types of Mycobacterium bovis BCG Tokyo 172. Reviewed International journal

    Keiichi Taniguchi, Daisuke Hayashi, Naomi Yasuda, Mao Nakayama, Kaori Yazawa, Shouta Ogawa, Yuji Miyatake, Saki Suda, Haruka Tomita, Miki Tokuda, Saotomo Itoh, Jun-Ichi Maeyama, Naoya Ohara, Saburo Yamamoto, Shigeaki Hida, Kikuo Onozaki, Takemasa Takii

    mSphere   6 ( 2 )   2021.3

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    Genomic analysis revealed that the vaccine seed lot of Mycobacterium bovis bacillus Calmette-Guérin (BCG) Tokyo 172 contains two subclones (types I and II), but their phenotypic differences have not been elucidated. In this study, we compared the susceptibility of bacilli types I and II to oxidative stress in vitro and within host cells. Notably, the subclones displayed similar superoxide dismutase activity; however, foam height in the catalase test and lysate catalase/peroxidase activity were higher for type I bacilli than for type II bacilli. Additionally, type I bacilli were less susceptible to hydrogen peroxide (H2O2) than type II bacilli. After exposure to H2O2, antioxidative stress response genes katG, ahpC, sodA, and trxA were more strongly induced in type I bacilli than in type II bacilli. Further, we investigated cell survival in macrophages. Fewer type II bacilli were recovered than type I bacilli. However, in the presence of apocynin, a specific inhibitor of NADPH oxidase, type II recovery was greater than that of type I. The production of interleukin 1β (IL-1β), IL-12 p40, and tumor necrosis factor alpha (TNF-α) was higher in type I bacillus-infected macrophages than in type II bacillus-infected macrophages. The proportions of type I and type II bacilli in vaccine lots over 3 years (100 lots) were 97.6% ± 1.5% and 2.4% ± 1.5%, respectively. The study results illustrated that type I bacilli are more resistant to oxidative stress than type II bacilli. Overall, these findings provide important information in terms of the quality control and safety of BCG Tokyo 172 vaccine.IMPORTANCE This study revealed the difference of in vivo and in vitro antioxidative stress properties of BCG Tokyo 172 types I and II as one of the bacteriological characteristics. In particular, the bacilli exhibited differences in catalase/peroxidase activity, which could explain their different protective effects against infection. The differences correlated with survival in the host cell and the production of proinflammatory cytokines to protect against infection by Mycobacterium tuberculosis The proportion of bacilli types I and II in all commercial lots of BCG Tokyo 172 over 3 years (100 lots) was constant. The findings also highlighted the importance of analyzing their content for quality control during vaccine production.

    DOI: 10.1128/mSphere.00111-21

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  • Point mutation in the stop codon of MAV_RS14660 increases the growth rate of Mycobacterium avium subspecies hominissuis Reviewed

    Tomomi Kawakita, Tetsu Mukai, Mitsunori Yoshida, Hiroyuki Yamada, Masaaki Nakayama, Yuji Miyamoto, Masato Suzuki, Noboru Nakata, Takemasa Takii, Akihide Ryo, Naoya Ohara, Manabu Ato

    Microbiology   in press ( 2 )   2020.12

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

    <italic>
    <named-content content-type="subspecies">
    <ext-link xmlns:xlink="http://www.w3.org/1999/xlink" ext-link-type="uri" xlink:href="http://doi.org/10.1601/nm.14878" xlink:type="simple">Mycobacterium avium</ext-link>
    </named-content>
    </italic> subspecies <italic>
    <named-content content-type="subspecies">
    <ext-link xmlns:xlink="http://www.w3.org/1999/xlink" ext-link-type="uri" xlink:href="http://doi.org/10.1601/nm.14878" xlink:type="simple">hominissuis</ext-link>
    </named-content>
    </italic> (MAH) is a pathogen that causes various non-tuberculous mycobacterial diseases in humans and animals worldwide. Among the genus, MAH is characterized by relatively slow growth. Here, we isolated a rapidly growing variant of the MAH 104 strain. The variant strain (named N104) exhibited an enhanced growth rate and higher motility compared to the parent MAH 104 strain (P104). Whole-genome sequencing analysis of N104 revealed the loss of the stop codon of <italic>MAV_RS14660</italic> due to a single nucleotide replacement, resulting in the substitution of the codon for tryptophan. Notably, exclusion of the stop codon ligated the open reading frames and caused the fusion of two adjacent proteins. A revertant parent strain, in which a mutation was introduced to restore the stop codon, revealed that elimination of the stop codon in <italic>MAV_RS14660</italic> was responsible for the N104 phenotype. Furthermore, we analysed the phenotypes of the parent and mutated strains by determining the functions of the <italic>MAV_RS14660</italic> and <italic>MAV_RS14655</italic> coding regions flanking the stop codon. The mutant strains, expected to express a fusion protein, exhibited increased resistance to antimicrobial drugs and exogenous copper toxicity compared to that of the parent strains. These findings suggest that the fusion of the <italic>MAV_RS14660</italic>- and <italic>MAV_RS14655</italic>-encoding regions in the mutant N104 strain could be related to the modified functions of these intrinsic proteins.

    DOI: 10.1099/mic.0.001007

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  • Construction and characterization of the PGN_0296 mutant of Porphyromonas gingivalis Reviewed

    Abu Saleh Muhammad Shahriar, Shintaro Ono, Masaaki Nakayama, Naoko Ohara, Naoya Ohara

    Journal of Oral Biosciences   62 ( 4 )   322 - 326   2020.12

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    Authorship:Last author, Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

    DOI: 10.1016/j.job.2020.09.007

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  • 抗酸菌のバイオフィルム形成と休眠時遺伝子発現比較

    西内 由紀子, 大田 篤, 矢野 大和, 岩本 朋忠, 阿戸 学, 松本 壮吉, 大原 直也, 丸山 史人

    BACTERIAL ADHERENCE & BIOFILM   33   49 - 52   2020.5

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    Language:Japanese   Publisher:日本バイオフィルム学会  

    環境から感染する肺非結核性抗酸菌症は世界中で著しく増加しているため公衆衛生上の解決すべき課題の一つである。私たちは主な起因菌であるMycobacterium avium subsp.hominissuis(MAH)は低酸素環境にするとバイオフィルム形成が促進されることを見出した。一方で類縁菌の結核菌は酸素欠乏で休眠する。結核菌は二成分制御系のDosS、DosTが酸素濃度の低下を感知し、DosRが休眠に必要なタンパク質の発現調節を行なっている。MAHも結核菌と同様に二成分制御系が働いているとの仮説を立て、トランスクリプトーム解析を行って検証した。MAHを大気条件下または5%酸素条件下で培養後RNAを抽出し、RNAseq解析を行った。その結果、141/109遺伝子の有意な発現増加/低下が認められた。MAH 104は17セットの2成分制御系がゲノム上認められる。このうち結核菌のDosS/DosRオルソログに相当する遺伝子(MAV_RS19700/MAV_RS19705)は変化しなかった。結核菌にはないセンサーヒスチジンキナーゼ遺伝子(MAV_RS11960)の発現が増加した。この遺伝子は低酸素を感知するPASドメイン構造を有し、DosSと相同性の高いヒスチジンキナーゼを有していることから、有力な候補遺伝子であると思われた。次に、低酸素環境に適応する共通の変化があるのではないかと考え、MAH発現増加遺伝子中にDosRレギュロンのオルソログがないか検索したところ、10遺伝子が該当した。これらの遺伝子は、低酸素環境に適応して生存を維持するために必要な遺伝子であると推察された。(著者抄録)

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  • Rescue from Stx2-Producing E. coli-Associated Encephalopathy by Intravenous Injection of Muse Cells in NOD-SCID Mice. Reviewed International journal

    Ryo Ozuru, Shohei Wakao, Takahiro Tsuji, Naoya Ohara, Takashi Matsuba, Muhammad Y Amuran, Junko Isobe, Morio Iino, Naoki Nishida, Sari Matsumoto, Kimiharu Iwadate, Noriko Konishi, Kaori Yasuda, Kosuke Tashiro, Misato Hida, Arisato Yadoiwa, Shinsuke Kato, Eijiro Yamashita, Sohkichi Matsumoto, Yoichi Kurozawa, Mari Dezawa, Jun Fujii

    Molecular therapy : the journal of the American Society of Gene Therapy   28 ( 1 )   100 - 118   2020.1

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    Shiga toxin-producing Escherichia coli (STEC) causes hemorrhagic colitis, hemolytic uremic syndrome, and acute encephalopathies that may lead to sudden death or severe neurologic sequelae. Current treatments, including immunoglobulin G (IgG) immunoadsorption, plasma exchange, steroid pulse therapy, and the monoclonal antibody eculizumab, have limited effects against the severe neurologic sequelae. Multilineage-differentiating stress-enduring (Muse) cells are endogenous reparative non-tumorigenic stem cells that naturally reside in the body and are currently under clinical trials for regenerative medicine. When administered intravenously, Musecells accumulate to the damaged tissue, where they exert anti-inflammatory, anti-apoptotic, anti-fibrotic, and immunomodulatory effects, and replace damaged cells by differentiating into tissue-constituent cells. Here, severely immunocompromised non-obese diabetic/severe combined immunodeficiency (NOD-SCID) mice orally inoculated with 9 × 109 colony-forming units of STEC O111 and treated 48 h later with intravenous injection of 5 × 104 Muse cells exhibited 100% survival and no severe after-effects of infection. Suppression of granulocyte-colony-stimulating factor (G-CSF) by RNAi abolished the beneficial effects of Muse cells, leading to a 40% death and significant body weight loss, suggesting the involvement of G-CSF in the beneficial effects of Muse cells in STEC-infected mice. Thus, intravenous administration of Muse cells could be a candidate therapeutic approach for preventing fatal encephalopathy after STEC infection.

    DOI: 10.1016/j.ymthe.2019.09.023

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  • Construction and characterization of a PGN_0297 mutant of Porphyromonas gingivalis: evidence of the contribution of PGN_0297 to gingipain activity. Reviewed

    Ono S, Nakayama M, Tachibana M, Shahriar ASM, Heling W, Takashiba S, Ohara N

    Acta Medica Okayama   73 ( 4 )   315 - 323   2019.8

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  • Sequential Sensing by TLR2 and Mincle Directs Immature Myeloid Cells to Protect against Invasive Group A Streptococcal Infection in Mice Reviewed

    Takayuki Matsumura, Tadayoshi Ikebe, Koji Arikawa, Masahito Hosokawa, Michio Aiko, Aoi Iguchi, Ikuko Togashi, Sayaka Kai, Sakiko Ohara, Naoya Ohara, Makoto Ohnishi, Haruo Watanabe, Kazuo Kobayashi, Haruko Takeyama, Sho Yamasaki, Yoshimasa Takahashi, Manabu Ato

    Cell Reports   27 ( 2 )   561 - 571.e6   2019.4

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    DOI: 10.1016/j.celrep.2019.03.056

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  • Discrimination of Mycobacterium leprae and Mycobacterium haemophilum in Clinical Isolates and Specimens by Multiplex PCR Assay and Prediction of Drug Susceptibility. Reviewed International journal

    Naoya Kitaoka, Hanako Fukano, Mitsunori Yoshida, Yuji Miyamoto, Shuichi Mori, Norihisa Ishii, Manabu Ato, Naoya Ohara, Yoshihiko Hoshino

    Journal of clinical microbiology   57 ( 2 )   e01760-18   2019.2

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    DOI: 10.1128/JCM.01760-18

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  • Mycobacterial infection induces eosinophilia and production of α-defensin by eosinophils in mice. Reviewed

    Khatun A, Sakurai M, Sakai Y, Tachibana M, Ohara N, Morimoto M

    The Journal of veterinary medical science   81 ( 1 )   138 - 142   2019.1

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    It has been well known in humans that eosinophil infiltration into the site of inflammation and eosinophilia occur in mycobacterial infections. However, the role of eosinophils against the mycobacterium is unclear. We showed in previous study that in situ mouse eosinophils infiltrated into tissues produce α-defensin, an anti-bacterial peptide. We investigated in this study whether eosinophils reacting to mycobacteria produce α-defensin in mice and whether it can be used as a model. We showed that mycobacterial infection induced blood eosinophilia and infiltration of α-defensin producing eosinophils that to surround mycobacteria at the site of infection. These findings were usually seen during human mycobacterial infection. We established a good model to study host defense mechanism against mycobacteria through α-defensin via eosinophils.

    DOI: 10.1292/jvms.18-0619

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  • C-terminal intrinsically disordered region-dependent organization of the mycobacterial genome by a histone-like protein Reviewed

    Anna Savitskaya, Akihito Nishiyama, Takehiro Yamaguchi, Yoshitaka Tateishi, Yuriko Ozeki, Masaaki Nameta, Tomohiro Kon, Shaban A. Kaboso, Naoya Ohara, Olga V. Peryanova, Sohkichi Matsumoto

    Scientific Reports   8 ( 1 )   2018.12

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    The architecture of the genome influences the functions of DNA from bacteria to eukaryotes. Intrinsically disordered regions (IDR) of eukaryotic histones have pivotal roles in various processes of gene expression. IDR is rare in bacteria, but interestingly, mycobacteria produce a unique histone-like protein, MDP1 that contains a long C-terminal IDR. Here we analyzed the role of IDR in MDP1 function. By employing Mycobacterium smegmatis that inducibly expresses MDP1 or its IDR-deficient mutant, we observed that MDP1 induces IDR-dependent DNA compaction. MDP1-IDR is also responsible for the induction of growth arrest and tolerance to isoniazid, a front line tuberculosis drug that kills growing but not growth-retardated mycobacteria. We demonstrated that MDP1-deficiency and conditional knock out of MDP1 cause spreading of the M. smegmatis genome in the stationary phase. This study thus demonstrates for the first time a C-terminal region-dependent organization of the genome architecture by MDP1, implying the significance of IDR in the function of bacterial histone-like protein.

    DOI: 10.1038/s41598-018-26463-9

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  • Interleukin-21 Induces Short-Lived Effector CD8(+) T Cells but Does Not Inhibit Their Exhaustion after Mycobacterium bovis BCG Infection in Mice Reviewed

    Noguchi Naoto, Nakamura Risa, Hatano Shinya, Yamada Hisakata, Sun Xun, Ohara Naoya, Yoshikai Yasunobu

    INFECTION AND IMMUNITY   86 ( 8 )   2018.8

  • Recovery of mycobacteriophages from archival stocks stored for approximately 50 years in Japan Reviewed

    Takako Ujihara, Jumpei Uchiyama, Tadahiro Nasukawa, Hiroki Ando, Hironobu Murakami, Naoya Ohara, Midori Ogawa, Toshio Yamazaki, Masanori Daibata, Masahiro Sakaguchi, Shigenobu Matsuzaki

    Archives of Virology   163 ( 7 )   1915 - 1919   2018.7

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    © 2018, Springer-Verlag GmbH Austria, part of Springer Nature. Mycobacteriophage archival stocks have been kept for ca. 20–50 years in Japan. In this study, we attempted to recover mycobacteriophages from 50 archival stocks and briefly analyzed the recovered phages. The phages were recovered from 72.2% (13/18) of the lyophilized stocks that had been stored for 47-56 years. Moreover, the analysis of 12 representative recovered phages led to their classification as belonging to the family Siphoviridae, and seven of them were typed by polymerase chain reaction (PCR) targeting the gene that encodes the tape measure protein. Considering these results, lyophilization seems to be suitable for phage archival storage.

    DOI: 10.1007/s00705-018-3788-8

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  • Genome sequences of 12 mycobacteriophages recovered from archival stocks in Japan Reviewed

    Jumpei Uchiyama, Keijiro Mizukami, Koji Yahara, Shin Ichiro Kato, Hironobu Murakami, Tadahiro Nasukawa, Naoya Ohara, Midori Ogawa, Toshio Yamazaki, Shigenobu Matsuzaki, Masahiro Sakaguchi

    Genome Announcements   6 ( 25 )   2018.6

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    © 2018 Uchiyama et al. Using Mycobacterium smegmatis mc2155, 12 siphoviruses were recovered from long-term archival stocks stored in Japan. Their genome sequences were 46.0 to 61.3 kbp with 63 to 68% G+C contents, which allowed them to be categorized within cluster W and subclusters A1, A2, B3, A7, I1, and K4.

    DOI: 10.1128/genomeA.00472-18

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  • The influence of zoledronate and teriparatide on gamma delta T cells in mice Reviewed

    Eiki Yamachika, Yuichi Matsui, Masakazu Matsubara, Tatsushi Matsumura, Naoki Nakata, Norifumi Moritani, Atsushi Ikeda, Hidetsugu Tsujigiwa, Naoya Ohara, Seiji Iida

    Journal of Dental Sciences   12 ( 4 )   333 - 339   2017.12

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:Association for Dental Sciences of the Republic of China  

    Background/purpose Few studies have investigated the possibility that bisphosphonate-related osteonecrosis of the jaw (BRONJ) might reflect an immune response
    however, gamma delta T cells have been shown to significantly decline in the blood of BRONJ patients. Additionally, there have been some reports of teriparatide usage for the treatment of BRONJ. In this study, we compared the effects of zoledronate and teriparatide on lymphocyte populations and inflammatory cytokine production in mice. Materials and methods Thirty female ICR mice were divided into three groups (n = 10 each): a vehicle, a zoledronate, and a teriparatide group. Drugs were administered for 8 weeks in each group. Lymphocytes in the blood and thymus were analyzed and femurs were used for histological observation and lymphocytes analysis of bone marrow. Cytokines were measured in separated serum using Milliplex® multiplex immunoassay analysis. Results Zoledronate decreased the T cell number in the bone marrow. Additionally, serum levels of interleukin (IL)-2, IL-7, IL-12, IL-15 and RANTES, which are cytokines that affect T cell activation, differentiation and/or proliferation, were significantly lower in zoledronate treated mice. Conversely, teriparatide treatment induced an increase in gamma delta T cells in peripheral blood. Conclusion Gamma delta T cells in the bone marrow are expected to decrease with zoledronate treatment and increase with teriparatide treatment. If BRONJ involves a loss of gamma delta T cells in the circulation or bone marrow, then the increase in gamma delta T cells that is induced by teriparatide may account for its ability to resolve BRONJ.

    DOI: 10.1016/j.jds.2017.03.007

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  • Molecular mechanisms of Porphyromonas gingivalis-host cell interaction on periodontal diseases Invited Reviewed

    Masaaki Nakayama, Naoya Ohara

    Japanese Dental Science Review   53 ( 4 )   134 - 140   2017.11

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    Porphyromonas gingivalis (P. gingivalis) is a major oral pathogen and associated with periodontal diseases including periodontitis and alveolar bone loss. In this review, we indicate that two virulence factors, which are hemoglobin receptor protein (HbR) and cysteine proteases “gingipains”, expressed by P. gingivalis have novel functions on the pathogenicity of P. gingivalis. P. gingivalis produces three types of gingipains and concomitantly several adhesin domains. Among the adhesin domains, hemoglobin receptor protein (HbR), also called HGP15, has the function of induction of interleukin-8 (IL-8) expression in human gingival epithelial cells, indicating the possibility that HbR is associated with P. gingivalis-induced periodontal inflammation. On bacteria-host cells contact, P. gingivalis induces cellular signaling alteration in host cells. Phosphatidylinositol 3-kinase (PI3K) and Akt are well known to play a pivotal role in various cellular physiological functions including cell survival and glucose metabolism in mammalian cells. Recently, we demonstrated that gingipains attenuate the activity of PI3K and Akt, which might have a causal influence on periodontal diseases by chronic infection to the host cells from the speculation of molecular analysis. In this review, we discuss new molecular and biological characterization of the virulence factors from P. gingivalis.

    DOI: 10.1016/j.jdsr.2017.06.001

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  • Novel function of Porphyromonas gingivalis gingipains in the PI3K/Akt signaling pathway Invited Reviewed

    Nakayama Masaaki, Ohara Naoya

    JOURNAL OF ORAL BIOSCIENCES   59 ( 3 )   131 - 134   2017.8

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    Language:English   Publisher:Japanese Association for Oral Biology  

    Background Porphyromonas gingivalis is s major oral bacterium closely associated with periodontal diseases including periodontitis and directly affects host cellular signaling. The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway plays multiple roles in various cell functions including cell survival and glucose metabolism. In this review, we describe the effect of gingipains on the PI3K/Akt signaling pathway in P. gingivalis infection. Highlight Gingipains inactivate PI3K and Akt in gingival epithelial cells infected with P. gingivalis. These events occur independently of invasion of this organism into the cells and are required for the enzymatic activity of gingipains. Furthermore, 3-Phosphoinositide-dependent protein kinase-1 (PDK1) failed to translocate to the plasma membrane from the cytosol following PI3K inactivation. Additionally, dephosphorylation of Akt downstream proteins, including glycogen synthase kinase 3 (GSK3), mammalian target of rapamycin (mTOR), and Bad, occurs in parallel with the dysregulation of PI3K/PDK1/Akt cascades. Conclusion This review describes the biological characterization of gingipains, which inactivate PI3K and Akt, and disorder the PI3K/Akt signaling pathway. Hence, gingipains may decrease cellular physiological functions, eventually disrupting the gingival epithelium and causing development of periodontal diseases.

    DOI: 10.1016/j.job.2017.05.003

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  • IL-21 inhibits IL-17A-producing T-cell response after infection with Bacillus Calmette-Guerin via induction of apoptosis Reviewed

    Yinxia Huang, Yumiko Matsumura, Shinya Hatano, Naoto Noguchi, Tesshin Murakami, Yoichiro Iwakura, Xun Sun, Naoya Ohara, Yasunobu Yoshikai

    INNATE IMMUNITY   22 ( 8 )   588 - 597   2016.11

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:SAGE PUBLICATIONS LTD  

    Innate T cells expressing V6 produce IL-17A at an early stage following infection with Mycobacterium bovis Bacillus Calmette-Guerin (BCG). In this study, we used IL-21 receptor knockout (IL-21R KO) mice and IL-21-producing recombinant BCG mice (rBCG-Ag85B-IL-21) to examine the role of IL-21 in the regulation of IL-17A-producing innate T-cell response following BCG infection. IL-17A-producing V6(+) T cells increased in the peritoneal cavity of IL-21R KO mice more than in wild type mice after BCG infection. In contrast, the number of IL-17A-producing V6(+) T cells was significantly lower after inoculation with rBCG-Ag85B-IL-21 compared with control rBCG-Ag85B. Notably, exogenous IL-21 selectively induced apoptosis of IL-17A-producing V6(+) T cells via Bim. Thus, these results suggest that IL-21 acts as a potent inhibitor of a IL-17A-producing T-cell subset during BCG infection.

