Faculty Profiles - OKAMOTO Kuniaki

MISC - OKAMOTO Kuniaki

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Application of Organoids and Extracellular Vesicles to Cancer Research

江口傑徳, 江口傑徳, 十川千春, 岡元邦彰

Journal of Oral Biosciences Supplement (Web) 2020 2020

分子シャペロントリオによるエクソソーム制御,腫瘍悪性化およびマクロファージ分極について

江口傑徳, 小野喜章, 小野喜章, 河合穂高, チャンチェンマン, 十川千春, 奥舎有加, 岡元邦彰

日本臨床ストレス応答学会大会抄録集 14th 2019

新しい腫瘍オルガノイド多元評価システムの開発

十川チハル, 江口傑徳, 江口傑徳, 大山和美, 奥舎有加, 中野敬介, 十川紀夫, 岡元邦彰

Journal of Oral Biosciences Supplement (Web) 2019 2019

ニコチンは口腔扁平上皮癌細胞のセツキシマブ耐性を促進する

清水理恵子, 伊原木聰一郎, 江口傑徳, 奥井達雄, 高畠清文, 河合穂高, 小野喜章, 岡元邦彰, 長塚仁, 佐々木朗

岡山歯学会雑誌 37 ( 2 ) 80 - 81 2018.12

口腔扁平上皮癌診断・治療における分子シャペロンHSP90含有細胞外小胞の可能性

小野喜章, 江口傑徳, 十川千春, 奥舎有加, 河合穂高, 中野敬介, 佐々木朗, 岡元邦彰, 小崎健一

Journal of Oral Biosciences Supplement 2018 333 - 333 2018.9

核内およびエクソソーム中に存在するMMPs/PEXの癌進展における役割とその抑制

奥舎有加, 江口傑徳, 十川千春, 小野喜章, 奥井達雄, 中野敬介, 岡元邦彰

Journal of Oral Biosciences Supplement 2018 150 - 150 2018.9

癌の治療抵抗性と転移におけるHSP90およびMMP3の役割

江口傑徳, 小野喜章, 奥舎有加, 十川千春, 内部健太, 中野敬介, 奥井達雄, 滝川正春, 岡元邦彰, カルダーウッド・スチュアート

Journal of Oral Biosciences Supplement 2018 142 - 142 2018.9

癌の治療抵抗性と転移におけるHSP90およびMMP3の役割

江口傑徳, 江口傑徳, 小野喜章, 小野喜章, 奥舎有加, 十川千春, 内部健太, 中野敬介, 中野敬介, 奥井達雄, 滝川正春, 岡元邦彰, CALDERWOOD SK

Journal of Oral Biosciences Supplement (Web) 2018 2018

急速転移性癌細胞株由来細胞外小胞による破骨細胞分化および癌細胞浸潤の制御

板垣まみ, 十川千春, 奥舎有加, 江口傑徳, 江口傑徳, 小野喜章, 岡元邦彰

Journal of Oral Biosciences Supplement (Web) 2018 2018

口腔扁平上皮癌診断・治療における分子シャペロンHSP90含有細胞外小胞の可能性

小野喜章, 江口傑徳, 江口傑徳, 十川千春, 奥舎有加, 河合穂高, 中野敬介, 佐々木朗, 岡元邦彰, 小崎健一

Journal of Oral Biosciences Supplement (Web) 2018 2018

破骨細胞分化を負に制御するRabタンパク質の同定

奥舎有加, 板垣まみ, 十川千春, 江口傑徳, 江口傑徳, 坂井詠子, 筑波隆幸, 岡元邦彰

Journal of Oral Biosciences Supplement (Web) 2018 2018

核内およびエクソソーム中に存在するMMPs/PEXの癌進展における役割とその抑制

奥舎有加, 江口傑徳, 江口傑徳, 十川千春, 小野喜章, 小野喜章, 奥井達雄, 中野敬介, 岡元邦彰

Journal of Oral Biosciences Supplement (Web) 2018 2018

転移性の異なる細胞株を用いた核内MMP3の機能解析とその役割

奥舎有加, 江口傑徳, 十川千春, 奥井達雄, 中野敬介, 小崎健一, 岡元邦彰

日本薬理学会近畿部会プログラム・要旨集 133rd 2018

新規高分子量Gタンパク質Rab44はCa²⁺流入及びNFATc1経路を介して破骨細胞分化を負に制御する

山口優, 坂井詠子, 岡元邦彰, 鍜治屋浩, 岡部幸司, 門脇知子, 筑波隆幸

日本生化学会大会(Web) 90th 2017

破骨細胞分化を負に制御する新規Kelchタンパク質の同定と機能解析

楢原峻, 楢原峻, 坂井詠子, 山口優, 楢原春菜, 門脇知子, 岡元邦彰, 朝比奈泉, 筑波隆幸

日本生化学会大会(Web) 90th 2017

破骨細胞分化を制御する新規Rabタンパク質の同定と解析

山口優, 坂井詠子, 岡元邦彰, 門脇知子, 筑波隆幸

日本病態プロテアーゼ学会学術集会プログラム抄録集 22nd 2017

口腔扁平上皮癌細胞由来エクソソームに含まれる分子シャペロンについての検討

小野喜章, 小野喜章, 江口傑徳, 十川千春, 村上純, 藤原敏史, 藤原敏史, 笠井智成, 妹尾昌治, 佐々木朗, 小崎健一, 岡元邦彰

臨床ストレス応答学会大会抄録集 12th 2017

癌や炎症性疾患において分子シャペロンHSPsが誘導される新たな経路:核内MMPとヘテロクロマチンタンパク質の相互作用

江口傑徳, 江口傑徳, 奥舎有加, 小崎健一, 岡元邦彰, CALDERWOOD Stuart K

臨床ストレス応答学会大会抄録集 12th 2017

Mmp遺伝子を標的とし,高転移性癌細胞の生存と転移を抑制する

奥舎有加, 江口傑徳, 十川千春, 中野敬介, 岡元邦彰, 小崎健一

Journal of Oral Biosciences Supplement (Web) 2017 2017

転移性の異なる癌細胞株の網羅的遺伝子発現解析による浸潤・転移促進因子の探索と機能解明

奥舎有加, 江口傑徳, 十川千春, 中野敬介, 小崎健一, 岡元邦彰

硬組織再生生物学会学術大会・総会プログラム・抄録集 26th 2017

破骨細胞分化を制御する新規Rabタンパク質の同定と機能解析

山口優, 坂井詠子, 岡元邦彰, 鍛治屋浩, 岡部幸司, 筑波隆幸

Journal of Oral Biosciences Supplement (Web) 2017 2017

tert-ButylhydroquinoneによるIrf8とMafBの発現増加を介した破骨細胞分化抑制機構

坂井詠子, 山口優, 福間裕, 西下一久, 岡元邦彰, 筑波隆幸

日本生化学会大会(Web) 89th 2016

Keap1/Nrf2の破骨細胞と骨芽細胞分化における役割

坂井詠子, 福間裕, 山口優, 西下一久, 岡元邦彰, 筑波隆幸

日本生化学会大会(Web) 88th 2015

歯科用モノマーによる破骨細胞分化に対する効果

稲光宏之, 岡元邦彰, 坂井詠子, 村田比呂司, 筑波隆幸

Journal of Oral Biosciences Supplement 2014 180 - 180 2014.9

Chemical constituent of Sanguisorba officinalis inhibits osteoclastogenesis

Eiko Sakai, Mayumi Iwatake, Yutaka Fukuma, Kazuhisa Nishishita, Kuniaki Okamoto, Takashi Tanaka, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 124 232P - 232P 2014

コバルトプロトポルフィリンの破骨細胞分化および活性化に対する影響

八島由佳, 八島由佳, 岡元邦彰, 坂井詠子, 西下一久, 筑波隆幸

Journal of Oral Biosciences Supplement (Web) 2014 ROMBUNNO.P2‐33 (WEB ONLY) 2014

Nrf2遺伝子欠損マウスにおける酸化ストレスと骨代謝

坂井詠子, 山口優, 福間裕, 菅原めぐみ, 菅原めぐみ, 西下一久, 岡元邦彰, 筑波隆幸

日本生化学会大会(Web) 87th 2014

Up-regulation mechanisms of endosomal/lysosomal proteins during osteoblast differentiation

Erika Yoneshima, Kuniaki Okamoto, Eiko Sakai, Noriaki Yoshida, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 121 170P - 170P 2013

Cafestol prevents osteoclastogenesis via impairment of NFATc1 expression and blocking of Erk phosphorylation