    DOI: 10.1177/1753425916664125

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  • Antitumor activity of recombinant Bacille Calmette-Guerin secreting interleukin-15-Ag85B fusion protein against bladder cancer Reviewed

    Ario Takeuchi, Masatoshi Eto, Katsunori Tatsugami, Masaki Shiota, Hisakata Yamada, Yoriyuki Kamiryo, Takashi Dejima, Eiji Kashiwagi, Keijiro Kiyoshima, Junichi Inokuchi, Ryosuke Takahashi, Akira Yokomizo, Naoya Ohara, Yasunobu Yoshikai

    INTERNATIONAL IMMUNOPHARMACOLOGY   35   327 - 331   2016.6

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

    Mycobacterium bovis Bacillus Calmette-Guerin (BCG) is used for the treatment of bladder cancer. The recruitment of neutrophlis to the bladder after BCG instillation exerts anti-tumor activity against bladder tumor. We have recently demonstrated that interleukin (IL)-17A produced by gamma delta T cells played a role in the recruitment of neutrophlis to the bladder after BCG instillation. IL-15 is known to play an important role in neutrophil migration during inflammation. We previously constructed a recombinant BCG strain expressing the fusion protein of IL-15 and Ag85B (BCG-IL-15) for prevention of Mycobacterium tuberculosis infection. Here we compared the efficacy of the BCG-IL-15 in protection against bladder cancer with that of rBCG-Ag85B (BCG).
    Six-week-old female C57BL/6 mice were inoculated with MB49 bladder tumor cells in the bladder and subsequently intravesically inoculated with BCG or BCG-IL-15.
    BCG-IL-15 treatment significantly prolonged survival of mice inoculated with bladder cancer cells compared with BCG treatment. Infiltration of neutrophils was significantly elevated in BCGB-IL-15 treated mice accompanied by increased chemokines (MIP-2 and MIP-1 alpha) in the bladder. Thus, BCG-IL-15 exerted additive effect on Infiltration of neutrophils in the bladder. BCG-IL-15 may be a promising drug for non-muscle invasive bladder cancer. (C) 2016 Elsevier B.V. All rights reserved.

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  • Recombinant Mycobacterium bovis bacillus Calmette-Guerin expressing Ag85B-IL-7 fusion protein enhances IL-17A-producing innate gamma delta T cells Reviewed

    Shinya Hatano, Toshiki Tamura, Masayuki Umemura, Goro Matsuzaki, Naoya Ohara, Yasunobu Yoshikai

    VACCINE   34 ( 22 )   2490 - 2495   2016.5

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    Interleukin 7 (IL-7) has an important function in the development and maintenance of IL-17A+ gamma delta T cells. We here constructed a recombinant Mycobacterium bovis bacillus Calmette-Guerin expressing antigen 85B (Ag85B)-IL-7 fusion protein (rBCG-Ag85B-IL-7). The Ag85B-IL-7 fusion protein and IL-7 were detected in the bacterial lysate of rBCG-Ag85B-IL-7. rBCG-Ag85B-IL-7 was the same in number as control rBCG expressing Ag85B (rBCG-Ag85B) in the lung at the early stage after intravenous inoculation, whereas the numbers of IL-17A+ gamma delta T cells and Ag-specific Th1 cells were significantly higher in the lungs of mice inoculated with rBCG-Ag85B-IL-7 than those inoculated with rBCG-Ag85B. The Ag-specific Thl cell response was impaired in mice lacking IL-17A+ gamma delta T cells after inoculation with rBCG-Ag85B-IL-7. Thus, rBCG-Ag85B-IL-7 increases the pool size of IL-17A+ gamma delta T cells, which subsequently augment the Thl response to mycobacterial infection. (C) 2016 Elsevier Ltd. All rights reserved.

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  • Involvement of an Skp-Like Protein, PGN_0300, in the Type IX Secretion System of Porphyromonas gingivalis Reviewed

    Yuko Taguchi, Keiko Sato, Hideharu Yukitake, Tetsuyoshi Inoue, Masaaki Nakayama, Mariko Naito, Yoshio Kondo, Konami Kano, Tomonori Hoshino, Koji Nakayama, Shogo Takashiba, Naoya Ohara

    INFECTION AND IMMUNITY   84 ( 1 )   230 - 240   2016.1

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    The oral Gram-negative anaerobic bacterium Porphyromonas gingivalis is an important pathogen involved in chronic periodontitis. Among its virulence factors, the major extracellular proteinases, Arg-gingipain and Lys-gingipain, are of interest given their abilities to degrade host proteins and process other virulence factors. Gingipains possess C-terminal domains (CTDs) and are translocated to the cell surface or into the extracellular milieu by the type IX secretion system (T9SS). Gingipains contribute to the colonial pigmentation of the bacterium on blood agar. In this study, Omp17, the PGN_0300 gene product, was found in the outer membrane fraction. A mutant lacking Omp17 did not show pigmentation on blood agar and showed reduced proteolytic activity of the gingipains. CTD-containing proteins were released from bacterial cells without cleavage of the CTDs in the omp17 mutant. Although synthesis of the anionic polysaccharide (A-LPS) was not affected in the omp17 mutant, the processing of and A-LPS modification of CTD-containing proteins was defective. PorU, a C-terminal signal peptidase that cleaves the CTDs of other CTD-containing proteins, was not detected in any membrane fraction of the omp17 mutant, suggesting that the defective maturation of CTD-containing proteins by impairment of Omp17 is partly due to loss of function of PorU. In the mouse subcutaneous infection experiment, the omp17 mutant was less virulent than the wild type. These results suggested that Omp17 is involved in P. gingivalis virulence.

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  • Deep sequencing analysis of the heterogeneity of seed and commercial lots of the bacillus Calmette-Guerin (BCG) tuberculosis vaccine substrain Tokyo-172 Reviewed

    Takayuki Wada, Fumito Maruyama, Tomotada Iwamoto, Shinji Maeda, Taro Yamamoto, Ichiro Nakagawa, Saburo Yamamoto, Naoya Ohara

    SCIENTIFIC REPORTS   5   17827   2015.12

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    BCG, only vaccine available to prevent tuberculosis, was established in the early 20th century by prolonged passaging of a virulent clinical strain of Mycobacterium bovis. BCG Tokyo-172, originally distributed within Japan in 1924, is one of the currently used reference substrains for the vaccine. Recently, this substrain was reported to contain two spontaneously arising, heterogeneous subpopulations (Types I and II). The proportions of the subpopulations changed over time in both distributed seed lots and commercial lots. To maintain the homogeneity of live vaccines, such variations and subpopulational mutations in lots should be restrained and monitored. We incorporated deep sequencing techniques to validate such heterogeneity in lots of the BCG Tokyo-172 substrain without cloning. By bioinformatics analysis, we not only detected the two subpopulations but also detected two intrinsic variations within these populations. The intrinsic variants could be isolated from respective lots as colonies cultured on plate media, suggesting analyses incorporating deep sequencing techniques are powerful, valid tools to detect mutations in live bacterial vaccine lots. Our data showed that spontaneous mutations in BCG vaccines could be easily monitored by deep sequencing without direct isolation of variants, revealing the complex heterogeneity of BCG Tokyo-172 and its daughter lots currently in use.

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  • Glycopeptidolipid of Mycobacterium smegmatis J15cs Affects Morphology and Survival in Host Cells Reviewed

    Nagatoshi Fujiwara, Naoya Ohara, Midori Ogawa, Shinji Maeda, Takashi Naka, Hatsumi Taniguchi, Saburo Yamamoto, Minoru Ayata

    PLOS ONE   10 ( 5 )   e0126813   2015.5

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    Mycobacterium smegmatis has been widely used as a mycobacterial infection model. Unlike the M. smegmatis mc(2)155 strain, M. smegmatis J15cs strain has the advantage of surviving for one week in murine macrophages. In our previous report, we clarified that the J15cs strain has deleted apolar glycopeptidolipids (GPLs) in the cell wall, which may affect its morphology and survival in host cells. In this study, the gene causing the GPL deletion in the J15cs strain was identified. The mps1-2 gene (MSMEG_0400-0402) correlated with GPL biosynthesis. The J15cs strain had 18 bps deleted in the mps1 gene compared to that of the mc2155 strain. The mps1-complemented J15cs mutant restored the expression of GPLs. Although the J15cs strain produces a rough and dry colony, the colony morphology of this mps1-complement was smooth like the mc2155 strain. The length in the mps1-complemented J15cs mutant was shortened by the expression of GPLs. In addition, the GPL-restored J15cs mutant did not survive as long as the parent J15cs strain in the murine macrophage cell line J774.1 cells. The results are direct evidence that the deletion of GPLs in the J15cs strain affects bacterial size, morphology, and survival in host cells.

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  • Attenuation of the Phosphatidylinositol 3-Kinase/Akt Signaling Pathway by Porphyromonas gingivalis Gingipains RgpA, RgpB, and Kgp Reviewed

    Masaaki Nakayama, Tetsuyoshi Inoue, Mariko Naito, Koji Nakayama, Naoya Ohara

    JOURNAL OF BIOLOGICAL CHEMISTRY   290 ( 8 )   5190 - 5202   2015.2

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    Porphyromonas gingivalis is a major pathogen of periodontal diseases, including periodontitis. We have investigated the effect of P. gingivalis infection on the PI3K/Akt (protein kinase B) signaling pathway in gingival epithelial cells. Here, we found that live P. gingivalis, but not heat-killed P. gingivalis, reduced Akt phosphorylation at both Thr-308 and Ser-473, which implies a decrease in Akt activity. Actually, PI3K, which is upstream of Akt, was also inactivated by P. gingivalis. Furthermore, glycogen synthase kinase 3 alpha/beta, mammalian target of rapamycin, and Bad, which are downstream proteins in the PI3K/Akt cascade, were also dephosphorylated, a phenomenon consistent with Akt inactivation by P. gingivalis. However, these events did not require direct interaction between bacteria and host cells and were independent of P. gingivalis invasion into the cells. The use of gingipain-specific inhibitors and a gingipain-deficient P. gingivalis mutant KDP136 revealed that the gingipains and their protease activities were essential for the inactivation of PI3K and Akt. The associations between the PI3K regulatory subunit p85 alpha and membrane proteins were disrupted by wild-type P. gingivalis. Moreover, PDK1 translocation to the plasma membrane was reduced by wild-type P. gingivalis, but not KDP136, indicating little production of phosphatidylinositol 3,4,5-triphosphate by PI3K. Therefore, it is likely that PI3K failed to transmit homeostatic extracellular stimuli to intracellular signaling pathways by gingipains. Taken together, our findings indicate that P. gingivalis attenuates the PI3K/Akt signaling pathway via the proteolytic effects of gingipains, resulting in the dysregulation of PI3K/Akt-dependent cellular functions and the destruction of epithelial barriers.

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  • Role of extracytoplasmic function sigma factors in biofilm formation of Porphyromonas gingivalis Reviewed

    Satosu Onozawa, Yuichiro Kikuchi, Kazuko Shibayama, Eitoyo Kokubu, Masaaki Nakayama, Tetsuyoshi Inoue, Keisuke Nakano, Yukinaga Shibata, Naoya Ohara, Koji Nakayama, Kazuyuki Ishihara, Toshiyuki Kawakami, Hiromasa Hasegawa

    BMC Oral Health   15   4   2015.1

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    Background: Porphyromonas gingivalis has been implicated as a major pathogen in the development and progression of chronic periodontitis. P. gingivalis biofilm formation in the subgingival crevice plays an important role in the ability of the bacteria to tolerate stress signals outside the cytoplasmic membrane. Some bacteria use a distinct subfamily of sigma factors to regulate their extracytoplasmic functions (the ECF subfamily). The objective of this study was to determine if P. gingivalis ECF sigma factors affect P. gingivalis biofilm formation.
    Methods: To elucidate the role of ECF sigma factors in P. gingivalis, chromosomal mutants carrying a disruption of each ECF sigma factor-encoding gene were constructed. Bacterial growth curves were measured by determining the turbidity of bacterial cultures. The quantity of biofilm growing on plates was evaluated by crystal violet staining.
    Results: Comparison of the growth curves of wild-type P. gingivalis strain 33277 and the ECF mutants indicated that the growth rate of the mutants was slightly lower than that of the wild-type strain. The PGN_0274- and PGN_1740-defective mutants had increased biofilm formation compared with the wild-type (p &lt; 0.001); however, the other ECF sigma factor mutants or the complemented strains did not enhance biofilm formation.
    Conclusion: These results suggest that PGN_0274 and PGN_1740 play a key role in biofilm formation by P. gingivalis.

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  • Characterization of the tripartite drug efflux pumps of Porphyromonas gingivalis ATCC 33277 Reviewed

    Tetsuyoshi Inoue, Masaaki Nakayama, Yuko Taguchi, Konami Kano, Miyuu Ono, Yurika Shimizu, Teruo Kuroda, Naoya Ohara

    NEW MICROBIOLOGICA   38 ( 1 )   101 - 107   2015.1

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    The periodontal pathogen, Porphyromonas gingivalis ATCC 33277 has six gene clusters that encode tripartite drug efflux pumps. To examine the effects of the drug efflux pumps on its antibiotic sensitivity, six mutants were constructed, each defective in the membrane fusion protein gene of each gene cluster. Compared to the wild-type strain, all mutants exhibited an elevated sensitivity to tetracycline, and two mutants with deletions in the PGN_1431 and PGN_1680 genes showed an increased sensitivity to various types of antibiotics. These results suggest that the activity of drug efflux systems may affect antibiotic sensitivity in P. gingivalis.

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  • Development of a novel plasmid vector pTIO-1 adapted for electrotransformation of Porphyromonas gingivalis Reviewed

    Junpei Tagawa, Tetsuyoshi Inoue, Mariko Naito, Keiko Sato, Tomomi Kuwahara, Masaaki Nakayama, Koji Nakayama, Takashi Yamashiro, Naoya Ohara

    JOURNAL OF MICROBIOLOGICAL METHODS   105   174 - 179   2014.10

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    We report here the construction of a plasmid vector designed for the efficient electrotransformation of the periodontal pathogen Porphyromonas gingivalis. The novel Escherichia coli-Bacteroides/P. gingivalis shuttle vector, designated pTIO-1, is based on the 11.0-kb E. coli-Bacteroides conjugative shuttle vector, pVAL-1 (a pB8-51 derivative). To construct pTIO-1, the pB8-51 origin of replication and erythromycin resistance determinant of pVAL-1 were cloned into the E. coli cloning vector pBluescript II SK(-) and non-functional regions were deleted. pTIO-1 has an almost complete multiple cloning site from pBluescript II SK(-). The size of pTIO-1 is 4.5 kb, which is convenient for routine gene manipulation. pTIO-1 was introduced into P. gingivalis via electroporation, and erythromycin-resistant transformants carrying pTIO-1 were obtained. We characterized the transformation efficiency, copy number, host range, stability, and insert size capacity of pTIO-1. An efficient plasmid electrotransformation of P. gingivalis will facilitate functional analysis and expression of P. gingivalis genes, including the virulence factors of this bacterium. (C) 2014 Elsevier B.V. All rights reserved.

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  • Hemoglobin Receptor Protein from Porphyromonas gingivalis Induces Interleukin-8 Production in Human Gingival Epithelial Cells through Stimulation of the Mitogen-Activated Protein Kinase and NF-kappa B Signal Transduction Pathways Reviewed

    Yuki Fujita, Masaaki Nakayama, Mariko Naito, Eiki Yamachika, Tetsuyoshi Inoue, Koji Nakayama, Seiji Iida, Naoya Ohara

    INFECTION AND IMMUNITY   82 ( 1 )   202 - 211   2014.1

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    Periodontitis is an inflammatory disease of polymicrobial origin affecting the tissues supporting the tooth. The oral anaerobic bacterium Porphyromonas gingivalis, which is implicated as an important pathogen for chronic periodontitis, triggers a series of host inflammatory responses that promote the destruction of periodontal tissues. Among the virulence factors of P. gingivalis, hemoglobin receptor protein (HbR) is a major protein found in culture supernatants. In this study, we investigated the roles of HbR in the production of inflammatory mediators. We found that HbR induced interleukin-8 (IL-8) production in the human gingival epithelial cell line Ca9-22. p38 mitogen-activated protein kinase (MAPK) and extracellular signal-related kinase 1/2 (Erk1/2) were activated in HbR-stimulated Ca9-22 cells. Inhibitors of p38 MAPK (SB203580) and Erk1/2 (PD98059) blocked HbR-induced IL-8 production. Additionally, HbR stimulated the translocation of NF-kappa B-p65 to the nucleus, consistent with enhancement of IL-8 expression by activation of the NF-kappa B pathway. In addition, small interfering RNA (siRNA) targeting activating transcription factor 2 (ATF-2) or cyclic AMP-response element-binding protein (CREB) inhibited HbR-induced IL-8 production. Moreover, pretreatment with SB203580 and PD98059 reduced HbR-induced phosphorylation of CREB and ATF-2, respectively. Combined pretreatment with an inhibitor of NF-kappa B (BAY11-7082) and SB203580 was more efficient in inhibiting the ability of HbR to induce IL-8 production than pretreatment with either BAY11-7082 or SB203580 alone. Thus, in Ca9-22 cells, the direct activation of p38 MAPK and Erk1/2 by HbR caused the activation of the transcription factors ATF-2, CREB, and NF-kappa B, thus resulting in the induction of IL-8 production.

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  • Evaluating efficacy of bacteriophage therapy against Staphylococcus aureus infections using a silkworm larval infection model Reviewed

    Iyo Takemura-Uchiyama, Jumpei Uchiyama, Shin ichiro Kato, Tetsuyoshi Inoue, Takako Ujihara, Naoya Ohara, Masanori Daibata, Shigenobu Matsuzaki

    FEMS Microbiology Letters   347 ( 1 )   52 - 60   2013.10

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    Silkworm larva has recently been recognized as an alternative model animal for higher mammals to evaluate the effects of antibiotics. In this study, we examined the efficacy of the bacteriophage (phage) therapy, which harnesses phages as antibacterial agents, against Staphylococcus aureus infections, using the silkworm larval infection model. Two newly isolated staphylococcal phages, S25-3 and S13′, were used as therapeutic phage candidates. They were assigned to two different lytic phage genera, Twort-like and AHJD-like viruses, based on their morphologies and the N-terminal amino acid sequences of the major capsid proteins. Both had a broad host range and strong lytic activity and showed preservative quality. Administration of these phages alone caused no adverse effects in the silkworm larvae. Moreover, the viruses showed life-prolonging effects in the silkworm larval infection model 10 min, 6 h, 12 h, and 24 h following infection. Such phage effects in the silkworm larval model were almost paralleled to the therapeutic efficacies in mouse models. These results suggest that phages S25-3 and S13′ are eligible as therapeutic candidates and that the silkworm larval model is valid for the evaluation of phage therapy as well as mouse models. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

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  • Antigen 85A and mycobacterial DNA-binding protein 1 are targets of immunoglobulin G in individuals with past tuberculosis. Reviewed

    Osada-Oka Mayuko, Tateishi Yoshitaka, Hirayama Yukio, Ozeki Yuriko, Niki Mamiko, Kitada Seigo, Maekura Ryoji, Tsujimura Kunio, Koide Yukio, Ohara Naoya, Yamamoto Taro, Kobayashi Kazuo, Matsumoto Sohkichi

    Microbiol Immunol   57 ( 1 )   30 - 37   2013.1

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    Development of accurate methods for predicting progression of tuberculosis (TB) from the latent state is recognized as vitally important in controlling TB, because a majority of cases develop from latent infections. Past TB that has never been treated has a higher risk of progressing than does latent Mycobacterium tuberculosis infection in patients who have previously received treatment. Antibody responses against 23 kinds of M. tuberculosis proteins in individuals with past TB who had not been medicated were evaluated. These individuals had significantly higher concentrations of antibodies against Antigen 85A and mycobacterial DNA-binding protein 1 (MDP1) than did those with active TB and uninfected controls. In addition, immunohistochemistry revealed colocalization of tubercle bacilli, antigen 85 and MDP1 inside tuberculous granuloma lesions in an asymptomatic subject, showing that M. tuberculosis in lesions expresses both antigen 85 and MDP1. Our study suggests the potential usefulness of measuring antibody responses to antigen 85A and MDP1 for assessing the risk of TB progression.

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  • 12-Methyltetradecanoic Acid, a Branched-Chain Fatty Acid, Represses the Extracellular Production of Surfactants Required for Swarming Motility in Pseudomonas aeruginosa PAO1 Reviewed

    Tetsuyoshi Inoue, Teruo Kuroda, Naoya Ohara

    JAPANESE JOURNAL OF INFECTIOUS DISEASES   65 ( 2 )   126 - 131   2012.3

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    Pseudomonas aeruginosa is known to produce surfactants that are involved in its swarming motility behavior, such as rhamnolipids and their precursors-3-(3-hydroxyalkanoyloxy) alkanoic acids (HAAs). In P. aeruginosa PAO1, swarming motility is inhibited by some fatty acids, including branched-chain fatty acids and unsaturated fatty acids. In the present study, addition of 12-methyltetradecanoic acid (12-MTA, anteiso-C15:0) to an agar medium markedly repressed surfactant activity in the extracellular fraction of a P. aeruginosa culture in a drop collapse assay. Further, an extracellular fraction of a culture of rhlA mutant P. aeruginosa, which did not produce both rhamnolipids and HAAs, showed a complete loss of surfactant activity and markedly reduced swarming activity. In contrast, an extracellular fraction of a culture of rhlB mutant P. aeruginosa, which produced HAAs but not rhamnolipids, showed moderate swarming activity and weak extracellular surfactant activity that was lost on the addition of 12-MTA to the agar medium. Expression of the rhlAB operon from the plasmid pMR2 restored normal swarming motility on 12-MTA-containing agar medium. Taken together, these findings indicate that 12-MTA reduced extracellular surfactant activity, thus resulting in a swarming defect in P. aeruginosa PAO1.

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  • Lipid Phenotype of Two Distinct Subpopulations of Mycobacterium bovis Bacillus Calmette-Guerin Tokyo 172 Substrain Reviewed

    Takashi Naka, Shinji Maeda, Mamiko Niki, Naoya Ohara, Saburo Yamamoto, Ikuya Yano, Jun-ichi Maeyama, Hisashi Ogura, Kazuo Kobayashi, Nagatoshi Fujiwara

    JOURNAL OF BIOLOGICAL CHEMISTRY   286 ( 51 )   44153 - 44161   2011.12

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    Bacillus Calmette-Guerin (BCG) Tokyo 172 is a predominant World Health Organization Reference Reagent for the BCG vaccine. Recently, the BCG Tokyo 172 substrain was reported to consist of two subpopulations with different colony morphologies, smooth and rough. Smooth colonies had a characteristic 22-bp deletion in Rv3405c of the region of difference (RD) 16 (type I), and rough colonies were complete in this region (type II). We hypothesized that the morphological difference is related to lipid phenotype and affects their antigenicity. We determined the lipid compositions and biosynthesis of types I and II. Scanning electron microscopy showed that type I was 1.5 times longer than type II. Phenolic glycolipid (PGL) and phthiocerol dimycocerosate (PDIM) were found only in type I. Although it has been reported that the RD16 is involved in the expression of PGL, type II did not possess PGL/PDIM. We examined the ppsA-E gene responsible for PGL/PDIM biosynthesis and found that the existence of PGL/PDIM in types I and II is caused by a ppsA gene mutation not regulated by the RD16. PGL suppressed the host recognition of total lipids via Toll-like receptor 2, and this suggests that PGL is antigenic and involved in host responses, acting as a cell wall component. This is the first report to show the difference between lipid phenotypes of types I and II. It is important to clarify the heterogeneity of BCG vaccine substrains to discuss and evaluate the quality, safety, and efficacy of the BCG vaccine.

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  • Effects of non-iron metalloporphyrins on growth and gene expression of Porphyromonas gingivalis Reviewed

    Hideharu Yukitake, Mariko Naito, Keiko Sato, Mikio Shoji, Naoya Ohara, Mamiko Yoshimura, Eiko Sakai, Koji Nakayama

    MICROBIOLOGY AND IMMUNOLOGY   55 ( 3 )   141 - 153   2011.3

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    The oral anaerobic bacterium Porphyromonas gingivalis, which is implicated as an important pathogen for chronic periodontitis, requires heme for its growth. Non-iron metalloporphyrins, In-PPIX and Ga-PPIX, were examined for antibacterial effects on P. gingivalis. Both In-PPIX and Ga-PPIX caused retardation of P. gingivalis growth in a dose-dependent fashion. Microarray and qPCR analyses revealed that In-PPIX treatment upregulated the expression of several genes encoding proteins including ClpB and ClpC, which are members of the Clp (caseinolytic protease, Hsp100) family, and aRNR, aRNR-activating protein and thioredoxin reductase, whereas In-PPIX treatment had no effect on the expression of genes encoding proteins involved in heme uptake pathways, Hmu-mediated, Iht-mediated and Tlr-mediated pathways. P. gingivalis ihtA and ihtB mutants were more resistant to In-PPIX than was the wild-type parent, whereas hmuR and tlr mutants did not show such resistance to In-PPIX. The results suggest that In-PPIX is incorporated by the Iht-mediated heme uptake pathway and that it influences protein quality control and nucleotide metabolism and retards growth of P. gingivalis.

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  • Tetratricopeptide Repeat Protein-Associated Proteins Contribute to the Virulence of Porphyromonas gingivalis Reviewed

    Yoshio Kondo, Naoya Ohara, Keiko Sato, Mamiko Yoshimura, Hideharu Yukitake, Mariko Naito, Taku Fujiwara, Koji Nakayama

    INFECTION AND IMMUNITY   78 ( 6 )   2846 - 2856   2010.6

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    Porphyromonas gingivalis is one of the most etiologically important microorganisms in periodontal disease. We found in a previous study that PG1385 (TprA) protein, a tetratricopeptide repeat (TPR) protein, was upregulated in P. gingivalis wild-type cells placed in a mouse subcutaneous chamber and that a tprA mutant was clearly less virulent in the mouse subcutaneous abscess model (M. Yoshimura et al., Oral Microbiol. Immunol. 23: 413-418, 2008). In the present study, we investigated the gene expression profile of tprA mutant cells placed in a mouse subcutaneous chamber and found that 9 genes, including PG2102 (tapA), PG2101 (tapB), and PG2100 (tapC) genes, were downregulated in the tprA mutant compared with those in the wild type. Expression of a cluster of tapA, tapB, and tapC genes of the mutant was also downregulated in an in vitro culture with enriched brain heart infusion medium. The TprA protein has three TPR motifs known as a protein-protein interaction module. Yeast two-hybrid system analysis and in vitro protein binding assays with immunoprecipitation and surface plasmon resonance detection revealed that the TprA protein could bind to TapA and TapB proteins. TprA and TapB proteins were located in the periplasmic space, whereas TapA, which appeared to be one of the C-terminal domain family proteins, was located at the outer membrane. We constructed tapA, tapB, and tapC single mutants and a tapA-tapB-tapC deletion mutant. In the mouse subcutaneous infection experiment, all of the mutants were less virulent than the wild type. These results suggest that TprA, TapA, TapB, and TapC are cooperatively involved in P. gingivalis virulence.

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  • Suppressed induction of mycobacterial antigen-specific T(h)1-type CD4(+) T cells in the lung after pulmonary mycobacterial infection Reviewed

    Ayano Yahagi, Masayuki Umemura, Toshiki Tamura, Ai Kariyone, M. Dilara Begum, Kazuyoshi Kawakami, Yuko Okamoto, Satoru Hamada, Kiyotetsu Oshiro, Hideyasu Kohama, Takeshi Arakawa, Naoya Ohara, Kiyoshi Takatsu, Goro Matsuzaki

    INTERNATIONAL IMMUNOLOGY   22 ( 4 )   307 - 318   2010.4

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    Although the importance of T(h)1-type immune response in protection against mycobacterial infection is well recognized, its regulatory mechanism in the Mycobacterium tuberculosis (Mtb)-infected lung is not well characterized. To address this issue, we analyzed kinetics of induction of mycobacterial antigen-specific CD4(+) T(h)1 T cells after mycobacterial infection in P25 TCR-transgenic (Tg) mice which express TCR alpha and beta chains from a mycobacterial Ag85B-specific MHC class II A(b)-restricted CD4(+) T-cell clone. To supply normal regulatory T-cell repertoire, we transferred normal spleen T cells into the P25 TCR-Tg mice before infection. High dose subcutaneous infection with Mtb or Mycobacterium bovis bacillus Calmette-Guerin (BCG) induced P25 TCR-Tg CD4(+) T(h)1 cells within a week. In contrast, high-dose Mtb or BCG infection into the lung failed to induce P25 TCR-Tg CD4(+) T(h)1 cells at the early stage of the infection. Furthermore, low-dose Mtb infection into the lung induced P25 TCR-Tg CD4(+) T(h)1 cells on day 21 in the mediastinal lymph node but not in the lung. IL-10 was partially involved in the suppression of T(h)1 induction in the lung because pretreatment of mice with anti-IL-10 antibody resulted in increase of P25 TCR-Tg CD4(+) T(h)1 cells in the Mtb-infected lung on day 21 of the infection, whereas neutralization of transforming growth factor-beta, another important suppressive cytokine in the lung, showed no effects on the T(h)1 induction. Our data suggest that induction of anti-mycobacterial CD4(+) T(h)1 cells is suppressed in the mycobacteria-infected lung partially by IL-10.

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  • Investigation of Contamination on Dental Surgical Goggles and Protective Spectacles by ATP Measuring Method Reviewed

    Norito Satoh, Akari Watanabe, Susumu Kokeguchi, Naoya Ohara

    Japanese Journal of Environmental Infections   25 ( 2 )   79 - 84   2010

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    Contamination of dental surgical goggles and protective spectacles was investigated using a measuring method to detect adenosine triphosphate (ATP). Mean concentration was 11 RLU (Relative Light Unit) at the beginning of the working day, but had increased to 11,638 RLU after 1 hour (t test: p&lt
    0.05). Mean concentration on the lecture measured for comparison was 46 RLU (p&lt
    0.05). Moreover, contamination of the back of the lens section of protective spectacles was 7 RLU at the beginning of the working day, but had increased to 306 RLU after 1 hour. The cleanliness factor was nearly worse than the back of the lens of dental surgical goggles (p&lt
    0.05). Choice of optic protection against transmission of infection should consider that spectacles do not offer complete protection, so dentists should wear dental surgical goggles to perform dentistry examinations. Contamination investigation using a measuring method which evaluates ATP is both simple and quick, so can be used effectively as an index of environmental contamination in a dental clinic. © 2010, Japanese Society for Infection Prevention and Control. All rights reserved.

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  • Hydrogen Sulfide Production From Cysteine and Homocysteine by Periodontal and Oral Bacteria Reviewed

    Akihiro Yoshida, Mamiko Yoshimura, Naoya Ohara, Shigeru Yoshimura, Shiori Nagashima, Tadamichi Takehara, Koji Nakayama

    JOURNAL OF PERIODONTOLOGY   80 ( 11 )   1845 - 1851   2009.11

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    Background: Hydrogen sulfide is one of the predominant volatile sulfur compounds (VSCs) produced by oral bacteria. This study developed and evaluated a system for detecting hydrogen sulfide production by oral bacteria.
    Methods: L-methionine-alpha-deamino-gamma-mercaptomethane-lyase (METase) and beta carbon-sulfur (beta C-S) lyase were used to degrade homocysteine and cysteine, respectively, to produce hydrogen sulfide. Enzymatic reactions resulting in hydrogen sulfide production were assayed by reaction with bismuth trichloride, which forms a black precipitate when mixed with hydrogen sulfide. The enzymatic activities of various oral bacteria that result in hydrogen sulfide production and the capacity of bacteria from periodontal sites to form hydrogen sulfide in reaction mixtures containing L-cysteine or DL-homocysteine were assayed.
    Results: With L-cysteine as the substrate, Streptococcus anginosus FW73 produced the most hydrogen sulfide, whereas Porphyromonas gingivalis American Type Culture Collection (ATCC) 33277 and W83 and Fusobacterium nucleatum ATCC 10953 produced similar to 35% of the amount produced by the P. gingivalis strains. Finally, the hydrogen sulfide found in subgingival plaque was analyzed. Using bismuth trichloride, the hydrogen sulfide produced by oral bacteria was visually detectable as a black precipitate.
    Conclusions: Hydrogen sulfide production by oral bacteria was easily analyzed using bismuth trichloride. However, further innovation is required for practical use. J Periodontol 2009, 80:1845-1851.

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  • Porphyromonas gingivalis mutant defective in a putative extracytoplasmic function sigma factor shows a mutator phenotype Reviewed

    Y. Kikuchi, N. Ohara, O. Ueda, K. Hirai, Y. Shibata, K. Nakayama, S. Fujimura

    ORAL MICROBIOLOGY AND IMMUNOLOGY   24 ( 5 )   377 - 383   2009.10

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    Introduction:
    Porphyromonas gingivalis is implicated as a major pathogen in the development and progression of chronic periodontitis. P. gingivalis must possess the ability to tolerate stress signals outside the cytoplasmic membrane by transcriptional activation of genes encoding proteins involved in defense or repair processes. Some bacteria utilize a distinct subfamily of sigma factors to regulate extracytoplasmic function (hence termed the ECF subfamily).
    Methods:
    To elucidate their role in P. gingivalis, a chromosomal mutant carrying a disruption of an ECF sigma factor PG1318-encoding gene was constructed. Hemagglutination and proteolytic activities were measured in the PG1318-defective mutant. Reverse transcription-polymerase chain reaction (RT-PCR) analysis and southern blot analysis were used to assess transcription of kgp in the PG1318-defective mutant. Frequency of spontaneous mutation that conferred resistance to l-trifluoromethionine was measured in the PG1318-defective mutant.
    Results:
    The PG1318-defective mutant formed non-pigmented colonies on blood agar plates at a relatively high frequency. Arginine-specific and lysine-specific proteinase activities of the non-pigmented variants were remarkably decreased compared with those of the parent strain and the pigmented variants. RT-PCR analysis showed that kgp was not transcribed in some non-pigmented variants and southern blot analysis revealed that there was a deletion in their kgp region. Frequency of mutation conferring resistance to l-trifluoromethionine was significantly higher in the PG1318-defective mutant than in the wild-type.
    Conclusion:
    These results suggest that PG1318 plays a role in the regulation of mutation frequency in the bacterium.

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  • Proteome analysis of Porphyromonas gingivalis cells placed in a subcutaneous chamber of mice Reviewed

    M. Yoshimura, N. Ohara, Y. Kondo, M. Shoji, S. Okano, Y. Nakano, Y. Abiko, K. Nakayama

    ORAL MICROBIOLOGY AND IMMUNOLOGY   23 ( 5 )   413 - 418   2008.10

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    Introduction: Porphyromonas gingivalis, an oral anaerobic bacterium, is considered a major pathogen for chronic periodontitis. Pathogenic bacteria usually upregulate or downregulate gene expression to combat the protective responses of their hosts.
    Methods: To determine what protein is regulated when P. gingivalis cells invade host tissues, we analyzed the proteome of P. gingivalis cells that were placed in a mouse subcutaneous chamber by two-dimensional gel electrophoresis and mass spectrometry.
    Results: Fourteen proteins were upregulated, while four proteins were downregulated. We focused on three upregulated proteins, PG1089 (DNA-binding response regulator RprY), PG1385 (TPR domain protein), and PG2102 (immunoreactive 61-kDa antigen), and constructed mutant strains that were defective in these proteins. Mouse abscess model experiments revealed that the mutant strain defective in PG1385 was clearly less virulent than the wild-type parent strain.
    Conclusion: These results indicate that the PG1385 protein is involved in P. gingivalis virulence and that the method used here is useful when investigating the P. gingivalis proteins responsible for virulence.

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  • Determination of the Genome Sequence of Porphyromonas gingivalis Strain ATCC 33277 and Genomic Comparison with Strain W83 Revealed Extensive Genome Rearrangements in P-gingivalis Reviewed

    Mariko Naito, Hideki Hirakawa, Atsushi Yamashita, Naoya Ohara, Mikio Shoji, Hideharu Yukitake, Keisuke Nakayama, Hidehiro Toh, Fuminobu Yoshimura, Satoru Kuhara, Masahira Hattori, Tetsuya Hayashi, Koji Nakayama

    DNA RESEARCH   15 ( 4 )   215 - 225   2008.8

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    The gram-negative anaerobic bacterium Porphyromonas gingivalis is a major causative agent of chronic periodontitis. Porphyromonas gingivalis strains have been classified into virulent and less-virulent strains by mouse subcutaneous soft tissue abscess model analysis. Here, we present the whole genome sequence of P gingivalis ATCC 33277, which is classified as a less-virulent strain. We identified 2090 protein-coding sequences (CDSs), 4 RNA operons, and 53 tRNA genes in the ATCC 3 3 2 7 7 genome. By genomic comparison with the virulent strain W83, we identified 461 ATCC 33277-specific and 415 W83-specific CDSs. Extensive genomic rearrangements were observed between the two strains: 175 regions in which genomic rearrangements have occurred were identified. Thirty-five of those genomic rearrangements were inversion or translocation and 140 were simple insertion, deletion, or replacement. Both strains contained large numbers of mobile elements, such as insertion sequences, miniature inverted-repeat transposable elements (MITEs), and conjugative transposons, which are frequently associated with genomic rearrangements. These findings indicate that the mobile genetic elements have been deeply involved in the extensive genome rearrangement of P gingivalis and the occurrence of many of the strain-specific CDSs. We also describe here a very unique feature of MITE400, which we renamed MITEPgRS (MITE of P gingivalis with Repeating Sequences).

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  • Efficacy of recombinant bacille Calmette-Guerin vaccine secreting interleukin-15/antigen 85B fusion protein in providing protection against Mycobacterium tuberculosis Reviewed

    Ce Tang, Hisakata Yamada, Kensuke Shibata, Naoyoshi Maeda, Shinichi Yoshida, Worawidh Wajjwalku, Naoya Ohara, Takeshi Yamada, Taroh Kinoshita, Yasunobu Yoshikai

    JOURNAL OF INFECTIOUS DISEASES   197 ( 9 )   1263 - 1274   2008.5

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    Protection against Mycobacterium tuberculosis not only depends on CD4(+) T helper type 1 (Th1) cells but, also, on CD8(+) T cells. Interleukin (IL)-15 has an important function in the maintenance of memory CD8(+) T cells. In the present study, we examined the efficacy of recombinant Mycobacterium bovis bacille Calmette-Guerin (rBCG) secreting fusion protein antigen (Ag) 85B murine IL-15 (rBCG-Ag85B-IL15) in providing protection against M. tuberculosis infection. The levels of major histocompatibility (MHC) class Ib (H2-M3)-binding TB2- or MHC class Ia (H-2D(b))-binding MPT64-specific CD8(+) T cells producing interferon (IFN)-gamma were significantly higher after immunization with rBCG-Ag85B-IL15 than after immunization with rBCG secreting Ag85B (rBCG-Ag85B). The levels of purified protein derivative-or Ag85B-specific CD4(+) Tcells producing IFN-gamma were also higher in mice immunized with rBCG-Ag85B-IL15 than in mice immunized with rBCG-Ag85B. Mice immunized with rBCG-Ag85B-IL15 exhibited CD8(+) and CD+ T cells responses that were stronger than those in mice immunized with rBCG-Ag85B, as well as robust protection in the lung against intratracheal challenge of M. tuberculosis. Thus, rBCG-Ag85B-IL15 vaccination capable of inducing efficient cell-mediated immunity might be used as an effective vaccine for tuberculosis.

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  • Evidence for association between a Toll-like receptor 4 gene polymorphism and moderate/severe periodontitis in the Japanese population Reviewed

    T. Fukusaki, N. Ohara, Y. Hara, A. Yoshimura, K. Yoshiura

    JOURNAL OF PERIODONTAL RESEARCH   42 ( 6 )   541 - 545   2007.12

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    Background and Objective: Chronic periodontitis is an inflammatory disease caused by bacteria in subgingival pockets. Because Toll-like receptor 2 and Toll-like receptor 4 have been shown to play an important role in the recognition of periodontal pathogens, we investigated the relevance of genetic variations in TLR2 and TLR4 to susceptibility to periodontitis.
    Material and Methods: A total of 97 patients with chronic periodontitis and 100 control subjects were examined for mutations in TLR2 and TLR4. Case-control analysis was performed using individual single nucleotide polymorphisms detected during the mutation search.
    Results: The missense mutations reported previously in TLR2 (677 Arg &gt; Trp and 753 Arg &gt; Gln) and in TLR4 (299 Asp &gt; Gly and 399 Thr &gt; Ile) were not detected in 97 of the Japanese patients with chronic periodontitis or in 100 of the Japanese control subjects. Nine single nucleotide polymorphisms were identified in exons of TLR2 and TLR4. The case-control analysis revealed that the frequency of the C/C genotype at base-pair position +3725 in TLR4 was significantly higher in both the moderate and the severe periodontitis patient group than in the control group.
    Conclusion: A genetic variation of TLR4 might be associated with moderate and severe periodontitis in the Japanese population.

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  • Molecular analysis of RANKL-independent cell fusion of osteoclast-like cells induced by TNF-alpha, lipopolysaccharide, or peptidoglycan. Reviewed International journal

    Hitoshi Hotokezaka, Eiko Sakai, Naoya Ohara, Yuka Hotokezaka, Carmen Gonzales, Ken-ichiro Matsuo, Yuji Fujimura, Noriaki Yoshida, Koji Nakayama

    Journal of cellular biochemistry   101 ( 1 )   122 - 34   2007.5

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    Focusing on the final step of osteoclastogenesis, we studied cell fusion from tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells into multinuclear cells. TRAP-positive mononuclear cells before generation of multinuclear cells by cell fusion were differentiated from RAW264.7 cells by treatment with receptor activator of nuclear factor kappa B ligand (RANKL), and then the cells were treated with lipopolysaccharide (LPS), followed by culturing for further 12 h. LPS-induced cell fusion even in the absence of RANKL. Similarly, tumor necrosis factor (TNF)-alpha and peptidoglycan (PGN) induced cell fusion, but M-CSF did not. The cell fusion induced by RANKL, TNF-alpha, and LPS was specifically blocked by osteoprotegerin (OPG), anti-TNF-alpha antibody, and polymyxin B, respectively. LPS- and PGN-induced cell fusion was partly inhibited by anti-TNF-alpha antibody but not by OPG. When TRAP-positive mononuclear cells fused to yield multinuclear cells, phosphorylation of Akt, Src, extracellular signal-regulated kinase (ERK), p38MAPK (p38), and c-Jun NH2-terminal kinase (JNK) was observed. The specific chemical inhibitors LY294002 (PI3K), PP2 (Src), U0126 (MAPK-ERK kinase (MEK)/ERK), and SP600125 (JNK) effectively suppressed cell fusion, although SB203580 (p38) did not. mRNA of nuclear factor of activated T-cells c1 (NFATc1) and dendritic cell-specific transmembrane protein (DC-STAMP) during the cell fusion was quantified, however, there was no obvious difference among the TRAP-positive mononuclear cells treated with or without M-CSF, RANKL, TNF-alpha, LPS, or PGN. Collectively, RANKL, TNF-alpha, LPS, and PGN induced cell fusion of osteoclasts through their own receptors. Subsequent activation of signaling pathways involving PI3K, Src, ERK, and JNK molecules was required for the cell fusion. Although DC-STAMP is considered to be a requisite for cell fusion of osteoclasts, cell fusion-inducing factors other than DC-STAMP might be necessary for the cell fusion.