Yutaka Fukuma, Eiko Sakai, Megumi Sugawara, Erika Yoneshima, Kazuhisa Nishishita, Kuniaki Okamoto, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 121 249P - 249P 2013

Identification of two transcrips and in vivo promoter analysis for cathepsin E

Kuniaki Okamoto, Yoshiko Okamoto, Eiko Sakai, Kazuhisa Nishishita, Kenji Yamamoto, Takayuki Tstikuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 118 259P - 259P 2012

Accumulation of NADPH oxidase (NOX2) and increased oxidative stress in cathepsin E-deficient macrophages

Takayuki Tsukuba, Tomoko Kadowaki, Kuniaki Okamoto, Kenji Yamamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 118 256P - 256P 2012

Fisetinl prevents osteoclastogenesis via impairment of NFATc1 expression and blocking of Erk phosphorylation

Megumi Sugawara, Eiko Sakai, Kazuhisa Nishishita, Yutaka Fukuma, Kuniaki Okamoto, Noriaki Yoshida, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 118 164P - 164P 2012

Receptor activator of NF-kappa B ligand dependent heme oxygenase-1 suppression is required for high mobility group box 1 release, caspase-3 activation, and subsequent osteoclastogenesis

Eiko Sakai, Megumi Shimada, Kazuhisa Nishishita, Yutaka Fukuma, Kuniaki Okamoto, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 115 219P - 219P 2011

Nrf2 activators inhibit osteoclastogenesis induced by receptor activator of NF-kappa B ligand

Reiko Fumimoto, Hiroshi Sakamoto, Yu Yamaguchi, Eiko Sakai, Kuniaki Okamoto, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 115 219P - 219P 2011

tert-Butylhydroquinoneの破骨細胞形成と骨吸収活性への影響

山口優, 坂井詠子, 岡元邦彰, 筑波隆幸

Journal of Oral Biosciences 53 ( Supplement ) 2011

Transcription factor Sp1 regulates the expression of the murine cathepsin E gene in gastric adenosarcoma cells

Kuniaki Okamoto, Yoshiko Okamoto, Tomoyo Kawakubo, Kenji Yamamoto, Takayuki Tsukuba

JOURNAL OF PHARMACOLOGICAL SCIENCES 112 216P - 216P 2010

Autophagy impairment associated with increased aberrant mitochondria and oxidative stress in cathepsin E-deficient macrophages

Takayuki Tsukuba, Michiyo Yanagawa, Tomoko Kadowaki, Yoshiko Okamoto, Kuniaki Okamoto, Kenji Yamamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 112 216P - 216P 2010

カテプシンE欠損における破骨細胞分化への影響

岡元邦彰, 坂井詠子, 西下一久, 福間裕, 坂元裕, 文元玲子, 山口優, 山本健二, 筑波隆幸

生化学 2010

Carbon dioxide laser irradiation stimulates mineralization in rat dental pulp cells

Y. Yasuda, E. Ohtomo, T. Tsukuba, K. Okamoto, T. Saito

INTERNATIONAL ENDODONTIC JOURNAL 42 ( 10 ) 940 - 946 2009.10

Language：English Publisher：WILEY-BLACKWELL PUBLISHING, INC

P>Aim
To examine the effect of carbon dioxide laser irradiation on mineralization in dental pulp cells.
Methodology
Rat dental pulp cells were irradiated with a carbon dioxide laser at 2 W output power for 20, 40 and 60 s, and were cultured in ascorbic acid and beta-glycerophosphate containing media. Cell viability was examined 24 h after laser irradiation by a modified MTT assay. Alizarin Red S staining was performed 10 days after laser irradiation. The amounts of secreted collagen from the cells after irradiation were quantified following Sirius Red staining. The expression levels of collagen type I and HSP47, collagen-binding stress protein, were analysed by real-time PCR. HSP47 protein expression was examined by Western blotting. Statistical analysis was performed using one-way analysis of variance (anova) followed by the Tukey's multiple comparison test.
Results
The cell viability was not affected by laser irradiation at 2 W for up to 40 s. However, it was significantly decreased by 20% at 60 s (P < 0.05). The amount of mineralization after 10 days of irradiation at 2 W for 40 s was significantly increased in comparison to the other conditions (P < 0.05). The extracellular collagen production was significantly increased by 73% on day 2 and 38% on day 4 after laser irradiation (P < 0.05). Although collagen type I gene expression was not changed by laser irradiation, HSP47 gene and protein expression was induced within 12 and 24 h, respectively.
Conclusions
These results suggested that carbon dioxide laser irradiation stimulated mineralization in dental pulp cells. The laser irradiation also increased HSP47 expression but not collagen gene expression.