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  • The hemoglobin receptor protein of porphyromonas gingivalis inhibits receptor activator NF-kappaB ligand-induced osteoclastogenesis from bone marrow macrophages. Reviewed International journal

    Yuji Fujimura, Hitoshi Hotokezaka, Naoya Ohara, Mariko Naito, Eiko Sakai, Mamiko Yoshimura, Yuka Narita, Hideki Kitaura, Noriaki Yoshida, Koji Nakayama

    Infection and immunity   74 ( 5 )   2544 - 51   2006.5

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    Extracellular proteinaceous factors of Porphyromonas gingivalis, a periodontal pathogen, that influence receptor activator of nuclear factor-kappaB (NF-kappaB) ligand (RANKL)-induced osteoclastogenesis from bone marrow macrophages were investigated. The culture supernatant of P. gingivalis had the ability to inhibit RANKL-induced in vitro osteoclastogenesis. A major protein of the culture supernatant, hemoglobin receptor protein (HbR), suppressed RANKL-induced osteoclastogenesis in a dose-dependent fashion. HbR markedly inhibited RANKL-induced osteoclastogenesis when present in the culture for the first 24 h after addition of RANKL, whereas no significant inhibition was observed when HbR was added after 24 h or later, implying that HbR might interfere with only the initial stage of RANKL-mediated differentiation. HbR tightly bound to bone marrow macrophages and had the ability to induce phosphorylation of ERK, p38, NF-kappaB, and Akt. RANKL-induced phosphorylation of ERK, p38, and NF-kappaB was not suppressed by HbR, but that of Akt was markedly suppressed. HbR inhibited RANKL-mediated induction of c-Fos and NFATc1. HbR could induce beta interferon (IFN-beta) from bone marrow macrophages, but the induction level of IFN-beta might not be sufficient to suppress RANKL-mediated osteoclastogenesis, implying presence of an IFN-beta-independent pathway in HbR-mediated inhibition of osteoclastogenesis. Since rapid and extensive destruction of the alveolar bone causes tooth loss, resulting in loss of the gingival crevice that is an anatomical niche for periodontal pathogens such as P. gingivalis, the suppressive effect of HbR on osteoclastogenesis may help the microorganism exist long in the niche.

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  • Superoxide dismutase-encoding gene of the obligate anaerobe Porphyromonas gingivalis is regulated by the redox-sensing transcription activator OxyR Reviewed

    N Ohara, Y Kikuchi, M Shoji, M Naito, K Nakayama

    MICROBIOLOGY-SGM   152 ( Pt 4 )   955 - 966   2006.4

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    Inspection of the genomic DNA sequence of the oral anaerobe Porphyromonas gingivalis reveals that the micro-organism possesses the peroxide-sensing transcription activator OxyR, but not the superoxide-sensing transcription factor SoxR. Investigatation of oxiclative-stress-responsive proteins in P. gingivalis by two-dimensional gel electrophoresis showed that two proteins were predominantly upregulated in oxidative conditions. In a P. gingivalis oxyR mutant these two proteins were not induced by treatment with hydrogen peroxide under aerobic conditions. By N-terminal amino acid sequencing, the two proteins were found to be superoxide dismutase and alkyl hydroperoxide reductase, encoded by sod and ahpC, respectively. Northern blot and lacZ fusion analyses revealed that P. gingivalis sod and ahpC were positively regulated by OxyR. Primer extension analysis located the promoter regions of sod and ahpC, and putative -35 boxes of these promoters were found immediately adjacent to their putative OxyR-binding sequences. Moreover, the promoter regions of sod and ahpC had the ability to bind P. gingivalis OxyR protein. These results demonstrate that P. gingivalis sod is one of the OxyR regulons, suggesting that OxyR functions as an intracellular redox sensor rather than a peroxide sensor in this organism. A sod gene of Bacteroides fragilis, which is taxonomically related to P. gingivalis, is inducible by redox stresses but not controlled by its OxyR. A DNA fragment including the B. fragilis sod promoter region could bind the P. gingivalis OxyR protein; however, a putative OxyR binding sequence within the DNA fragment was 14 bases distant from a putative -35 box of its promoter.

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  • Mutant Escherichia coli enterotoxin as a mucosal adjuvant induces specific Th1 responses of CD4(+) and CD8(+) T cells to nasal killed-bacillus calmette-guerin in mice Reviewed

    H Takahashi, K Sasaki, M Takahashi, N Shigemori, S Honda, H Arimitsu, S Ochi, N Ohara, T Tsuji

    VACCINE   24 ( 17 )   3591 - 3598   2006.4

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    On single nasal immunization of mice with killed-bacillus calmette-guerin (BCG) plus a mutant Escherichia coli enterotoxin, delayed-type hypersensitivity was induced and BCG-infection decreased. Spleen cells, particularly CD4(+) T cells among them produced IL-2, IFN gamma and TNF alpha in response to the killed-BCG or purified protein derivatives. CD8(+) T cells including cytotoxic T lymphocytes produced IFN gamma and TNF alpha. However, both types of T cells reacted a little to Ag85B.
    The mutant induces cellular immunity to nasal killed-BCG vaccine and decreases BCG-infection. CD4(+) and CD8(+) T cells produce cytokines effective for tuberculosis. Although killed-BCG loses some antigens like Ag85B, nasal killed-BCG plus the mutant is useful for tuberculosis. (c) 2006 Elsevier Ltd. All rights reserved.

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  • Dissecting the role of Rho-mediated signaling in contractile ring Formation Reviewed

    K Kamijo, N Ohara, M Abe, T Uchimura, H Hosoya, JS Lee, T Miki

    MOLECULAR BIOLOGY OF THE CELL   17 ( 1 )   43 - 55   2006.1

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    In anaphase, microtubules provide a specification signal for positioning of the contractile ring. However, the nature of the signal remains unknown. The small GTPase Rho is a potent regulator of cytokinesis, but the involvement of Rho in contractile ring formation is disputed. Here, we show that Rho serves as a microtubule-dependent signal that specifies the position of the contractile ring. We found that Rho translocates to the equatorial region before furrow ingression. The Rho-specific inhibitor C3 exoenzyme and small interfering RNA to the Rho GDP/GTP exchange factor ECT2 prevent this translocation and disrupt contractile ring formation, indicating that active Rho is required for contractile ring formation. ECT2 forms a complex with the GTPase-activating protein MgcRacGAP and the kinesinlike protein MKLP1 at the central spindle, and the localization of ECT2 at the central spindle depends on MgcRacGAP and MKLP1. In addition, we show that the bundled microtubules direct Rho-mediated signaling molecules to the furrowing site and regulate furrow formation. Our study provides strong evidence for the requirement of Rho-mediated signaling in contractile ring formation.

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  • Porphyromonas gingivalis-induced platelet aggregation in plasma depends on Hgp44 adhesin but not Rgp proteinase Reviewed

    M Naito, E Sakai, YX Shi, H Ideguchi, M Shoji, N Ohara, K Yamamoto, K Nakayama

    MOLECULAR MICROBIOLOGY   59 ( 1 )   152 - 167   2006.1

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    Evidence from recent epidemiological studies suggests a link between periodontal infections and increased risk of atherosclerosis and related cardiovascular and cerebrovascular events in human subjects. One of the major pathogens of periodontitis, Porphyromonas gingivalis, has the ability to aggregate human platelets in platelet-rich plasma (PRP). Mechanism of P. gingivalis-induced platelet aggregation in PRP was investigated. Proteinase inhibitors toward Arg-gingipain (Rgp) and Lys-gingipain (Kgp) did not suppress P. gingivalis-induced platelet aggregation in PRP, whereas the Rgp inhibitor markedly inhibited P. gingivalis-induced platelet aggregation using washed platelets. Mutant analysis revealed that P. gingivalis-induced platelet aggregation in PRP depended on Rgp-, Kgp- and haemagglutinin A (HagA)-encoding genes that intragenically coded for adhesins such as Hgp44. Hgp44 adhesin on the bacterial cell surface, which was processed by Rgp and Kgp proteinases, was essential for P. gingivalis-induced platelet aggregation in PRP. P. gingivalis cell-reactive IgG in plasma, and Fc gamma RIIa receptor and to a lesser extent GPIb alpha receptor on platelets were found to be a prerequisite for P. gingivalis-induced platelet aggregation in PRP. These results reveal a novel mechanism of platelet aggregation by P. gingivalis.

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  • Analysis of amphotericin B-induced cell signaling with chemical inhibitors of signaling molecules. Reviewed International journal

    Kenichiro Matsuo, Hitoshi Hotokezaka, Naoya Ohara, Yuji Fujimura, Atsutoshi Yoshimura, Yukio Okada, Yoshitaka Hara, Noriaki Yoshida, Koji Nakayama

    Microbiology and immunology   50 ( 4 )   337 - 47   2006

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    Although amphotericin B (AmB) is a major polyene antibiotic against invasive fungal infection, administration to patients sometimes causes inflammatory side effects, which limits the usage of the antibiotic. We studied the intracellular signaling that was induced by AmB. p65 (RelA) of nuclear factor-kappaB (NF-kappaB), a well-known signaling molecule as an inducer of proinflammatory cytokines, was phosphorylated by AmB in RAW264.7 cells, a monocyte-like cell line. Among chemical inhibitors of signaling molecules, U-73122 (phospholipase C (PLC) inhibitor), Gö6976 (protein kinase C (PKC) inhibitor), BAPTA-AM (calcium chelator), LFM-A13 (Bruton's tyrosine kinase (Btk)-specific inhibitor), and PP2 (c-Src kinase inhibitor) suppressed AmB-induced phosphorylation of p65 and translocation of p65 into the nucleus. U-73122 and Gö6976 reduced AmB-mediated induction of proinflammatory cytokines (tumor necrosis factor (TNF)-alpha and interleukin (IL)-6) in RAW264.7 cells. Furthermore, AmB-induced activation of NF-kappaB was observed in toll-like receptor (TLR) 2-expressed cells, and the activation of NF-kappaB was inhibited by U-73122, whereas peptidoglycan-induced NF-kappaB activation, which was also dependent on TLR2, was not inhibited by U-73122. Finally, U-73122 partially suppressed in vivo production of TNF-alpha and IL-6 induced by AmB administration in BALB/c mice. These results suggested that the signaling from AmB stimulation to proinflammatory cytokine production is mediated by TLR2, Btk, PLC, PKC, c-Src and NF-kappaB. These signaling molecules may become a target for chemotherapy suppressing AmB-induced proinflammatory cytokine production.

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  • [Study on antigenicity and pathogenicity of mycobacteria]. Reviewed

    Ohara N

    Nihon saikingaku zasshi. Japanese journal of bacteriology   60 ( 2 )   349 - 356   2005.5

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  • Identification of a new membrane-associated protein that influences transport/maturation of gingipains and adhesins of Porphyromonas gingivalis Reviewed

    K Sato, E Sakai, PD Veith, M Shoji, Y Kikuchi, H Yukitake, N Ohara, M Naito, K Okamoto, EC Reynolds, K Nakayama

    JOURNAL OF BIOLOGICAL CHEMISTRY   280 ( 10 )   8668 - 8677   2005.3

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    The dual membrane envelopes of Gram-negative bacteria provide two barriers of unlike nature that regulate the transport of molecules into and out of organisms. Organisms have developed several systems for transport across the inner and outer membranes. The Gram-negative periodontopathogenic bacterium Porphyromonas gingivalis produces proteinase and adhesin complexes, gingipains/adhesins, on the cell surface and in the extracellular milieu as one of the major virulence factors. Gingipains and/or adhesins are encoded by kgp, rgpA, rgpB, and hagA on the chromosome. In this study, we isolated a P. gingivalis mutant (porT), which showed very weak activities of gingipains in the cell lysates and culture supernatants. Subcellular fractionation and immunoblot analysis demonstrated that precursor forms of gingipains and adhesins were accumulated in the periplasmic space of the porT mutant cells. Peptide mass fingerprinting and N-terminal amino acid sequencing of the precursor proteins and the kgp'-'rgpB chimera gene product in the porT mutant indicated that these proteins lacked the signal peptide regions, consistent with their accumulation in the periplasm. The PorT protein seemed to be membrane-associated and exposed to the periplasmic space, as revealed by subcellular fractionation and immunoblot analysis using anti-PorT antiserum. These results suggest that the membrane-associated protein PorT is essential for transport of the kgp, rgpA, rgpB, and hagA gene products across the outer membrane from the periplasm to the cell surface, where they are processed and matured.

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  • Novel stationary-phase-upregulated protein of Porphyromonas gingivalis influences production of superoxide dismutase, thiol peroxidase and thioredoxin Reviewed

    Y Kikuchi, N Ohara, K Sato, M Yoshimura, H Yukitake, E Sakai, M Shoji, M Naito, K Nakayama

    MICROBIOLOGY-SGM   151 ( Pt 3 )   841 - 853   2005.3

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    Porphyromonas gingivalis, an obligately anaerobic bacterium, is implicated as a major pathogen in the development and progression of chronic periodontitis. Although expression of several virulence factors of the bacterium has been found to be affected by environmental stress such as entrance into the stationary growth phase and heat, there is relatively little information on the mechanisms that may operate in the bacterium in response to environmental stress. In this study, a novel protein (UstA) was investigated that was initially identified following two-dimensional gel analysis. Expression of UstA was upregulated in stationary phase or by exposure to atmospheric oxygen. N-terminal sequencing and database analysis with the P. gingivalis genome sequence revealed that the UstA-encoding gene (ustA) was located upstream of a homologue of the usp gene encoding the universal stress protein on the chromosome. The ustA gene appeared to be transcribed in a monocistronic fashion, as revealed by primer extension and Northern blot analysis. To elucidate the role of UstA in the bacterium, chromosomal mutants carrying a disruption of the ustA gene were constructed. The ustA mutant grew slower than the wild-type parent strain in rich medium, resulting in a lower yield in stationary phase. Furthermore, in this mutant, expression levels of the P. gingivalis homologues of superoxide dismutase, thiol peroxidase and thioredoxin were markedly higher than those in the wild-type, especially in stationary phase. The ustA mutant was more resistant to diamide, a thiol-specific oxidant, than the wild-type. In addition, the ustA mutation suppressed hypersensitivities of the oxyR mutant to diamide, metronidazole and mitomycin C. These results suggest that UstA may play a significant role in oxidative stress responses in the bacterium.

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  • Novel recombinant BCG and DNA-vaccination against tuberculosis in a cynomolgus monkey Reviewed

    Y Kita, T Tanaka, S Yoshida, N Ohara, Y Kaneda, S Kuwayama, Y Muraki, N Kanamaru, S Hashimoto, H Takai, C Okada, Y Fukunaga, Y Sakaguchi, IN Furukawa, K Yamada, Y Inoue, Y Takemoto, M Naito, T Yamada, M Matsumoto, DN McMurray, EC Dela Cruz, EV Tan, RM Abalos, JA Burgos, R Gelber, Y Skeiky, S Reed, M Sakatani, M Okada

    VACCINE   23 ( 17-18 )   2132 - 2135   2005.3

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    We have developed two novel tuberculosis (TB) vaccines: a DNA vaccine combination expressing mycobacterial heat shock protein 65 (Hsp65) and interleukin-12 (IL-12) by using the hemagglutinating virus of Japan (HVJ)-liposome (HSP65 + IL-12/HVJ) and a recombinant BCG harboring the 72f fusion gene (72f rBCG). These vaccines provide remarkable protective efficacy in mouse and guinea pig models, as compared to the current by available BCG vaccine. In the present study, we extended our studies to a cynomolgus monkey model, which is currently the best animal model of human tuberculosis, to evaluate the HSP65 + IL-12/HVJ and 72f rBCG vaccines. Vaccination with HSP65 + IL-12/HVJ as well as 72f rBCG vaccines provided better protective efficacy as assessed by the Erythrocyte Sedimentation Rate, chest X-ray findings and immune responses than BCG. Most importantly, HSP65 + IL-12/HVJ resulted in an increased survival for over a year. This is the first report of successful DNA vaccination and recombinant BCG vaccination against M. tuberculosis in the monkey model. (c) 2005 Elsevier Ltd. All rights reserved.

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  • The major structural components of two cell surface filaments of Porphyromonas gingivalis are matured through lipoprotein precursors Reviewed

    M Shoji, M Naito, H Yukitake, K Sato, E Sakai, N Ohara, K Nakayama

    MOLECULAR MICROBIOLOGY   52 ( 5 )   1513 - 1525   2004.6

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    Bacterial cell surface filaments play significant roles in adherence to and invasion of host cells. They are generated by the chaperone/usher pathway system (class I fimbriae), the type II secretion system (type IV pili) and the nucleation-dependent polymerization system (Curli filaments) that are categorized by their modes of expression and assembly. In this study, we found that the periodontal pathogen Porphyromonas gingivalis expressed the major structural components of two cell surface filaments (fimbrilin and the 75 kDa protein) that had extremely long prosequences in their primary gene products. N-terminal amino acid sequencing of the prosequences, treatment of P. gingivalis cells with globomycin, an inhibitor for lipoprotein-specific signal peptidase, amino acid substitution of the cysteine residue of the prosequence of fimbrilin and [H-3]-palmitic acid labelling implied that fimbrilin and the 75 kDa protein were matured through their lipoprotein precursor forms. Accumulation of precursor forms of fimbrilin and the 75 kDa protein on the cell surface of the gingipain-null mutant revealed that Arg-gingipain processed these precursors on the surface to yield their mature forms, which subsequently assembled into the filamentous structures, suggesting that the transport and assembly of the major component proteins appear to be novel.

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  • Infection-induced up-regulation of the costimulatory molecule 4-1BB in osteoblastic cells and its inhibitory effect on M-CSF/RANKL-induced in vitro osteoclastogenesis. Reviewed International journal

    Kan Saito, Naoya Ohara, Hitoshi Hotokezaka, Satoshi Fukumoto, Kenji Yuasa, Mariko Naito, Taku Fujiwara, Koji Nakayama

    The Journal of biological chemistry   279 ( 14 )   13555 - 63   2004.4

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    Bacterial infection sometimes impairs bone metabolism. In this study, we infected the osteoblastic cell line MC3T3-E1 with Mycobacterium bovis bacillus Calmette-Guérin (BCG) and identified genes that were up-regulated in the BCG-infected cells by the suppression subtractive hybridization method. A gene encoding 4-1BB (CD137), a member of the tumor necrosis factor-alpha receptor family, was found to be one of the up-regulated genes. Up-regulation of 4-1BB was also observed by infection with Escherichia coli, Salmonella typhimurium, and Staphylococcus aureus, and by treatment with lipopolysaccharides and heat-killed BCG. Bone marrow cells and the macrophage-like cell lines J774 and RAW264.7 were found to express 4-1BB ligand (4-1BBL). Recombinant 4-1BB (r4-1BB) that was immobilized on culture plates strongly inhibited macrophage colony stimulating factor (M-CSF)/receptor activator of nuclear factor-kappaB ligand (RANKL)-induced in vitro osteoclast formation from bone marrow cells. Anti-4-1BBL antibody also inhibited osteoclast formation to a lesser extent, indicating involvement of reverse signaling through 4-1BBL during inhibition of osteoclast formation. A casein kinase I (CKI) inhibitor markedly suppressed the inhibitory effect of r4-1BB on M-CSF/RANKL-induced osteoclast formation, suggesting that CKI might be involved in 4-1BB/4-1BBL reverse signaling. r4-1BB showed no effects on M-CSF- or RANKL-induced phosphorylation of I-kappaB, ERK1/2, p38, or JNK, whereas RANKL-induced phosphorylation of Akt, a downstream target of phosphatidylinositol 3-kinase (PI3K), was completely abolished by r4-1BB, suggesting that 4-1BB/4-1BBL reverse signaling may interfere with PI3K/Akt pathway. r4-1BB also abolished RANKL-mediated induction of nuclear factor of activated T cells-2. This study may elucidate a novel role of 4-1BB in cell metabolism, especially osteoclastogenesis.

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  • Induction of protective cellular immunity against Mycobacterium tuberculosis by recombinant attenuated self-destructing Listeria monocytogenes strains harboring eukaryotic expression plasmids for antigen 85 complex and MPB/MPT51 Reviewed

    K Miki, T Nagata, T Tanaka, YH Kim, M Uchijima, N Ohara, S Nakamura, M Okada, Y Koide

    INFECTION AND IMMUNITY   72 ( 4 )   2014 - 2021   2004.4

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    We report here the induction of specific protective cellular immunity against Mycobacterium tuberculosis by the employment of vaccination with recombinant attenuated Listeria monocytogenes strains. We constructed self-destructing attenuated L. monocytogenes Delta2 strains carrying eukaryotic expression plasmids for the antigen 85 complex (Ag85A and Ag85B) and for MPB/MPT51 (mycobacterial protein secreted by M. bovis BCG/mycobacterial protein secreted by M. tuberculosis) molecules. Infection of these recombinant bacteria allowed expression of the genes in the J774A.1 murine macrophage cell line. Intraperitoneal vaccination of C57BL/6 mice with these recombinant bacteria,was capable of inducing purified protein derivative-specific cellular immune responses, such as foot pad reactions, proliferative responses of splenocytes, and gamma interferon production from splenocytes, suggesting the efficacy of vaccination against mycobacterial infection by use of these recombinant L. monocytogenes strains. Furthermore, intravenous vaccination with recombinant bacteria carrying expression plasmids for Ag85A, Ag85B, or MPB/MPT51 in BALB/c mice elicited significant protective responses, comparable to those evoked by a live Mycobacterium bovis BCG vaccine. Notably, this is the first report to show that MPB/MPT51 is a major protective antigen in addition to Ag85A and Ag85B, which have been reported to be major mycobacterial protective antigens.

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  • Deregulation and mislocalization of the cytokinesis regulator ECT2 activate the Rho signaling pathways leading to malignant transformation Reviewed

    S Saito, XF Liu, K Kamijo, R Raziuddin, T Tatsumoto, Okamoto, I, XY Chen, CC Lee, MV Lorenzi, N Ohara, T Miki

    JOURNAL OF BIOLOGICAL CHEMISTRY   279 ( 8 )   7169 - 7179   2004.2

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    The human ECT2 protooncogene encodes a guanine nucleotide exchange factor for the Rho GTPases and regulates cytokinesis. Although the oncogenic form of ECT2 contains an N-terminal truncation, it is not clear how the structural abnormality of ECT2 causes malignant transformation. Here we show that both the removal of the negative regulatory domain and alteration of subcellular localization are required to induce the oncogenic activity of ECT2. The transforming activity of oncogenic ECT2 was strongly inhibited by dominant negative Rho GTPases, suggesting the involvement of Rho GTPases in ECT2 transformation. Although deletion of the N-terminal cell cycle regulator-related domain (N) of ECT2 did not activate its transforming activity, removal of the small central domain (S), which contains two nuclear localization signals (NLSs), significantly induced the activity. The ECT2 N domain interacted with the catalytic domain and significantly inhibited the focus formation by oncogenic ECT2. Interestingly, the introduction of the NLS mutations in the S domain of N-terminally truncated ECT2 dramatically induced the transforming activity of this otherwise nononcogenic derivative. Among the known Rho GTPases expressed in NIH 3T3 cells, RhoA was predominantly activated by oncogenic ECT2 in vivo. Therefore, the mislocalization of structurally altered ECT2 might cause the untimely activation of cytoplasmic Rho GTPases leading to the malignant transformation.

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  • MgcRacGAP regulates cortical activity through RhoA during cytokinesis Reviewed

    JS Lee, K Kamijo, N Ohara, T Kitamura, T Miki

    EXPERIMENTAL CELL RESEARCH   293 ( 2 )   275 - 282   2004.2

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    Although Rho GTPases regulate multiple cellular events, their role in cell division is still obscure. Here we show that expression of a GTPase-activating protein (GAP)-deficient mutant (R386A) of the Rho regulator MgcRacGAP induces abnormal cortical activity during cytokinesis in U20S cells. Multiple large blebs were observed in cells expressing MgcRacGAP R386A from the onset of anaphase to the late stage of cell division. When mitotic blebbing was excessive, cytokinesis was inhibited, and cells with micronuclei were generated. It has been reported that blebbing is caused by abnormal cortical activity. The MgcRacGAP R386A-induced abnormal cortical activity was inhibited by the dominant negative form of RhoA, but not Rac1 or Cdc42. Moreover, expression of constitutively active RhoA also induced drastic cortical activity during cytokinesis. Unlike apoptotic blebbing, MgcRacGAP R386A-induced blebbing was not inhibited by the ROCK inhibitor Y-27632, suggesting that MgcRacGAP regulates cortical activity during cytokinesis through a novel signaling pathway. We propose that MgcRacGAP plays a pivotal role in cytokinesis by regulating cortical movement through RhoA. (C) 2003 Elsevier Inc. All rights reserved.

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  • Novel (recombinant BCG- and DNA-) vaccination against tuberculosis using cynomolgus monkey Reviewed

    Kita, I, T Tanaka, S Yoshida, N Ohara, Y Kaneda, Kuwayama, I, Muraki, I, Kanamaru, I, S Hashimoto, H Takai, C Okada, Y Fukunaga, Y Sakaguchi, Furukawa, I, K Yamada, Inoue, I, Takemoto, I, M Naito, T Yamada, M Matsumoto, EC Cruz, EV Tan, RM Abalos, LJ Young, JA Burgos, R Gelber, Y Skeiky, S Reed, M Sakatani, M Okada

    IMMUNOLOGY 2004: IMMUNODEFICIENCY, INFECTIOUS DISEASES, IMMUNOMODULATION, AND VACCINES   403 - 406   2004

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    HVJ-liposome / HSP65 DNA + IL-12 DNA vaccination were 100 fold more efficient than BCG on the elimination of Mycobacterium tuberculosis (M,TB) in lungs, liver, and spleen in he BALB/c mice. Cytotoxic T cells activity against M.TB in the mice was augmented. The recombinant(r) 72f BCG vaccine as well as HSP65 DNA + IL-12 DNA vaccine showed stronger anti-TB immunity than BCG in the mice, and guinea pigs. By using these new vaccines (HSP65 DNA + IL-12 DNA, r72f BCG and 72f fusion protein + BCG) and the cynomolgus monkey models which are very similar to human tuberculosis, the prophylactic effect (survival, Erythrocyte Sedimentation Rate, chest X-P finding, immune responses) of vaccines was observed. Thus, these novel vaccines should provide a useful tool for the prevention of human TB infection.

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  • Changed activation of HIV-1 LTR in monocytoid cells by mycobacteria with temporal progression of infection Reviewed

    H Kitaura, N Ohara, N Nakao, N Yoshida, T Yamada

    MICROBIOLOGICA   25 ( 3 )   357 - 361   2002.7

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    Coincubation of monocytoid cell line U937 cells cotransfected with HIV-1 LTR CAT plasmid and Tat expression plasmid, with Mycobacterium smegmatis, M. avium, M. bovis BCG and M. tuberculosis enhanced chloramphenicol acetyltransferase (CAT) production, indicating that these mycobacteria could activate the LTR in this cell line. The amount of CAT in the cells coincubated with M. smegmatis was higher than that infected with the other mycobacteria after 12, 24 and 48 hour time periods. However, the amount of CAT production in the cells cocultured with M. tuberculosis was higher than those coincubated with the other mycobacteria at 72 hours. These findings indicated that avirulent mycobacteria such as M. smegmatis may activate HIV replication at an early time and its effects are gradually decreased, while the effect of virulent M. tuberculosis increased gradually, and lasted for a long time resulting in an acceleration of HIV disease in patients.

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  • Discovery of immunostimulatory CpG-DNA and its application to tuberculosis vaccine development Reviewed

    S Yamamoto, T Yamamoto, Y Nojima, K Umemori, S Phalen, DN McMurray, E Kuramoto, S Iho, R Takauji, Y Sato, T Yamada, N Ohara, S Matsumoto, Y Goto, K Matsuo, T Tokunaga

    JAPANESE JOURNAL OF INFECTIOUS DISEASES   55 ( 2 )   37 - 44   2002.4

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    DNA containing an unmethylated CpG motif has a potent immunostimulatory effect on the vertebrate immune system. Because such CpG motifs are relatively common in bacterial DNA, but rare in mammalian animal and plant DNA, they may be an evolutionary adaptation augmenting innate immunity, most likely in response to pathogens that replicate within the host cells, such as viruses and intracellular bacteria. Microbial infection induces innate immunity by triggering pattern-recognition systems. The infected cells produce proinflammatory cytokines that directly combat microbial invaders and express costimulating surface molecules, which develop adaptive immunity by inducing distinct T cell differentiation. Bacterial DNA with unmethylated CpG-DNA stimulates vertebrate immature immune cells to induce maturation and to produce TNF-alpha as well as Th1-type cytokines, IL-12 and IFN-gamma. Therefore, CpG-DNA functions as an adjuvant for regulating the initiation of Th1 differentiation. The roles of immunostimulatory CpG motifs in DNA vaccine developments and in therapeutic applications have been discussed.

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  • Expression of alkaline phosphatase induces rapid and artificial mineralization in specific transformed Escherichia coli Reviewed

    N Ohara, N Ohara, K Yanagiguchi, S Yamada, IL Viloria, Y Hayashi

    MICROBIOLOGICA   25 ( 1 )   107 - 110   2002.1

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    Matrix vesicles (MV) having high alkaline phosphatase (ALP) activity act as initiators of biological mineralization. Although bacteria have similar membranous structures to MV. ALP mediated mineralization has not been studied in bacterial cells. Escherichia coli was transformed with a bacterial ALP gene in this study. Recombinant E. coli overproducing ALP induced mineralization through hydrolysis of calcium-glycerophosphate (Ca-GP). Fourier transform infrared spectroscopy and electron microscopy combined with electron diffraction revealed newly formed hydroxyapatite mineral deposits. These findings suggest that hydrolysis of Ca-GP through ALP induced high Ca and Pi concentrations within bacterial cells followed by complete bacterial mineralization.

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  • TNF-alpha-mediated activation of HIV-1 LTR in monocytoid cells by mycobacteria Reviewed

    H Kitaura, N Ohara, K Kobayashi, T Yamada

    FEMS IMMUNOLOGY AND MEDICAL MICROBIOLOGY   31 ( 2 )   97 - 103   2001.8

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    Mycobacterial infection occurs commonly in patients with acquired immune deficiency syndrome. Incubation of monocytoid cell line U937 cells, which was cotransfected HIV-1 long terminal repeat sequence (LTR) chloramphenicol acetyltransferase (CAT) plasmid and Tat expression plasmid, with mycobacterium smegmatis, Mycobacterium avium, Mycobacterium bovis BCG and Mycobacterium tuberculosis resulted in enhancement of CAT production, indicating that these mycobacteria could activate LTR in this cell line. The amount of CAT in the cells coexisting with M. smegmatis was higher than that infected with other mycobacteria. The amounts of CAT production in the cells coculturing with M. avium and M. bovis BCG were intermediate. M. tuberculosis slightly stimulated CAT production. The amount of tumor necrosis factor (TNF)-alpha produced by transfected U937 cells was correlated with the amount of CAT production. The interleukin (IL)-1 beta and IL-6 levels in the supernatant from coculturing with all species were similar. The antibody to TNF-alpha inhibited CAT production induced by mycobacterial infections. The anti-IL-1 beta and anti-IL-6 antibodies, however, scarcely influenced stimulation of LTR by mycobacteria. In addition, U937 cells transfected with full length LTR CAT plasmid showed increased CAT production by activation with mycobacteria, but the cells transfected with mutant LTR CAT constructs from which the nuclear factor (NF)-KB binding site was deleted did not show activation. These findings indicated that activation of Mycobacterium-induced LTR CAT is NF-KB dependent. These findings suggested that activation of HIV-1 LTR by mycobacteria was mainly mediated by NF-KB-induced secondary release of cytokine TNF-alpha. (C) 2001 Federation of European Microbiological Societies. Published by Elsevier Science BN. All rights reserved.

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  • Recombinant BCG vaccines Invited Reviewed

    N Ohara, T Yamada

    VACCINE   19 ( 30 )   4089 - 4098   2001.7

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  • TNF-alpha-mediated multiplication of human immunodeficiency virus in chronically infected monocytoid cells by mycobacterial infection Reviewed

    H Kitaura, N Ohara, K Kobayashi, T Yamada

    APMIS   109 ( 7-8 )   533 - 540   2001.7

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    Mycobacterial infection is a common occurrence in patients with acquired immune deficiency syndrome. Incubation of U1, a chronically HIV-1-infected human promonocytic cell line, with Mycobacterium smegmatis, M. avium, M. bovis BCG and M. tuberculosis resulted in enhancement of p24 antigen release in the supernatant, indicating that these mycobacteria could activate HIV replication from this cell line. The amount of p24 in the culture infected with M. smegmatis was higher than in cultures infected with other mycobacteria. The amounts of p24 release in cultures infected with M. avium and M. bovis BCG were intermediate. M. tuberculosis slightly stimulated HIV replication. The amount of TNF-alpha produced by U1 cells was correlated with the amount of p24 antigen release. The IL-1 beta and IL-6 levels in the supernatant from cultures infected with all species were the same. The antibody to TNF-alpha inhibited p24 release induced by mycobacterial infections. The anti-IL-1 beta and anti-IL-6 antibodies, however, scarcely influenced stimulation of HIV replication by mycobacterial infection. These data suggested that activation of HIV replication by mycobacteria mainly occurred by secondary release of cytokine TNF-alpha.

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  • Protective responses against experimental Mycobacterium leprae infection in mice induced by recombinant Bacillus Calmette-Guerin over-producing three putative protective antigen candidates Reviewed

    N Ohara, M Matsuoka, H Nomaguchi, M Naito, T Yamada

    VACCINE   19 ( 15-16 )   1906 - 1910   2001.2

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    The components of Ag85 (Ag85A, Ag85B, and Ag85C) are putative protective antigen candidates against mycobacterial infection. A recombinant Mycobacterium boris Bacillus Calmette-Guerin (rBCG) over-producing Ag85A, Ag85B, and MPB51 (rBCG/BA51) was constructed. rBCG/BA51 could secrete these antigens at levels more than five times higher than parental BCG. Immunization of C57BL/6 and BALB/c mice with this rBCG reduced the multiplication of Mycobacterium leprae in the foot pads of both strains of mice. The inhibition by rBCG/BA51 was more evident than that by parental BCG. (C) 2001 Elsevier Science Ltd. All rights reserved.

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  • Identification and characterization of the ribosome-associated protein, HrpA, of Bacillus Calmette-Guerin Reviewed

    Y Tabira, N Ohara, T Yamada

    MICROBIAL PATHOGENESIS   29 ( 4 )   213 - 222   2000.10

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    HrpA was found as a ribosome-associated protein which appeared in heat-stressed Mycobacterium bovis Bacillus Calmette-Guerin. Here, we have studied the function of HrpA in vitro. HrpA is a heat shock protein belonging to a small heat shock protein family. The putative molecular mass was 17784.86 kDa. Recombinant HrpA formed large complexes of nonamer or dodecamer. HrpA prevented the aggregation of enzymes under heat shock conditions, and it formed stable complexes with partially denatured enzymes. HrpA was induced temporarily by oxygen repletion after anaerobic condition. (C) 2000 Academic Press.

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  • Fibronectin-binding proteins secreted by Mycobacterium avium Reviewed

    H Kitaura, N Ohara, M Naito, K Kobayashi, T Yamada

    APMIS   108 ( 9 )   558 - 564   2000.9

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    Mycobacterium avium is an intracellular pathogen and a major opportunistic infectious agent observed in patients with acquired immune deficiency syndrome (AIDS). Fibronectin is an extracellular matrix protein and is a virulence factor for several extracellular pathogenic bacteria binding to mucosal surfaces. We investigated the fibronectin (FN)-binding proteins in the culture filtrate of nl. avium by two-dimensional electrophoresis (2DE). Proteins in Sauton medium of nl. nl avium after 3 weeks were separated by 2DE. The proteins were blotted onto polyvinylidene difluoride membrane and incubated with FN. FN-binding proteins were detected by Western blotting using anti-FN antibody FN bound to five spots (33 kDa, 32 kDa, 31 kDa, 30 kDa and 25 kDa). N-terminal amino acids of these were determined. The 33 kDa spot corresponded to antigen 85 (Ag 85) C. The 31 and 31 kDa spots were either Ag 85 A or Ag 85 B. The 30 kDa spot corresponded to Ag 85 B of nl. avium. The 25 kDa spot corresponded to MPA51 (M. avium MPB51). Thus, FN bound exclusively to the Ag 85 complex and MPA51.

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  • Inhibition of multiplication of Mycobacterium leprae in mouse foot pads by recombinant Bacillus Catmette-Guerin (BCG) Reviewed

    N Ohara, M Matsuoka, H Nomaguchi, M Naito, T Yamada

    VACCINE   18 ( 14 )   1294 - 1297   2000.1

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    Immunization of mice with recombinant Mycobacterium bovis Bacillus Calmette-Guerin (rBCG) which over-produces a putative protective antigen candidate, the A component of antigen 85 complex (Ag85A), reduced the multiplication of Mycobacterium leprae in the foot pads of mice. The inhibition by this rBCG (rBCG/85A) was more evident than that with parental BCG. Repeated rBCG/85A immunization significantly could reduce M. leplae multiplication in mice. This is first report of rBCG to control mycobacterial infection in animal model. Therefore, rBCG technique may be useful for the development of a more effective mycobacteria vaccine. (C) 2000 Elsevier Science Ltd. All rights reserved.

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  • The antigen 85 complex vaccine against experimental Mycobacterium leprae infection in mice Reviewed

    M Naito, M Matsuoka, N Ohara, H Nomaguchi, T Yamada

    VACCINE   18 ( 9-10 )   795 - 798   1999.12

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    The proteins in culture filtrate derived from Bacillus Calmette-Guerin (BCG) were examined for protection against infection by Mycobacterium leprae. Immunization with the major secreted proteins, antigen 85 complex (Ag 85) A: B and C, induced effective protective immunity against multiplication of M. leprae in the foot pads of mice. The most effective protection was observed when mice were immunized with Ag 85A. A single immunization with Ag 85 could induce antigen-specific interferon gamma (IFN gamma) synthesis and more effective protection than live BCG vaccine. This study demonstrates that Ag 85 is an important immunoprotective molecule against leprosy infection. (C) 1999 Elsevier Science Ltd. All rights reserved.

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  • Species-specific B-cell epitope on the C-terminal region of the alpha antigen from Mycobacterium intracellulare in mice Reviewed

    H Kitaura, N Ohara, M Naito, K Kobayashi, T Yamada

    VETERINARY MICROBIOLOGY   65 ( 1 )   9 - 19   1999.2

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    The alpha antigen, which is an immunodominant antigen, is a 30 kDa protein secreted by mycobacterial species. The C-terminal regions of alpha antigens are quite divergent. We investigated the question of whether the C-tenninal regions of Mycobacterium avium alpha antigen (A-alpha), M. intracellulare alpha antigen (I-alpha) and M. bovis BCG alpha antigen (B-alpha) contained species-specific B-cell epitopes. We investigated the reactions of these peptides with anti-A-alpha, anti-I-alpha and anti-B-alpha sera prepared from BALB/c in a Western blot assay and ELISA. The C-terminal regions of I-alpha reacted exclusively with anti-I-alpha serum. The results of the inhibition assay of antibodies binding to I-alpha by peptides of C-A-alpha, C-I-alpha, and C-B-alpha are that only C-I-alpha inhibited the binding of antibodies to C-I-alpha. We found that the C-terminal region was B-cell epitope-specific to I-alpha in BALB/c mice. (C) 1999 Elsevier Science B.V. All rights reserved.

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  • Host immune responses to ribosome, ribosomal proteins, and RNA from Mycobacterium bovis bacille de Calmette-Guerin Reviewed

    C Miyazaki, N Ohara, H Yukitake, M Kinomoto, K Matsushita, S Matsumoto, A Mizuno, T Yamada

    VACCINE   17 ( 3 )   245 - 251   1999.1

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    The ribosomes from BCG strongly induced delayed type hypersensitivity (DTH) skin reactions in guinea pigs immunized with live BCG or heat killed Mycobacterium tuberculosis H37Rv, and also induced lymphocyte proliferative response in mice immunized with ribosomes. In contrast, neither ribosomal proteins nor RNA alone induced both DTH skin reactions and lymphocyte proliferative responses. Particle form consisted of ribosomal proteins and RNAs might be absolutely required for the activation of immune responses. (C) 1998 Elsevier Science Ltd. All rights reserved.

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  • Serological analysis of C-terminal region of alpha antigen from Mycobacterium avium-intracellulare complex and Mycobacterium tuberculosis Reviewed

    H Kitaura, N Ohara, M Naito, K Kobayashi, T Yamada

    APMIS   106 ( 9 )   893 - 900   1998.9

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    The alpha antigen, which is a 30 kDa protein secreted by mycobacterial species, is an immunodominant antigen. The C-terminal regions of alpha antigens are highly divergent, though there are regions where the amino acid sequence of a antigen is conserved. We investigated whether the C-terminal regions of the Mycobacterium avium alpha antigen, M. intracellular alpha antigen and M. tuberculosis alpha antigen contain sequence-specific B-cell epitopes. The C-terminal regions of M. avium alpha antigen and M. intracellulare alpha antigen reacted to anti-M. avium alpha antigen but not to anti-M. tuberculosis alpha antigen derived from rabbits. Thus, M. avium and M. intracellulare have an antigenic determinant in common with rabbit. The C-terminal region of M. tuberculosis alpha antigen did not react to anti-M. avium alpha antigen or anti-M. tuberculosis alpha antigen. An enzyme-linked immunosorbent assay revealed that only the C-terminal region of M. avium alpha antigen reacted to the sera of two of six patients with M. avium-intracellulare (MAC) but not to the sera of patients with M. tuberculosis. In contrast, the C-terminal regions of M. intracellulare alpha antigen and M. tuberculosis a antigen were not recognized by the sera from patients with MAC or M. tuberculosis. This region of M. avium alpha antigen can produce a sequence-specific B-cell epitope in humans.

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  • Immunological characterization of alpha antigen of Mycobacterium kansasii: B-cell epitope mapping Reviewed

    M Naito, N Ohara, S Matsumoto, T Yamada

    SCANDINAVIAN JOURNAL OF IMMUNOLOGY   48 ( 1 )   73 - 78   1998.7

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    Mapping of B-cell epitopes on alpha antigen of Mycobacterium kansasii (K-alpha) was carried out by using recombinant truncated K-alpha fusion peptides. We observed that two immunodominant B-cell epitopes (amino acids 222-268 and 267-306) and one minor epitope (amino acid 249-286) were located in the C-terminal region of K-alpha. The other three minor B-cell epitopes were mapped in N-terminal (amino acids 80-98 and 99-166) and central (amino acid 174-203) regions of K-alpha. All defined epitopes were common to Mycobacterium tuberculosis and M. kansasii. Besides these common epitopes, a region in K-alpha (amino acid 290-319) revealed different reactivities between antibodies against K-alpha and alpha antigen of M. tuberculosis. These findings may provide a basis for development of serodiagnosis that can distinguish between M. kansasii and M. tuberculosis infections.