DOI： 10.1111/j.1365-2591.2009.01598.x

Impaired chemotaxis and cell adhesion due to decrease in several cell-surface receptors in cathepsin E-deficient macrophages

Takayuki Tsukuba, Michiyo Yanagawa, Kuniaki Okamoto, Yoshiko Okamoto, Yoshiyuki Yasuda, Keiichi I. Nakayama, Tomoko Kadowaki, Kenji Yamamoto

JOURNAL OF BIOCHEMISTRY 145 ( 5 ) 565 - 573 2009.5

Language：English Publisher：OXFORD UNIV PRESS

Cathepsin E is an endo-lysosomal aspartic proteinase exclusively present in immune system cells. Previous studies have shown that cathepsin E-deficient (CatE(/)) mice display aberrant immune responses such as atopic dermatitis and higher susceptibility to bacterial infection. However, the mechanisms underlying abnormal immune responses induced by cathepsin E deficiency are still unclear. In this study, we found that the cell-surface levels of chemotactic receptors, including chemokine receptor (CCR)-2 and N-formyl peptide receptors (FPRs), were clearly diminished in CatE(/)macrophages compared with those in wild-type cells. Consistently, chemotaxis of CatE(/)macrophages to MCP-1 and N-formyl-methionyl-leucyl-phenylalanine was also decreased. Similar to the chemotactic receptors, the surface expressions of the adhesion receptors CD18 (integrin (2)) and CD 29 (integrin (1)) in CatE(/) macrophages were significantly decreased, thereby reducing cell attachment of CatE(/) macrophages. These results indicate that the defects in chemotaxis and cell adhesion are likely to be involved in the imperfect function of CatE(/)macrophages.

DOI： 10.1093/jb/mvp016

カテプシンE欠損マクロファージにおけるオートファジーの低下とそれに伴うミトコンドリア機能異常と酸化ストレスの上昇

筑波隆幸, 門脇知子, 坂井詠子, 岡元邦彰, 山本健二

Journal of Oral Biosciences 51 ( Supplement ) 2009

Determination of active site of lysine-specific cysteine proteinase (Lys-gingipain) by use of a Porphyromonas gingivalis plasmid system

Yutaka Ishida, JinPing Hu, Eiko Sakai, Tomoko Kadowaki, Kenji Yamamoto, Takayuki Tsukuba, Yuzo Kato, Koji Nakayama, Kuniaki Okamoto

ARCHIVES OF ORAL BIOLOGY 53 ( 6 ) 538 - 544 2008.6

Language：English Publisher：PERGAMON-ELSEVIER SCIENCE LTD

Porphyromonas gingivalis, a major etiological bacterium of periodontal diseases, produces a unique lysine-specific cysteine proteinase (Lys-gingipain, Kgp) implicated in the virulence of this organism. Our observations show the expression of a catalytically active recombinant Kgp in a P. gingivalis Kgp-null mutant and the restoration of its functions by the use of a shuttle plasmid vector stable in P. gingivalis. The Kgp-expressing mutant exhibited a similar catalytic activity to that of the wild-type strain. This mutant also restored the ability to form black-pigmented colonies on blood agar plates and to generate a 19-kDa haemoglobin receptor protein responsible for haemoglobin binding. In order to establish the importance of the active-site Cys residue and elucidate its role in bacterial black pigmentation we constructed three Kgp mutants with changed potential active-site Cys residues. The cells expressing a single mutation (C476A) showed the high Kgp activity and the black pigmentation. In contrast, the cells expressing the single mutant (C477A) and the double mutant (C476A/C477A) exhibited neither Kgp activity nor black pigmentation. These results indicate that the 477th Cys residue is essential for both the Kgp activity and the black pigmentation of P. gingivalis. (C) 2008 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.archoralbio.2008.01.004

Association of cathepsin E deficiency with the increased territorial aggressive response of mice