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  • The 16-kDa alpha-crystallin-like protein of Mycobacterium bovis BCG is produced under conditions of oxygen deficiency and is associated with ribosomes Reviewed

    Y Tabira, N Ohara, N Ohara, H Kitaura, S Matsumoto, M Naito, T Yamada

    RESEARCH IN MICROBIOLOGY   149 ( 4 )   255 - 264   1998.4

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    A 16-kDa protein, identical to the a-crystallin-like stress protein, was induced under O-2-deficient culture conditions and bound principally to the 30S ribosomal subunits of Mycobacterium bovis BCG substrain Tokyo (BCG). The 16-kDa protein was shown to be tightly associated with the ribosome.

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  • Differentiation of clinical isolates of Actinobacillus actinomycetemcomitans using an insertion sequence, ISAa1. Reviewed International journal

    H Hayashida, H Hotokezaka, N Ohara, T Koseki, T Nishihara, O Takagi, T Yamada

    Oral microbiology and immunology   13 ( 2 )   120 - 3   1998.4

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    We previously identified an IS200-like sequence (ISAa1) in the genome of Actinobacillus actinomycetemcomitans FDC Y4. One or more hybridizing bands to the ISAa1 probe were detected in each of several reference strains, representing three of the serotypes (a through c) of A. actinomycetemcomitans. In this study, we examined whether a restriction fragment-length polymorphism (RFLP) with ISAa1 as a probe could differentiate clinical isolates. One or more hybridizing bands were detected in each of the 27 strains examined, which could be divided into seven groups according to restriction fragment-length polymorphism pattern. Several strains were observed with identical restriction fragment-length polymorphism types but with different serotypes. Conversely, strains were also observed with differing restriction fragment-length polymorphism types and identical serotypes.

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  • The novel fibronectin-binding motif and key residues of mycobacteria Reviewed

    M Naito, N Ohara, S Matsumoto, T Yamada

    JOURNAL OF BIOLOGICAL CHEMISTRY   273 ( 5 )   2905 - 2909   1998.1

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    The binding motifs of the immunodominant antigen (Ag) alpha-Ag (Ag 85 complex B) of Mycobacterium kansasii for human fibronectin were examined using digested fragments. We defined two fibronectin-binding epitopes on 27 amino acids from 84 to 110 and on 20 amino acids from 211 to 230, The epitopes were almost conserved in the closely related Ag 85 complex of other mycobacteria species. Inhibition of fibronectin binding to intact alpha-Ag molecules was observed with peptide-(84-110), but not with peptide-(211-230). Peptide (84-110) could also inhibit fibronectin binding to all components of the Ag 85 complex of Bacillus Calmette-Guerin (Ag 85A, Ag 85B, and Ag 85C). Further study with synthetic peptides defined 11 residues from 98 to 108 as the minimum motif. Six residues ((98)FEWYYQ(103)) were critical for interacting with fibronectin. The motif revealed no homology to other known prokaryotic and eukaryotic fibronectin-binding proteins. The defined motif of alpha-Ag is novel and unique for mycobacteria.

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  • The ribosomes contents of mycobacteria Reviewed

    T Mineda, N Ohara, H Yukitake, T Yamada

    MICROBIOLOGICA   21 ( 1 )   1 - 7   1998.1

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    Mycobacterium bovis BCG (BCG) contains a low number of ribosomes compared to rapidly growing Escherichia coli.

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  • PCR amplification of 16S ribosomal RNA gene for detection of bacterial infection in root canal Reviewed

    Dentistry in Japan   34   21 - 23   1998

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  • Shotgun cloning and characterization of the thymidylate synthase-encoding gene from Mycobacterium bovis BCG Reviewed

    S Matsumoto, H Yukitake, N Ohara, T Dairi, H Kanbara, T Yamada

    MICROBIOLOGY AND IMMUNOLOGY   42 ( 1 )   15 - 21   1998

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    The shotgun cloning of a Mycobacterium bovis BCG (BCG) genome into pBluescript SK (+) successfully yielded a 0.9 kbp fragment, confirming the ability of Escherichia coli thyA mutant MH2702 to grow in a thymine-depleted medium. This DNA fragment contained a gene homologous to the thymidylate synthase (TS)-encoding genes (thyA) of other organisms. An inverted repeat sequence and open reading frame (ORF) were observed at the upstream region of the thyA. A computer analysis revealed that the protein encoded by this ORF possessed a structure unique for a DNA binding protein.

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  • Characterization of the transcriptional initiation regions of genes for the major secreted protein antigens 85C and MPB51 of Mycobacterium bovis BCG Reviewed

    N Ohara, T Nishiyama, N Ohara-Wada, S Matsumoto, T Matsuo, T Yamada

    MICROBIAL PATHOGENESIS   23 ( 5 )   303 - 310   1997.11

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    The component of mycobacterial 85 complex (85A, 85B, and 85C) and MPB51 are very important from immunological, biochemical, and antimycobacterial points of view. In this study, the transcriptional properties of genes encoding three components of 85 complex and MPB51 from BCG were analysed. The authors' analyses revealed that genes for 85A and MPB51 were transcribed as a single unit despite the one operon-like structure and these four genes were probably under a different regulatory control. These findings may help to understand the immunological and physiological roles of these antigens. (C) 1997 Academic Press Limited.

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  • HrpA, a new ribosome-associated protein which appears in heat-stressed Mycobacterium bovis Bacillus Calmette-Guerin Reviewed

    N Ohara, N Ohara, M Naito, C Miyazaki, S Matsumoto, Y Tabira, T Yamada

    JOURNAL OF BACTERIOLOGY   179 ( 20 )   6495 - 6498   1997.10

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    A novel 18-kDa heat shock protein, HrpA, has been identified from Mycobacterium bovis BCG. HrpA was rapidly synthesized in membrane and ribosome fractions but not in the cytoplasmic fraction under heat shock stress. HrpA bound tightly to 70S ribosomes, mainly in 30S subunits. HrpA might be involved in the initiation step of translation at high temperature.

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  • Analysis of the genes encoding the antigen 85 complex and MPT51 from Mycobacterium avium Reviewed

    N Ohara, N OharaWada, H Kitaura, T Nishiyama, S Matsumoto, T Yamada

    INFECTION AND IMMUNITY   65 ( 9 )   3680 - 3685   1997.9

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    The components of the fibronectin-binding antigen 85 complex (85A, 85B, and 85C) and the related protein MPB/MPT51 are major secreted proteins in Mycobacterium tuberculosis and Mycobacterium bovis BCT;, The fbpA, fbpC, and mpt51 genes encoding 85A, 85C, and MPT51, respectively, were isolated from Mycobacterium avium and sequenced in this study, The structures of these genes, and that of the fbpB gene encoding the 85B protein, were conserved in these three species. The secreted amounts of 85A, 85B, 85C, and MPB/MPT51 were compared for M. tuberculosis, BCG, and M, avium, These four proteins were found in large amounts in the culture filtrates from M. tuberculosis and BCG, In contrast, in the culture filtrate from M. avium, 85B and MPT51 were abundant whereas 85A and 85C were hardly found, in spite of the presence of the encoding genes, The difference in the secretion amounts might be regulated at the transcription level, These facts might reflect host immunopathogenesis, the protective immunities against infections, and the drug susceptibilities of these organisms.

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  • Transcriptional analysis of the groESL operon from Porphyromonas gingivalis. Reviewed International journal

    H Hotokezaka, N Ohara, H Hayashida, S Matsumoto, T Matsuo, M Naito, K Kobayashi, T Yamada

    Oral microbiology and immunology   12 ( 4 )   236 - 9   1997.8

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    Transcriptional analysis of the groESL operon from Porphyromonas gingivalis, one of the obligative anaerobic oral microorganisms implicated in adult periodontitis, was performed. P. gingivalis 381 cultured at 37 degrees C was shifted to 42 degrees C, 45 degrees C or 48 degrees C for 10 mins. Northern hybridization analysis revealed that a band with 2.1-kb (kilo base pair) was observed, and the transcripts increased greatly by heat shock. Primer extension and S1 mapping detected four different 5'-ending sites of the mRNAs at the upstream region of the groES. Three sites out of the four were heat-inducible. There were inverted repeats and a Escherichia coli sigma 32-recognizing consensus sequence in the promoter region of the groESL, which may be relevant to the regulation of transcription of groESL operon in P. gingivalis. Both a heat shock promoter and inverted repeats may be relevant to the transcriptional regulation of the groESL operon in P. gingivalis.

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  • Inhibition of multiplication of Mycobacterium leprae in mouse foot pads by immunization with ribosomal fraction and culture filtrate from Mycobacterium bovis BCG Reviewed

    M Matsuoka, H Nomaguchi, H Yukitake, N Ohara, S Matsumoto, K Mise, T Yamada

    VACCINE   15 ( 11 )   1214 - 1217   1997.8

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    Immunization of mice with the ribosomal fraction from ruptured Mycobacterium bovis Bacillus Calmette-Guerin (BCG) and the culture filtrate reduced remarkably the multiplication of Mycobacterium leprae in the foot pads of mice, This is the first reported case of the protective activity against M, leprae multiplication in mice of the BCG ribosomal fraction and culture filtrate. The inhibition was more evident with the culture filtrate than with the ribosomal fraction. When the ribosomal proteins separated from ribosomal RNA were injected into mice, only slight inhibition was observed Ribosomal RNA alone did not inhibit at all, in contrast to the conclusion reported by Youmans and Youmans. (C) 1997 Elsevier Science Ltd.

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  • Species-specific B-cell epitope on the C-terminal region of the alpha antigen from Mycobacterium scrofulaceum Reviewed

    M Takano, N Ohara, M Naito, S Matsumoto, T Matsuo, R Shirai, A Mizuno, T Yamada

    MICROBIAL PATHOGENESIS   23 ( 2 )   95 - 100   1997.8

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    In the amino acid (AA) sequences of alpha antigen from mycobacteria, C-terminal regions were variable among a variety of mycobacterial species though the N-terminal regions were relatively conserved. These regions may possess some species-specific antigenic determinants of the alpha antigen from Mycobacterium scrofulaceum (S-alpha). AAs288-300 of S-alpha fused to beta-galactosidase was reactive with the antisera raised against S-alpha. The same fused peptide did not react with the antisera raised against the alpha antigen from Mycobacterium avium (A-alpha) and Mycobacterium bovis BCG (B-alpha). B-cell epitope mapping then was performed focusing on the C-terminal region of S-alpha using the synthetic peptides. Their reactivities with antisera raised against the alpha antigens of three different mycobacterial species were assessed by ELISA. AAs279-286 were a cross-reactive common immunodominant region among three mycobacterial species. This region may be one of the crossreactive common epitopes in mycobacterial species. And AAs291-300 were reactive only with the antisera raised against S-alpha. This region may possess a species-specific epitope. (C) 1997 Academic Press Limited.

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  • Cloning and sequencing of part of the S10 operon from Actinobacillus actinomycetemcomitans FDC Y4. Reviewed International journal

    H Hayashida, H Hotokezaka, N Ohara, M Kimura, O Takagi, T Yamada

    Oral microbiology and immunology   12 ( 3 )   174 - 7   1997.6

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    We have cloned and sequenced the 5.2 kb EcoRI fragment that contained part of the S10 operon from Actinobacillus actinomycetemcomitans FDC Y4. The order of the ribosomal protein genes was identical to that of the S10 operon of Haemophilus influenzae and Escherichia coli. The deduced amino acid sequences of ribosomal proteins in this operon displayed significant homologies (65.3%-100%) to those of H. influenzae, E. coli, Yersinia enterocolitica and Yersinia pseudotuberculosis. Phylogenetic trees obtained for these ribosomal proteins were similar to that obtained for 16S rRNA.

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  • Molecular analysis of a new insertion sequence from Actinobacillus (Haemophilus) actinomycetemcomitans FDC Y4. Reviewed International journal

    H Hayashida, H Hotokezaka, N Ohara, M Kimura, O Takagi, T Yamada

    Microbiology (Reading, England)   142 ( Pt 9)   2449 - 52   1996.9

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    We have found a new insertion sequence (IS), designated ISAa1, downstream of the S10 operon in Actinobacillus (Haemophilus) actinomycetemcomitans FDC Y4. ISAa1, the first IS element characterized in this organism, is 705 bp long and lacks terminal inverted repeats. This element displayed significant homology with IS200. Hybridization patterns of genomic DNA of seven A. actinomycetemcomitans strains with an internal ISAa1 probe varied depending on the serotypes, suggesting that ISAa1 might be a useful tool for epidemiological studies.

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  • Cloning and sequencing of an MPB70 homologue corresponding to MPB83 from Mycobacterium bovis BCG Reviewed

    T Matsuo, H Matsuo, N Ohara, S Matsumoto, H Kitaura, A Mizuno, T Yamada

    SCANDINAVIAN JOURNAL OF IMMUNOLOGY   43 ( 5 )   483 - 489   1996.5

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    MPB70 is secreted in high concentrations by Mycobacterium bovis BCG substrain Tokyo (BCG Tokyo), but little by substrains Pasteur (BCG Pasteur) and M. tuberculosis. The gene encoding a MPB70 homologue secreted by BCG Tokyo was found at the upstream region of the gene encoding MPB70, with approximately 2.3 kilobase pairs (kbp) spacing: the same gene was also found in BCG Pasteur. This gene was cloned and sequenced from BCG Tokyo. The DNA sequence which contained a 663 base pair (bp) open reading frame beginning at position 1 and ending with a TAA codon at position 661 was found. Its theoretical molecular mass was calculated to be 22.068 kDa. This gene was highly homologous to the coding region of mpb70 and the deduced amino acid sequence was very similar to MPB83 reported by Harboe et al. It was speculated that the gene the authors characterized probably corresponded to the mpb83 gene.

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  • Long-lasting immune response induced by recombinant Bacillus Calmette-Guerin (BCG) secretion system Reviewed

    N Wada, N Ohara, M Kameoka, Y Nishino, S Matsumoto, T Nishiyama, M Naito, H Yukitake, Y Okada, K Ikuta, T Yamada

    SCANDINAVIAN JOURNAL OF IMMUNOLOGY   43 ( 2 )   202 - 209   1996.2

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    The recombinant bacillus Calmette-Guerin (rBCG) secretion system utilizing an extracellular alpha antigen of Mycobacterium kansasii (alpha-K) was characterized biochemically and immunologically. The human immunodeficiency virus type1 (HIV-1)p17(gag) B cell epitope fused to alpha-K was secreted in extremely large amounts. At least three mice out of seven inoculated with rBCG generated high titres of antibody to the epitope. The long-lasting antibody production persisted more than 14 months.

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  • Stable expression and secretion of the B-cell epitope of rodent malaria from Mycobacterium bovis BCG and induction of long-lasting humoral response in mouse Reviewed

    S Matsumoto, T Yanagi, N Ohara, N Wada, H Kanbara, T Yamada

    VACCINE   14 ( 1 )   54 - 60   1996.1

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    The live bacterial vaccine Mycobacterium bovis BCG (BCG) is a vehicle worth noticing for various protective antigens. The gene encoding the B-cell epitope of the oligopeptide repeating in the circumsporozoite protein (C.S. protein) of the rodent malaria parasite, Plasmodium yoelii, was inserted into the plasmid vector under the control of an expression cassette carrying the promoter and signal sequence of the a antigen derived from Mycobacterium kansasii (k-a). The B-cell epitope was successfully expressed and secreted from BCG as a fusion protein with k-a. This recombinant BCG was administered subcutaneously into BALB/c mice and the antibody production was measured by the enzyme-linked immunosorbent assay (ELISA). Long lasting humoral response was found in one of seven mice.

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  • BCG vaccination to Mycobacterium leprae infection in mice Reviewed

    H. Nomaguchi, Y. Fukutomi, S. Nagai, Y. Yogi, H. Okamura, N. Ohara, Matsuoka M, K. Nagata, T. Yamada

    Japanese Journal of Leprosy   65 ( 2 )   106 - 112   1996

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    BCG vaccine (Tokyo strain) was given in BALB/cA mice intradermally 1 or 3 months before Mycobacterium leprae (M. leprae) challenge as modified Shepard's method. The vaccine dosage was 107-8 or 106. The BCG gave good protection in both dosages and both challenges against M. leprae infection. Lymphocytes proliferations of BCG-vaccinated splenocyte cultures in response to M. leprae lysate or BCG components (hsp65, 38kD, 30kD or 12kD protein) were tested, and potent proliferative responses were seen in the cultures with M. leprae lysate and hsp65. Furthermore, γ-IFN productions were positive in the cultures with M. leprae lysate or hsp65, but negative with other antigens. The production of γ-IFN with hsp65 was never inhibited with polymyxin B, but inhibited with IL 10. These results show that BCG (Tokyo strain) is a useful vaccine for M. leprae infection in mice, and one of the compounds of BCG, hsp65, may be a effective antigen component for protection of M. leprae infection inducing Th1 type cytokine.

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  • AN INTERNAL CONTROL FOR THE RAPID DETECTION OF MYCOBACTERIA BY AMPLIFICATION OF A SEGMENT OF THE GENE ENCODING ALPHA-ANTIGEN Reviewed

    H KITAURA, N OHARA, T MATSUO, T YAMADA

    MICROBIOLOGICA   18 ( 4 )   429 - 433   1995.10

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    The internal control of DNA for the rapid detection of mycobacteria by PCR is described. The 1100bp fragment for internal control was produced from Streptomyces lividans DNA with the primers used for the rapid detection of mycobacteria by PCR. The amplified reaction consequently produced two products with 782bp for mycobacteria and 1100bp for the internal control extracted from all mycobacterial DNAs containing internal control so far examined. The 1100bp amplified fragment proved to be useful as an internal control with the same primer-binding sequence for the detection of mycobacteria.

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  • DIFFERENTIAL TRANSCRIPTION OF THE MPB70 GENES IN 2 MAJOR GROUPS OF MYCOBACTERIUM-BOVIS BCG SUBSTRAINS Reviewed

    T MATSUO, S MATSUMOTO, N OHARA, H KITAURA, A MIZUNO, T YAMADA

    MICROBIOLOGY-UK   141   1601 - 1607   1995.7

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    Substrains of Mycobacterium bovis BCG (BCG) have been divided into two major groups, high and low producers, on the basis of the amount of secretion of the MPB70 protein. The antigen is produced in high concentration by BCG Tokyo, Moreau, Russia and Sweden (high-producer substrains), whereas in BCG Pasteur, Copenhagen and Tice (low-producer substrains) it is detected at 1% (w/w) or less of the concentration of BCG Tokyo. To investigate why this protein is secreted differently, the MPB70 genes of BCG Tokyo and Pasteur were cloned, sequenced and compared. The MPB70 genes in two substrains showed exactly the same sequence. Even the upstream and downstream regions of the MPB70 gene were identical. MPB70 gene expression was assessed by means of Northern hybridization analysis and reverse transcriptase polymerase chain reaction. The mRNA was clearly detected in BCG Tokyo, but at a very low level in BCG Pasteur. On the basis of these results, the difference in the secretion of the MPB70 protein between BCG Tokyo and Pasteur was attributed to differential transcription efficiencies.

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  • Characterization of the gene encoding the MPB51, one of the major secreted protein antigens of Mycobacterium bovis BCG, and identification of the secreted protein closely related to the fibronectin binding 85 complex. Reviewed International journal

    N Ohara, H Kitaura, H Hotokezaka, T Nishiyama, N Wada, S Matsumoto, T Matsuo, M Naito, T Yamada

    Scandinavian journal of immunology   41 ( 5 )   433 - 42   1995.5

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    The secreted protein MPB51 is one of the major proteins in the culture filtrate of Mycobacterium bovis BCG (BCG) and is a protein immunologically cross-reacting with the fibronectin binding 85 complex secreted by this bacterium. The gene encoding MPB51 (mpb51) was cloned, sequenced, and expressed in Escherichia coli. The mpb51 gene was mapped downstream of the gene for 85A component with 179 bp spaces. The mpb51 gene encoded 299 amino acids, including 33 amino acids for the signal peptide, followed by 266 amino acids for the mature protein with a molecular mass of 27807.37 Da. This is the first complete sequence of MPB51. MPB51 showed 37-43% homology to the components of 85 complex. Two-dimensional electrophoresis of culture fluids of BCG and Western blotting indicated the existence of the other novel protein(s) which strongly cross-reacted with the alpha antigen (85B) and MPB51.

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  • CLONING AND NUCLEOTIDE-SEQUENCE OF THE GENE-CLUSTER ENCODING RIBOSOMAL-PROTEINS S12 AND S7 FROM MYCOBACTERIUM-BOVIS BCG Reviewed

    S IWANAGA, N OHARA, T KARIU, M KIMURA, N YAMASAKI, T YAMADA

    BIOCHEMISTRY AND MOLECULAR BIOLOGY INTERNATIONAL   36 ( 1 )   209 - 218   1995.5

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    A pair of oligonucleotide primers, based on the experimentally determined amino terminal sequence of Mycobacterium bovis BCG ribosomal protein S12 (MboS12) and a highly conserved sequence found in all mycobacterial ribosomal S12 proteins, was used for polymerase chain reaction (PCR) with M. bovis genomic DNA as template. The nucleotide sequence of the 338 bp fragment thus produced confirmed its origin in MboS12 gene. A 5.0 kb EcoRI fragment of M. bovis DNA hybridizing to this fragment was cloned. Its sequencing analysis revealed the presence of two open reading frames in the same strand. Their amino acid sequences deduced from DNA sequence showed high homology with Escherichia coli ribosomal proteins S12 and S7. However, the intercistronic region between S12 and S7 genes, which plays an important role for autoregulation for the str operon in E. coli, is completely absent in M. bovis.

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  • Cloning and sequencing of a unique antigen MPT70 from Mycobacterium tuberculosis H37Rv and expression in BCG using E. coli-mycobacteria shuttle vector. Reviewed International journal

    S Matsumoto, T Matsuo, N Ohara, H Hotokezaka, M Naito, J Minami, T Yamada

    Scandinavian journal of immunology   41 ( 3 )   281 - 7   1995.3

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    MPB70 is known to be an immunogenic mycobacterial protein secreted in large amounts from Mycobacterium bovis BCG (BCG) Tokyo. The analogous gene for MPT70 was cloned from Mycobacterium tuberculosis H37Rv which produces this protein in only a small amount. The gene encoding 193 amino acid residues including 30 amino acids for the signal peptide, the promoter-like sequence, and the ribosome-binding site, was completely identical to that of BCG Tokyo. Computer analysis revealed that the carboxy-terminal half of MPT70 was homologous to amino acid sequences of fasciclin I, osteoblast-specific factor 2 (OSF-2), and human transforming growth factor-beta induced gene product (beta IG-H3). Escherichia coli (E. coli) -mycobacteria shuttle vectors containing mpt70 or mpb70 genes 0.7kbp upstream of the 5' end of them were able to be expressed in BCG Pasteur which is a MPB70 low-producer, but the extent of the expression was not that of a high-producer.

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  • Biochemical and Immunological Characterization of Ribosomal Fraction and Culture Filtrate from Mycobacterium Reviewed

    Yamada T, Ohara N, Wada N, Matsumoto S, Yukitake E, Matsuo T, Kitaura H, Takano M, Nishiyama T, Nomaguchi H, Matsuo M

    Actinomycetol   9   228 - 235   1995

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  • 1. The role of some cellular components of bacterial parasites in determining the incidence of tuberculosis: Studies on mycobacterial antigens, with special reference to mycobacterial immunoreactive ribosomal and secreted proteins Reviewed

    T. Yamada, N. Ohara, S. Matsumoto, T. Matsuo, H. Kitaura, H. Yukitake, N. Wada, T. Nishiyama, M. Naito, M. Kinomoto, M. Kimura

    Kekkaku   70 ( 11 )   639 - 644   1995

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    Tuberculosis remains as major disease, affecting more than 20 million people. The elimination of the disease with vaccination, rapid diagnosis, and efficient therapy is an important objective of our study. To realize the objective, the characterization of antigens is essential. We have chosen two kinds of antigens for our study, the ribosomal antigens and an antigenic proteins secreted by mycobacteria. The biochemical and immunological characterization of ribosomal fraction was carried out. Ribosomal proteins were purified and assessed for DTH reaction. The N-terminal amino acids sequences were determined. Total structures of S19, S7 and S12 in 30S and L7/L12 in 50S subunits were elucidated. L7/L12 had 66% homology with analogue from S. griseus which showed GTPase activity in protein synthesis. This protein was secreted in culture medium and induced strong DTH. Secreted antigenic proteins are of great interest for us. Secreted antigens may he recognized rapidly by immune system and therefore may induce rapid and high level immune response. It is also expected that it may contain protective antigens, since live BCG protect disease more efficiently than heat killed BCG. We have determined and published the total structure of four proteins (MPB64, MPB70, MPB57, anti α antigen). We attempted to utilize this antigen for the diagnosis and the design of vaccine. The structures of a antigens from M. avium, M. intracellulare, M. scrofulaceum, M. kansasii and BCG were determined and its potential for application to diagnosis was presented. Using the operon of M. kansasii, α antigen anti V3 reagion of HIV-1 were expressed by recombinant BCG which induced CTL in mice.

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  • Biochemical and immunological characterization of ribosomal fraction and culture filtrate from BCG Reviewed

    Yamada T, Ohara N, Matsumoto S, Matsuo T, Kitaura H, Takano M, Nishiyama T, Yukitake E, Miyazaki R, Nomaguchi H, Mastuoka M

    Proceedings to 30th Joint Reserch Conference on Leprosy and Tuberculosis   30   246 - 249   1995

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  • Cloning and sequencing of the groESL homologue from Porphyromonas gingivalis. Reviewed International journal

    H Hotokezaka, H Hayashida, N Ohara, H Nomaguchi, K Kobayashi, T Yamada

    Biochimica et biophysica acta   1219 ( 1 )   175 - 8   1994.9

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    The homologue of groESL from Porphyromonas gingivalis was cloned and sequenced. Nucleotide sequencing suggested an operon containing two open reading frames (ORFs) homologous to groESL operon of Escherichia coli. The upstream ORF consisted of 267 bp corresponding to 89 amino acid residues. The downstream ORF consisted of 1635 bp corresponding to 545 amino acid residues.

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  • Cloning and sequencing of the groESL homologue from Porphyromonas gingivalis

    Hitoshi Hotokezaka, Hideaki Hayashida, Naoya Ohara, Hiroko Nomaguchi, Kazuhide Kobayashi, Takeshi Yamada

    BBA - Gene Structure and Expression   1219   175 - 178   1994.9

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    The homologue of groESL from Porphyromonas gingivalis was cloned and sequenced. Nucleotide sequencing suggested an operon containing two open reading frames (ORFs) homologous to groESL operon of Escherichia coli. The upstream ORF consisted of 267 bp corresponding to 89 amino acid residues. The downstream ORF consisted of 1635 bp corresponding to 545 amino acid residues. © 1994.

    DOI: 10.1016/0167-4781(94)90265-8

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  • CLONING, SEQUENCING AND EXPRESSION IN ESCHERICHIA-COLI OF THE GENE FOR ALPHA-ANTIGEN FROM MYCOBACTERIUM-SCROFULACEUM Reviewed

    M TAKANO, N OHARA, A MIZUNO, T YAMADA

    SCANDINAVIAN JOURNAL OF IMMUNOLOGY   40 ( 2 )   165 - 170   1994.8

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    The gene for the extracellular a antigen of Mycobacterium scrofulaceum (S-a) was cloned by using the a antigen gene fragments of Mycobacterium bovis BCG as probes. The complete nucleotide sequence was determined. The gene was expressed in Escherichia coli. The gene encodes 330 amino acids, including 40 amino acids for the signal peptide, followed by 290 amino acids for the mature protein. The deduced amino acid sequences were highly homologous to the a antigen of the species of other mycobacteria. Interestingly, the a antigens of MAIS complex (Mycobacterium avium-Mycobacterium intracellulare- M. scrofulaceum) were closely homologous even at the C-terminal regions which were variable among those of M. bovis BCG, Mycobacterium leprae and Mycobacterium kansasii.

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  • CYTOTOXIC T-LYMPHOCYTE RESPONSE IN MICE INDUCED BY A RECOMBINANT BCG VACCINATION WHICH PRODUCES AN EXTRACELLULAR ALPHA ANTIGEN THAT FUSED WITH THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 ENVELOPE IMMUNODOMINANT DOMAIN IN THE V3 LOOP Reviewed

    M KAMEOKA, Y NISHINO, K MATSUO, N OHARA, T KIMURA, A YAMAZAKI, T YAMADA, K IKUTA

    VACCINE   12 ( 2 )   153 - 158   1994.2

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    The host immune response of cell-mediated immunity, particularly that of cytotoxic T lymphocytes (CTLs), is a major immune defence mechanism which may provide resistance to a human immunodeficiency virus type 1 (HIV-1) spread leading to acquired immune deficiency syndrome (AIDS). To prevent the accompanying activity of HIV-1 proteins responsible for the loss of helper T-lymphocyte function, it is crucial to develop a live attenuated recombinant vaccine expressing only T- or both T- and B-cell epitopes. Here, we examined the expression of the HIV-1 Env protein V3 region (15 amino acids from Arg(315) to Lys(329)) in Mycobacterium bovis BCG as a fused form with an extracellular alpha antigen of Mycobacterium kansasii. Balb/c mice inoculated with this recombinant BCG (rBCG), rapidly induced V3 peptide-specific CTLs. Target cell lysis was restricted to the murine class I major histocompatibility complex, H-2(d). A similar CTL response was also elicited after Balb/c mice were immunized with the same rBCG even when pre-inoculated with non-recombinant BCG. Thus, the rapid induction of HIV-1-specific CTLs indicates that this vaccine may be a therapeutic approach to preventing progression to AIDS.

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  • Structure and immunological properties of major secreted antigen a and related antigens of mycobacteria. Reviewed

    Ohara N, Kitaura H, Naito M, Nishiyama T, Wada N, Matsumoto S, Matsuo T, Hotokezaka H, Hayashida H, Yamada T

    Procceedings to 4th Western Pacifuc Congress on Chemotherapy and Infectious Diseases   10 ( 3 )   628 - 629   1994

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  • Regulation of MPB70 gene expression between two major BCG substrains. Reviewed

    Matsuo T, Ohara N, Matsumoto S, Kitaura H, Naito M, Wada N, Nishiyama T, Hotokezaka H, Hayashida H, Mizuno A, Yamada T

    Procceedings to 4th Western Pacifuc Congress on Chemotherapy and Infectious Diseases   10 ( 3 )   590 - 591   1994

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  • Study of a antigen from Mycobacterium intracellulare and use of PCR for the rapid identification of Mycobacterium intracellulare. Reviewed

    Kitaura H, Ohara N, Naito M, Nishiyama T, Wada N, Matsumoto S, Matsuo T, Hotokezaka H, Hayashida H, Kobayashi K, Yamada T

    Procceedings to 4th Western Pacific Congress on Chemotherapy and Infectious Diseases   10 ( 3 )   585 - 586   1994

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    DOI: 10.1006/bbrc.1993.2417

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  • CLONING, SEQUENCING AND EXPRESSION OF THE GENE FOR ALPHA-ANTIGEN FROM MYCOBACTERIUM-INTRACELLULARE AND USE OF PCR FOR THE RAPID IDENTIFICATION OF MYCOBACTERIUM-INTRACELLULARE Reviewed

    H KITAURA, N OHARA, T MATSUO, H TASAKA, K KOBAYASHI, T YAMADA

    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS   196 ( 3 )   1466 - 1473   1993.11

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    DOI: 10.1006/bbrc.1993.2417

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  • IMMUNOLOGICAL PROPERTIES OF RIBOSOMAL-PROTEINS FROM MYCOBACTERIUM-BOVIS BCG Reviewed

    S TANTIMAVANICH, S NAGAI, H NOMAGUCHI, M KINOMOTO, N OHARA, T YAMADA

    INFECTION AND IMMUNITY   61 ( 9 )   4005 - 4007   1993.9

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    Two proteins with molecular mass 65 kDa, a heat shock protein, and an S1-like protein were found in a 30S ribosomal subunit from Mycobacterium bovis BCG. The 17-kDa protein in the 30S subunit was homologous to alpha-crystallin heat shock protein, and the 16-kDa protein in the 50S subunit was homologous to the L7/L12 protein. The latter provoked a strong delayed-type hypersensitivity reaction in the sensitized guinea pigs. The GroES-like protein (12 kDa) loosely associated with ribosomes.

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  • ISOLATION AND AMINO-ACID-SEQUENCE OF THE 30S-RIBOSOMAL PROTEIN-S19 FROM MYCOBACTERIUM-BOVIS BCG Reviewed

    N OHARA, M KIMURA, Y HIGASHI, T YAMADA

    FEBS LETTERS   331 ( 1-2 )   9 - 14   1993.9

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    The 30S ribosomal proteins from Mycobacterium bovis BCG were separated by reverse phase-high performance liquid chromatography (RP-HPLC). The isolated proteins were analyzed by SDS-PAGE, blotted on PVDF-membranes and subjected to sequence analyses using a gas-phase sequencer to correlate them to those of the well studied Escherichia coli and Bacillus stearothermophilus ribosomes. Moreover, the internal amino acid sequence of one ribosomal protein, MboS19, which is homologous to E. coli ribosomal protein S19 (EcoS19) and B. stearothermophilus ribosomal protein S19 (BstS19), was further analyzed by sequencing its internal peptides and two segments from the N- and C-termini of the protein were selected to deduce the sequence of two oligonucleotide primers which were used in a polymerase chain reaction. Using the amplified DNA fragment thus obtained as a hybridization probe, the gene encoding protein S19 was identified and cloned. Sequence analysis of the DNA fragment, together with peptide sequence analysis could determine the complete amino acid sequence of MboS19. This sequence proved to be 64% and 71% identical to those of the corresponding S19 proteins from the eubacteria E coli, and B. stearothermophilus, respectively; 33% of the residues of MboS19 were identical to those in the archaebacteral ribosomal protein HmaS19.

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  • CLONING AND SEQUENCING OF THE GENE ENCODING THE RIBOSOMAL L7/L12-LIKE PROTEIN OF MYCOBACTERIUM-BOVIS BCG Reviewed

    N OHARA, M KIMURA, N WADA, T YAMADA

    NUCLEIC ACIDS RESEARCH   21 ( 15 )   3579 - 3579   1993.7

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  • CLONING AND SEQUENCING OF THE GENE FOR ALPHA-ANTIGEN FROM MYCOBACTERIUM-AVIUM AND MAPPING OF B-CELL EPITOPES Reviewed

    N OHARA, K MATSUO, R YAMAGUCHI, A YAMAZAKI, H TASAKA, T YAMADA

    INFECTION AND IMMUNITY   61 ( 4 )   1173 - 1179   1993.4

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    The complete nucleotide sequence of alpha antigen secreted from Mycobacterium avium (A-alpha) was determined. The gene encodes 330 amino acids, including 40 amino acids for the signal peptide, followed by 290 amino acids for the mature protein with a molecular mass of 30,811 Da. This is the first sequence of A-alpha. Comparisons between A-alpha and alpha antigens of Mycobacterium leprae, Mycobacterium bovis BCG, and Mycobacterium kansasii showed highly homologous regions which suggested a conserved functional domain and two less-homologous regions. Serological analysis of recombinant A-alpha, expressed by a series of deletion constructs, indicated the possibility that A-alpha carries at least six B-cell epitopes. The three antigenic determinants were common to Mycobacterium tuberculosis, M. kansasii, and M. avium. The results also suggested the possibility that there are three species-specific epitopes.

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  • III-2. Study on recombinant BCG Reviewed

    K. Matsuo, R. Yamaguchi, A. Yamazaki, K. Terasaka, S. Nagai, H. Tasaka, C. Abe, M. Totsuka, H. Yukitake, K. Kobayashi, N. Ohara, T. Yamada

    Kekkaku   66 ( 9 )   615 - 619   1991

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  • The comparison of recovery rate and distribution of Brugia pahangi in mogolian jirds (Meriones unguiculatus) kept at different levels of ambient temperature and ultra violet irradiation Reviewed

    Fujiwara M, Ohara N, Shigeno S, Kimura E, Aoki Y

    Trop Med   27   17 - 22   1985

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Books

  • 口腔微生物学・免疫学 第5版

    大原直也( Role: Joint editor)

    医歯薬出版  2021.11 

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  • Interface Oral Health Science〈2007〉

    Kondo Y, Yoshimura M, Ohara N, Shoji M, Yukitake H, Naito M, Fujiwara T, Nakayama K( Role: Joint author)

    Springer  2008  ( ISBN:9784431766896

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    Responsible for pages:271-272   Language:English

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MISC

  • 四半世紀に及んだ腸管出血性大腸菌感染症の戦いと未来 感染症ミューズ細胞治療の挑戦(Quarter century battle against EHEC infectious disease; Muse cell therapy challenge on infectious diseases)

    藤井 潤, 出澤 真理, 尾鶴 亮, 若尾 昌平, 辻 高寛, 松葉 隆司, 黒沢 洋一, 大原 直也, 松本 壮吉, 安田 香央里, 飯野 守男

    日本細菌学雑誌   76 ( 1 )   126 - 126   2021.2

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  • Comparison of differential gene expression in mycobacterial biofilm formation vs dormancy

    西内由紀子, 大田篤, 矢野大和, 岩本朋忠, 阿戸学, 松本壮吉, 大原直也, 丸山史人

    Bacterial Adherence & Biofilm   33   2020

  • Porphyromonas gingivalisのPGN_0297の機能解析

    小野 晋太郎, 中山 真彰, 大原 直也, 高柴 正悟

    日本歯周病学会会誌   61 ( 秋季特別 )   124 - 124   2019.10

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  • 非結核性抗酸菌におけるpYT系プラスミド利用の可能性

    野崎 高儀, 中山 真彰, 小川 みどり, 吉田 志緒美, 阿戸 学, 大原 直也

    日本細菌学雑誌   74 ( 1 )   88 - 88   2019.3

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  • 急速に増殖するMycobacterium aviumの亜種のhominissuis 104株の分析(Analysis of a rapid growing Mycobacterium avium subspecies hominissuis 104 strain)

    河喜多 智美, 吉田 光範, 鈴木 仁人, 中田 登, 瀧井 猛将, 中山 真彰, 梁 明秀, 星野 仁彦, 阿戸 学, 大原 直也

    日本細菌学雑誌   74 ( 1 )   61 - 61   2019.3

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  • マイコファージによる非結核性抗酸菌症の予防法と治療法の確立

    島守 祐月, 大原 直也, 露口 一成, 大屋 賢司, 吉田 志緒美, 武田 茂樹, 永井 宏樹, 安藤 弘樹

    日本細菌学雑誌   74 ( 1 )   51 - 51   2019.3

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  • マイコファージによる非結核性抗酸菌症の予防法と治療法の確立

    島守 祐月, 大原 直也, 露口 一成, 大屋 賢司, 吉田 志緒美, 武田 茂樹, 永井 宏樹, 安藤 弘樹

    日本細菌学雑誌   74 ( 1 )   51 - 51   2019.3

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  • 非結核性抗酸菌におけるpYT系プラスミド利用の可能性

    野崎 高儀, 中山 真彰, 小川 みどり, 吉田 志緒美, 阿戸 学, 大原 直也

    日本細菌学雑誌   74 ( 1 )   88 - 88   2019.3

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  • pYTプラスミドの非結核性抗酸菌における応用の可能性の検討

    野崎 高儀, 中山 真彰, 小川 みどり, 吉田 志緒美, 阿戸 学, 大原 直也

    結核   94 ( 3 )   293 - 293   2019.3

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  • 非結核性抗酸菌のバイオフィルム形成条件における遺伝子発現解析(Detection of Differential Gene Expression in Biofilm-Forming Mycobacterium avium subsp. hominissuis)

    西内 由紀子, 大田 篤, 岩本 朋忠, 阿戸 学, 松本 壮吉, 大原 直也, 丸山 史人

    日本細菌学雑誌   74 ( 1 )   109 - 109   2019.3

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  • トリ型結核菌Mycobacterium aviumの酸性環境下における適応機構の解析

    瀧井 猛将, 小川 翔大, 大原 直也, 小川 賢二, 八木 哲也, 藤原 永年, 前田 伸司, 伊藤 佐生智, 肥田 重明, 小野崎 菊夫

    日本細菌学雑誌   74 ( 1 )   55 - 55   2019.3

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  • Porphyromonas gingivalisのジンジパインの機能発現におけるPGN_0297の役割

    小野 晋太郎, 高柴 正悟, 大原 直也

    岡山歯学会雑誌   37 ( 2 )   81 - 82   2018.12

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  • チミジル酸合成酵素遺伝子過剰発現によるBCGの増殖速度の促進とその機序の解明

    大原直也, 大原直也, 和田崇之, 阿戸学, 鈴木定彦

    結核   93 ( 4 )   290   2018.4

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  • マウスにおいてIL-21はBCG感染後に短命のエフェクターCD8+T細胞を誘導する(IL-21 induces short-lived effector CD8+ T cells after BCG infection in mice) Reviewed

    野口 直人, 中村 梨沙, 畑野 晋也, 山田 久方, Sun Xun, 大原 直也, 吉開 泰信

    日本細菌学雑誌   73 ( 1 )   132 - 132   2018.2

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  • BCGの増殖率促進に関与する遺伝子の調査(Investigation of genes involved in promoting the growth rate of BCG)

    竜門 亜矢子, 中山 真彰, 和田 崇之, 橘 理人, 阿戸 学, 中島 千絵, 鈴木 定彦, 小崎 弘貴, 大原 直也

    日本細菌学雑誌   73 ( 1 )   68 - 68   2018.2

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  • Investigation of genes involved in promoting the growth rate of BCG

    竜門亜矢子, 中山真彰, 中山真彰, 和田崇之, 橘理人, 阿戸学, 中島千絵, 鈴木定彦, 小崎弘貴, 大原直也, 大原直也

    日本細菌学雑誌(Web)   73 ( 1 )   2018

  • 抗酸菌における誘導発現系を用いたmycobacterial DNA-binding protein 1の機能解析(Conditional expression of mycobacterial DNA-binding protein 1 function in mycobacteria)

    Savitskaya Anna G, 西山 晃史, 大原 直也, 松本 壮吉

    日本細菌学雑誌   72 ( 1 )   97 - 97   2017.2

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  • BCG thyXを用いた抗酸菌のPAS耐性機序の解析

    大原直也, 大原直也, 阿戸学, 鈴木定彦, 小林和夫

    結核   91 ( 3 )   325   2016.3

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  • 次世代シーケンサーを用いたBCG Tokyo 172のシードロットおよび市販ロットにおけるヘテロ変異検出

    和田 崇之, 岩本 朋忠, 前田 伸司, 山本 太郎, 山本 三郎, 大原 直也, 中川 一路, 丸山 史人

    結核   91 ( 3 )   326 - 326   2016.3

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  • 抗酸菌のPAS耐性機序の解明 BCG thyX欠損株および過剰発現株の作製

    有村 友紀, 中山 真彰, 妹尾 昌紀, 田川 淳平, 阿戸 学, 中島 千絵, 鈴木 定彦, 中山 浩次, 小林 和夫, 大原 直也

    日本細菌学雑誌   71 ( 1 )   142 - 142   2016.2

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  • 次世代シーケンサーによるBCGワクチンのシードロットと市販ロットにおけるヘテロ変異の検出

    和田 崇之, 丸山 史人, 岩本 朋忠, 山本 太郎, 中川 一路, 山本 三郎, 大原 直也

    日本細菌学雑誌   71 ( 1 )   134 - 134   2016.2

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  • Porphyromonas gingivalis ECFシグマ因子変異株における菌体表層性状の解析

    菊池有一郎, 菊池有一郎, 国分栄仁, 国分栄仁, 柴山和子, 大原直也, 中山浩次, 石原和幸, 石原和幸

    Journal of Oral Biosciences Supplement (Web)   2016   2016

  • The deletion of glycopeptidolipid in Mycobacterium smegmatis J15cs strain affects morphology and survival in host cells