Naoki Shigematsu, Takaichi Fukuda, Tsuneyuki Yamamoto, Tsuyoshi Nishioku, Taku Yamaguchi, Masaru Himeno, Keiichi I. Nakayama, Takayuki Tsukuba, Tomoko Kadowaki, Kuniaki Okamoto, Shun Higuchi, Kenji Yamamoto

JOURNAL OF NEUROCHEMISTRY 105 ( 4 ) 1394 - 1404 2008.5

Language：English Publisher：WILEY

Cathepsin E is an endolysosomal aspartic proteinase predominantly expressed in cells of the immune system, but physiological functions of this protein in the brain remains unclear. In this study, we investigate the behavioral effect of disrupting the gene encoding cathepsin E in mice. We found that the cathepsin E-deficient (CatE-/-) mice were behaviorally normal when housed communally, but they became more aggressive compared with the wild-type littermates when housed individually in a single cage. The increased aggressive response of CatE-/- mice was reduced to the level comparable to that seen for CatE+/+ mice by pretreatment with an NK-1-specific antagonist. Consistent with this, the neurotransmitter substance P (SP) level in affective brain areas including amygdala, hypothalamus, and periaqueductal gray was significantly increased in CatE-/- mice compared with CatE+/+ mice, indicating that the increased aggressive behavior of CatE-/- mice by isolation housing followed by territorial challenge is mainly because of the enhanced SP/NK-1 receptor signaling system. Double immunofluorescence microscopy also revealed the co-localization of SP with synaptophysin but not with microtubule-associated protein-2. Our data thus indicate that cathepsin E is associated with the SP/NK-1 receptor signaling system and thereby regulates the aggressive response of the animals to stressors such as territorial challenge.

DOI： 10.1111/j.1471-4159.2008.05242.x

Dentin phosphophoryn promotes cellular migration of human dental pulp cells

Yoshiyuki Yasuda, Masanobu Izumikawa, Kuniaki Okamoto, Takayuki Tsukuba, Takashi Saito

JOURNAL OF ENDODONTICS 34 ( 5 ) 575 - 578 2008.5

Language：English Publisher：ELSEVIER SCIENCE INC

Dentin phosphophoryn (DPP) is a dentin sialophosphoprotein gene product that has an RGD motif and repeat sequences of aspartic acid and phosphoserine. To date, the function of DPP in the early stage of reparative dentin formation still remains unclear. The objective of this study was to evaluate the effects of DPP on pulp cell migration and proliferation. DPP promoted cell migration in a concentration-dependent manner, thus increasing it by about 3-fold at 1000 ng/mL compared with the control, but it had no effect on cell proliferation. Dephosphorylated DPP also promoted cell migration, similarly to DPP. However, cell migration was significantly suppressed by the addition of alpha v beta 3 integrin antibody to the medium. Furthermore, porcine DPP-derived RGD peptide, but not its mutant RAD peptide, significantly promoted cell migration. These results indicated that the RGD motif of DPP plays an important role in the migration of human dental pulp cells.

DOI： 10.1016/j.joen.2008.02.018

Berberine inhibits RANKL-induced osteoclast formation and survival through suppressing the NF-kappa B and Akt pathways

Jin-Ping Hu, Kazuhisa Nishishita, Eiko Sakai, Hajime Yoshida, Yuzo Kato, Takayuki Tsukuba, Kuniaki Okamoto

EUROPEAN JOURNAL OF PHARMACOLOGY 580 ( 1-2 ) 70 - 79 2008.2

Language：English Publisher：ELSEVIER SCIENCE BV

Berberine, an isoquinoline alkaloid isolated from several medicinal plants, has been reported to possess anti-bacterial, anti-inflammatory and antitumor properties. Although berberine also inhibits osteoclastogenesis and bone resorption, the molecular machinery for its inhibitory effects remains unknown. This study focused on the suppressive effects of berberine on receptor activator of nuclear factor kappa B (NF-kappa B) ligand (RANKL)induced osteoclastogenesis and survival. Berberine inhibited RANKL-mediated osteoclast fort-nation and survival while having no cytotoxic effects on bone marrow macrophages or osteoblastic cells. Berberine attenuated RANKL-induced activation of NF-kappa B through inhibiting phosphorylation at the activation loop of I kappa B alpha kinase, phosphorylation and degradation of I kappa B alpha, and NF-kappa B p65 nuclear translocation. RANKL-induced Akt phosphorylation was strongly inhibited by berberine; however, neither monocyte/macrophage-colony stimulating factor (M-CSF)-induced nor insulin-induced Akt activation was inhibited by the drug. Under M-CSF- and RANKL-deprived condition, berberine increased the active form of caspase-3 in osteoclasts. By contrast, berberine did not potentiate the activation of caspase-3 in M-CSF-deprived bone marrow macrophages. These findings indicate that berberine inhibits osteoclast formation and survival through suppression of NF-kappa B and Akt activation and that both pathways in the osteoclast lineage are highly sensitive to berberine treatment. (C) 2007 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.ejphar.2007.11.013