    N. Fujiwara, M. Ayata, N. Ohara, M. Ogawa, T. Naka, H. Taniguchi, S. Yamamoto, S. Maeda

    FEBS JOURNAL   282   238 - 238   2015.7

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  • 抗酸菌におけるパラアミノサリチル酸に対する新たな耐性機序の可能性

    ZHAO NA, NAKAYAMA MASAAKI, SEKIZUKA TSUYOSHI, KURODA MAKOTO, HONDA NAOKO, ATO MANABU, NAKAJIMA CHIE, SUZUKI YASUHIKO, OHARA NAOYA

    日本細菌学雑誌   70 ( 1 )   193   2015.2

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  • 結核に対する新規ワクチンIL-7産生BCGの開発(Development of a recombinant BCG expressing murine IL-7 as a novel vaccine against tuberculosis)

    畑野 晋也, 柴田 健輔, 山田 久方, 大原 直也, 田村 敏生, 吉開 泰信

    日本細菌学雑誌   70 ( 1 )   235 - 235   2015.2

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  • 結核菌の潜伏、持続感染の分子遺伝学

    大原直也

    化学療法の領域   31 ( 9 )   101 - 106   2015

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  • Porphyromonas gingivalis ECFシグマ因子変異株の性状解析

    菊池有一郎, 柴山和子, 国分栄仁, 国分栄仁, 大原直也, 中山浩次, 石原和幸, 石原和幸

    Journal of Oral Biosciences Supplement (Web)   2015   2015

  • バイオフィルム形成におけるPorphyromonas gingivalisECFシグマ因子の役割

    菊池有一郎, 菊池有一郎, 柴山和子, 国分栄仁, 国分栄仁, 大原直也, 中山浩次, 石原和幸, 石原和幸

    Journal of Oral Biosciences Supplement (Web)   2014   2014

  • 電気穿孔法に適したPorphyromonas gingivalis用新規プラスミドベクターの構築

    田川 淳平, 井上 哲圭, 大原 直也, 山城 隆

    日本矯正歯科学会大会プログラム・抄録集   72回   210 - 210   2013.10

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  • Porphyromonas gingivalis PGN_1796は薬剤感受性に関与する

    田口 裕子, 井上 哲圭, 大原 直也, 前田 博史, 高柴 正悟

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   139回   95 - 95   2013.10

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  • Porphyromonas gingivalisにおいて電気穿孔法で導入可能なプラスミドベクターの構築

    田川 淳平, 井上 哲圭, 佐藤 啓子, 内藤 真理子, 中山 真彰, 中山 浩次, 山城 隆, 大原 直也

    Journal of Oral Biosciences Supplement   2013   113 - 113   2013.9

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  • 口腔癌に対するBCG生菌療法による抗腫瘍、延命効果の検討

    村上 純, 大原 直也, 中山 真彰, 辻極 秀次, 長塚 仁, 此内 浩信, 柳 文修, 畦坪 輝寿, 浅海 淳一

    Journal of Oral Biosciences Supplement   2013   222 - 222   2013.9

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  • Porphyromonas gingivalisのPGN_1202(rpoN)は必須遺伝子である

    加野 小奈美, 井上 哲圭, 田口 裕子, 田川 淳平, 中山 真彰, 山城 隆, 大原 直也

    Journal of Oral Biosciences Supplement   2013   169 - 169   2013.9

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  • Porphyromonas gingivalisにおいて電気穿孔法で導入可能なプラスミドベクターの構築

    田川 淳平, 井上 哲圭, 佐藤 啓子, 内藤 真理子, 中山 真彰, 中山 浩次, 山城 隆, 大原 直也

    Journal of Oral Biosciences Supplement   2013   167 - 167   2013.9

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  • New insights into the virulence factors of Mycobacterium tuberculosis

    246 ( 6 )   465 - 469   2013.8

  • バイオフィルム形成におけるPorphyromonas gingivalis ECFシグマ因子の役割

    菊池有一郎, 菊池有一郎, 柴山和子, 国分栄仁, 国分栄仁, 大原直也, 中山浩次, 石原和幸

    Journal of Oral Biosciences Supplement (Web)   2013   2013

  • [Mycobacterium tuberculosis]. Reviewed

    Ohara N

    Nihon rinsho. Japanese journal of clinical medicine   70 ( 2 )   243 - 246   2012.2

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  • 国際共同基盤研究に応用する抗酸菌感染症研究の整備 結核菌の分子遺伝学

    谷口初美, 藤原永年, 大原直也, 福田和正, 小川みどり

    国際共同基盤研究に応用する抗酸菌感染症研究の整備 平成22年度 総括・分担研究報告書   35 - 38   2011

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    J-GLOBAL

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  • 抗酸菌感染症の発症・診断・治療・新世代予防技術に係わる分子機構に関する研究 結核菌潜伏感染の分子機構と治療・予防戦略

    小林和夫, 前倉亮治, 北田清悟, 松本壮吉, 藤原永年, 阿戸学, 大西和夫, 高橋宜聖, 岡部真裕子, 大原直也

    抗酸菌感染症の発症・診断・治療・新世代予防技術に係わる分子機構に関する研究 平成21年度 総括・分担研究報告書   17 - 21   2010

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    J-GLOBAL

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  • Porphyromonas gingivalis ECFシグマ因子PG0162変異株の性状解析 Reviewed

    菊池 有一郎, 大原 直也, 上田 青海, 平井 要, 柴田 幸永, 雪竹 英治, 中山 浩次, 藤村 節夫

    日本細菌学雑誌   64 ( 1 )   162 - 162   2009.2

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  • 持続性結核菌感染の病原性や発症に関わる分子機構の解明及び治療・予防の基礎研究 休眠抗酸菌と宿主応答

    小林和夫, 藤原永年, 大原直也, 阿戸学, 大西和夫, 高橋宜聖, 岡部真裕子

    持続性結核菌感染の病原性や発症に関わる分子機構の解明及び治療・予防の基礎研究 平成20年度 総括・分担研究報告書   11 - 14   2009

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    J-GLOBAL

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  • 抗酸菌感染症の発症・診断・治療・新世代予防技術に係わる分子機構に関する研究 結核菌潜伏感染の分子機構と治療・予防戦略

    小林和夫, 前倉亮治, 北田清悟, 松本壮吉, 藤原永年, 大原直也, 阿戸学, 大西和夫, 高橋宜聖, 岡部真裕子

    抗酸菌感染症の発症・診断・治療・新世代予防技術に係わる分子機構に関する研究 平成20年度 総括・分担研究報告書   15 - 19   2009

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    J-GLOBAL

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  • 結核菌による臓器侵襲の分子機構

    大原直也

    平成20年度社会保障国際協力推進研究事業(国際医学研究事業)日米医学協力研究計画結核・ハンセン病専門部会年次報告書 (研究代表者 結核研究所 菅原勇)   2009

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  • 日本株BCG Tokyo 172-1 Ⅰ型とⅡ型の遺伝学的検討と新規BCG宿主ベクター系の作製

    大原直也

    平成20年度政策創薬総合研究事業 細菌性ベクター及び粘膜アジュバントを用いた新興・再興感染症に対する新規予防・治療法の開発研究報告書(研究代表者 国立感染症研究所前山順一)   2009

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  • 抗酸菌(マイコバクテリア)感染症

    大原直也, 小林和夫

    コンパクト内科学   418 - 420   2009

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  • Porphyromonas gingivalis ECFシグマ因子PG0162変異株の性状解析 Reviewed

    菊池 有一郎, 大原 直也, 上田 青海, 平井 要, 柴田 幸永, 中山 浩次, 藤村 節夫

    Journal of Oral Biosciences   50 ( Suppl. )   212 - 212   2008.9

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  • 結核菌による臓器侵襲の分子機構

    大原直也

    平成19年度社会保障国際協力推進研究事業日米医学協力研究計画結核・ハンセン病専門部会年次報告書 (研究代表者 結核研究所 菅原勇)   2008

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  • 新規結核ワクチンとしてのリコンビナントBCGに関する研究

    大原直也

    平成19年度新興・再興感染症研究事業(アジア地域との研究ネットワークの活用による多剤耐性結核の制御に関する研究)研究報告書 (研究代表者 近畿中央胸部疾患センター 岡田全司)   2008

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  • 組換えBCGワクチンの改良・開発の研究

    大原直也

    新興・再興感染症研究事業(アジア地域との研究ネットワークの活用による多剤耐性結核の制御に関する研究)平成17年度~19年度総合研究報告書 (研究代表者 近畿中央胸部疾患センター 岡田全司)   2008

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  • 新規結核ワクチンとしてのリコンビナントBCGに関する研究

    大原直也

    平成19年度新興・再興感染症研究事業(有用な結核対策(BCG及び結核感染特異的診断に関する費用対効果分析等)に関する研究)研究報告書 (研究代表者 近畿中央胸部疾患センター 坂谷光則)   2008

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  • 結核菌

    大原直也, 小林和夫

    バイオセーフティの事典―病原微生物とバイオハザード対策の実際―   194 - 197   2008

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  • 組換えBCGワクチンの改良・開発の研究

    大原直也

    新興・再興感染症研究事業(有用な結核対策(BCG及び結核感染特異的診断に関する費用対効果分析等)に関する研究)平成17年度~19年度総合研究報告書 (研究代表者 近畿中央胸部疾患センター 坂谷光則)   2008

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  • 多価ワクチンとしての活用を目指したBCG宿主-ベクター系の改良

    大原直也

    平成19年度政策創薬総合研究事業 細菌性ベクター及び粘膜アジュバントを用いた新興・再興感染症に対する新規予防・治療法の開発研究報告書(研究代表者 国立感染症研究所前山順一)   2008

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  • ハンセン病

    大原直也

    微生物の事典   470 - 471   2008

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  • 新規結核ワクチンとしてのリコンビナントBCGに関する研究

    大原直也

    平成18年度新興・再興感染症研究事業(アジア地域との研究ネットワークの活用による多剤耐性結核の制御に関する研究)研究報告書 (研究代表者 近畿中央胸部疾患センター 岡田全司)   2007

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  • 結核菌による臓器侵襲の分子機構

    大原直也

    平成18年度国際医学研究協力事業日米医学協力研究計画結核・ハンセン病専門部会年次報告書 (研究代表者 結核研究所 菅原勇)   2007

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  • 結核

    岡部真裕子, 大原直也, 小林和夫

    保健の科学   49 ( 10 )   691 - 697   2007

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  • TNF-α, lipopolysaccharide, and peptidoglycan induce cell fusion independently of RANKL at the latest step of differentiation of RAW264.7 cells into osteoclast-like cells. Reviewed

    Hotokezaka H, Sakai E, Ohara N, Hotokezaka Y, Gonzales C, Matsuo, K, Fujimura Y, Yoshida N, Nakayama K

    J Cell Biochem.   277 ( 21 )   18545 - 18551   2007

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  • TNF-alpha, lipopolysaccharide, and peptidoglycan induce cell fusion independently of RANKL at the latest step of differentiation of RAW264.7 cells into osteoclast-like cells

    Hotokezaka H, Sakai E, Ohara N, Hotokezaka Y, Gonzales C, Matsuo K, Fujimura Y, Yoshida N, Nakayama K

    Journal of Cellular Biochemistry   101 ( 1 )   122 - 134   2007

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  • 新規結核ワクチンとしてのリコンビナントBCGに関する研究

    大原直也

    平成18年度新興・再興感染症研究事業(有用な結核対策(BCG及び結核感染特異的診断に関する費用対効果分析等)に関する研究)研究報告書 (研究代表者 近畿中央胸部疾患センター 坂谷光則)   2007

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  • A TLR4 gene polymorphism is associated with moderate and severe periodontitis in the Japanese population

    Fukusaki Tsuyoshi, Yoshimura Atsutoshi, Ohara Naoya, Muraoka Yuki, Yoshiura koh-ichiro, Hara Yoshitaka

    Program and Abstracts of Annual Meeting of the Japanese Society of Periodontology   2006 ( 0 )   48 - 48   2006

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    Publisher:特定非営利活動法人 日本歯周病学会  

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  • Induction and prevention of virus-associated malignant lymphoma by serial transmission of EBV-related virus from cynomolgus by blood transfusion in rabbits.

    Koirala TR, Hayashi K, Z Jin, Onoda S, Tanaka T, Oda W, Ichimura K, Ohara N, Oka T, Yamada M, Yoshino T

    Acta Med Okayama   2004

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  • 新しい結核ワクチンの開発とELISPOT assay(自動解析)を用いたT細胞活性化によるワクチン効果の解析

    田中 高生, 喜多 洋子, 井上 義一, 坂谷 光則, 岡田 全司, 桑山 さち子, 村木 裕美子, 稲永 由紀子, 金丸 典子, 橋元 里実, 高井 寛子, 渡邊 悠子, 岡田 知佳, 森 珠里, 石崎 邦子, 松本 久美, 岡 美穂, 黒川 恵理, 吉田 栄人, 金田 安史, 大原 直也, 内藤 真理子, 山田 毅, 松本 壮吉, Reed Steven, Skeiky Yasir, Gillis Steven

    結核   78 ( 3 )   278 - 278   2003.3

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  • Novel DNA and recombinant BCG vaccinations against tuberculosis by the augmentation of cytotoxic activity

    M Okada, S Yoshida, N Ohara, T Yamada, Y Kaneda, T Tanaka, Y Kita, S Kuwayama, Y Muraki, Y Inanaga, N Kanamaru, Y Inoue, M Matsumoto, K Kimura, M Sakatani, T Mori

    FASEB JOURNAL   16 ( 4 )   A308 - A309   2002.3

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    Web of Science

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  • DNA and recombinant BCG vaccination against tuberculosis by the augmentation of cytotoxic activity

    M Okada, T Tanaka, S Yoshida, N Ohara, T Yamada, Y Inoue, S Minamoto, M Sakatani, T Mori

    FASEB JOURNAL   15 ( 5 )   A1008 - A1008   2001.3

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    Web of Science

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  • ヒトマラリアの防御抗原を発現する組換えBCGワクチンの作製

    雪竹 英治, 大原 直也, 神原 廣二, 山田 毅, 松本 壮吉

    日本細菌学雑誌   56 ( 1 )   316 - 316   2001.2

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  • 抗酸菌抗原DNAワクチンとrBCGワクチンのモルモット免疫応答及び結核防御効果

    山本 三郎, 野島 康弘, 梅森 清子, 野間口 博子, 大原 直也, 山田 毅, 山本 十糸子, 佐藤 由紀夫, 松本 壮吉, 松尾 和浩

    日本細菌学雑誌   56 ( 1 )   315 - 315   2001.2

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  • Analysis of CTL activity in the patients with multi-drug resistant tuberculosis and development of new vaccination by the induction of CTL using murine model.

    M Okada, Y Katayama, Y Inoue, M Yotsumoto, K Yasumitsu, S Hosoe, S Yoshida, N Ohara, T Yamada, M Sakatani, T Mori

    FASEB JOURNAL   14 ( 6 )   A1061 - A1061   2000.4

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  • 抗酸菌の重感染によるHIV-1複製亢進とサイトカインについて

    北浦 英樹, 大原 直也, 山田 毅

    日本細菌学雑誌   55 ( 2 )   211 - 211   2000.4

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  • 顎関節症患者の関節滑液中の抗体が認識する抗酸菌の44kDa蛋白質について

    安達 紀子, 松本 壮吉, 松尾 長光, 大原 直也, 内藤 真理子, 雪竹 英治, 山田 毅

    日本細菌学雑誌   54 ( 1 )   312 - 312   1999.2

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  • 分泌型リボゾーム蛋白質L7/L12の遅延型アレルギー反応について

    北浦 英樹, 大原 直也, 西山 毅, 木之本 雅通, 山田 毅

    日本細菌学雑誌   54 ( 1 )   318 - 318   1999.2

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  • 結核発病における菌側の因子;抗酸菌の抗原の研究の意味

    山田毅, 大原直也, 松本壮吉, 松尾長光, 北浦英樹, 高野美貴子, 雪竹英治, 和田直子, 西山毅, 内藤真理子, 木ノ本雅道, 木村誠

    結核   70   639 - 644   1995

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Presentations

  • チミジル酸合成酵素ThyXの過剰発現はBCGの増殖を促進する

    竜門亜矢子、中山真彰、有村友紀、阿戸学、中島千絵、鈴木定彦、小崎弘貴、大原直也

    第69回日本細菌学会中国・四国支部総会  2016 

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    Event date: 2016.10.15 - 2016.10.16

    Language:Japanese   Presentation type:Oral presentation (general)  

    Venue:岡山  

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  • PI3K/Akt経路に対するP. gingivalis ジンジパインの役割

    中山真彰、内藤真理子、中山浩次、大原直也

    第58回歯科基礎医学会学術集会・総会  2016 

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    Event date: 2016.8.24 - 2016.8.26

    Language:English   Presentation type:Poster presentation  

    Venue:札幌  

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  • Porphyromonas gingivalis ECFシグマ因子変異株における菌体表層性状の解析

    菊池有一郎、国分栄二、柴山和子、大原直也、中山浩次、石原和幸

    第58回歯科基礎医学会学術集会・総会  2016 

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    Event date: 2016.8.24 - 2016.8.26

    Language:English   Presentation type:Poster presentation  

    Venue:札幌  

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  • Porphyromonas gingivalis ECFシグマ因子変異株の性状解析

    第57回歯科基礎医学会学術集会・総会  2015 

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  • Porphyromonas gingivalis ジンジパインによるPI3K/Akt経路の抑制効果とその意義

    第57回歯科基礎医学会学術集会・総会  2015 

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  • BCG thyX欠損株の作製とその性状解析

    第57回歯科基礎医学会学術集会・総会  2015 

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  • BCGにおけるcyclic-di-GMP菌体内合成の影響

    第57回歯科基礎医学会学術集会・総会  2015 

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  • BCGにおけるcyclic-di-GMP菌体内過剰発現の影響

    第68回日本細菌学会中国・四国支部総会  2015 

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  • BCG thyX変異株の作製とその性状解析

    第68回日本細菌学会中国・四国支部総会  2015 

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  • P. gingivalisジンジパインによるPI3K/Akt経路攪乱の分子解析

    第68回日本細菌学会中国・四国支部総会  2015 

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  • Mycobacterium smegmatis バクテリオファージの長期保存状態における生存率

    第68回日本細菌学会中国・四国支部総会  2015 

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  • BCG Tokyo 172-1に存在する2種類のサブポピュレーション間での遺伝子発現の比較検討

    第62回日本細菌学会中国四国支部総会  2009 

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  • BCGにおけるチミジル酸合成酵素ThyXの意義

    第81回日本細菌学会総会  2009 

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  • Porphyromonas gingivalis のPG1385(TPR domain protein)の解析

    第81回日本細菌学会総会  2009 

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  • Porphyromonas gingivalis ECFシグマ因子PG0162変異株の性状解析

    第81回日本細菌学会総会  2009 

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  • BCG Tokyo 172-I型および-II型株におけるsliding能の差異

    第81回日本細菌学会総会  2009 

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  • Construction of a host-vector system in Mycobacterium using alternative thymidylate synthase ThyX

    第44回日米医学研究協力会議結核・ハンセン病専門部会合同会議  2009 

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  • Porphyromonas gingivalis ECFシグマ因子PG0162変異株の性状解析

    第50回歯科基礎医学会学術大会ならびに総会  2009 

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  • thyX遺伝子欠損株の作製による抗酸菌チミジル酸合成酵素の解析

    第78回実験結核研究会  2008 

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  • Contribution of NADPH oxidative activity of thymidylate synthase ThyX to growth of BCG

    第43回日米医学研究協力会議結核・ハンセン病専門部会合同会議  2008 

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  • CD30L/CD30 signaling by T-T cell interaction augments Th1 responses

    第37回日本免疫学会総会・学術集会  2008 

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  • MIP-T3のC末端は微小管結合と蛋白質の安定性制御に重要である

    第31回日本分子生物学会年会、第81回日本生化学会大会合同大会  2008 

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  • Association between a TLR4 gene polymorphism and moderate/severe periodontitis in the Japanese population

    第36回日本免疫学会総会・学術集会  2007 

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  • Porphyromonas gingivalis ATCC33277株の全ゲノム塩基配列決定

    第1回日本ゲノム微生物学会  2007 

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  • 細菌による破骨細胞分化の制御

    第80回日本細菌学会総会  2007 

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  • Porphyromonas gingivalis ATCC33277株の全ゲノム塩基配列決定

    第80回日本細菌学会総会  2007 

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  • One of the bacterial tetratricopeptide repeat-containing proteins is involved in virulence of the periodontal pathogen Porphyromonas gingivalis

    第2回インターフェイス口腔健康科学国際シンポジウム  2007 

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  • Porphyromonas gingivalis のTPRドメイン蛋白質欠損株の解析

    第80回日本細菌学会総会  2007 

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  • MIP-T3と微小管との結合機構の解析

    平成19年度日本生化学会九州支部例会  2007 

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  • Porphyromonas gingivalis ECFシグマ因子群の遺伝子挿入変異株の作製

    第49回歯科基礎医学会学術大会ならびに総会  2007 

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  • Porphyromonas gingivalis ATCC33277株の全ゲノム塩基配列決定

    第49回歯科基礎医学会学術大会ならびに総会  2007 

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  • Construction and characterization of the thymidylate synthase mutants derived from BCG

    中日米結核セミナーおよび42回日米医学研究協力会議結核・ハンセン病専門部会合同会議  2007 

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  • Efficacy of recombinant BCG vaccine secreting IL-15/Ag85B fusion protein on protection against Mycobacterium tuberculosis

    中日米結核セミナーおよび42回日米医学研究協力会議結核・ハンセン病専門部会合同会議  2007 

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  • Recombinant BCG vaccines: current status. 2nd Nagasaki Symposium on Tropical and Emerging Infectious Diseases

    第2回長崎熱帯医学新興感染症シンポジウム  2007 

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