Berberine inhibits RANKL-induced osteoclast formation and survival through suppressing the NF-kappa B and Akt pathways

Jin-Ping Hu, Kazuhisa Nishishita, Eiko Sakai, Hajime Yoshida, Yuzo Kato, Takayuki Tsukuba, Kuniaki Okamoto

EUROPEAN JOURNAL OF PHARMACOLOGY 580 ( 1-2 ) 70 - 79 2008.2

Language：English Publisher：ELSEVIER SCIENCE BV

Berberine, an isoquinoline alkaloid isolated from several medicinal plants, has been reported to possess anti-bacterial, anti-inflammatory and antitumor properties. Although berberine also inhibits osteoclastogenesis and bone resorption, the molecular machinery for its inhibitory effects remains unknown. This study focused on the suppressive effects of berberine on receptor activator of nuclear factor kappa B (NF-kappa B) ligand (RANKL)induced osteoclastogenesis and survival. Berberine inhibited RANKL-mediated osteoclast fort-nation and survival while having no cytotoxic effects on bone marrow macrophages or osteoblastic cells. Berberine attenuated RANKL-induced activation of NF-kappa B through inhibiting phosphorylation at the activation loop of I kappa B alpha kinase, phosphorylation and degradation of I kappa B alpha, and NF-kappa B p65 nuclear translocation. RANKL-induced Akt phosphorylation was strongly inhibited by berberine; however, neither monocyte/macrophage-colony stimulating factor (M-CSF)-induced nor insulin-induced Akt activation was inhibited by the drug. Under M-CSF- and RANKL-deprived condition, berberine increased the active form of caspase-3 in osteoclasts. By contrast, berberine did not potentiate the activation of caspase-3 in M-CSF-deprived bone marrow macrophages. These findings indicate that berberine inhibits osteoclast formation and survival through suppression of NF-kappa B and Akt activation and that both pathways in the osteoclast lineage are highly sensitive to berberine treatment. (C) 2007 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.ejphar.2007.11.013

カテプシンE欠損はオートファジーの低下とそれに伴うミトコンドリア機能異常と酸化ストレスを引き起こす。

筑波隆幸, 柳川三千代, 門脇知子, 岡本美子, 岡元邦彰, 中山敬一, 山本健二

生化学 2008

カテプシンE欠損マウスにおける高脂血症および脂肪肝の誘導

門脇知子, 岡元邦彰, 山本健二, 筑波隆幸

Journal of Oral Biosciences 50 ( Supplement ) 2008

カテプシンE欠損による遊走能および細胞接着能の低下

筑波隆幸, 柳川三千代, 岡元邦彰, 門脇知子, 山本健二

Journal of Oral Biosciences 50 ( Supplement ) 2008

Cathepsin E prevents tumor growth and metastasis by catalyzing the proteolytic release of soluble TRAIL from tumor cell surface

Tomoyo Kawakubo, Kuniaki Okamoto, Jun-Ichi Lwata, Masashi Shin, Yoshiko Okamoto, Atsushi Yasukochi, Keiichi I. Nakayama, Tomoko Kadowaki, Takayuki Tsukuba, Kenji Yamamoto

CANCER RESEARCH 67 ( 22 ) 10869 - 10878 2007.11

Language：English Publisher：AMER ASSOC CANCER RESEARCH

The aspartic proteinase cathepsin E is expressed predominantly in cells of the immune system and highly secreted by activated phagocytes, and deficiency of cathepsin E in mice results in a phenotype affecting immune responses. However, because physiologic substrates for cathepsin E have not yet been identified, the relevance of these observations to the physiologic functions of this protein remains speculative. Here, we show that cathepsin E specifically induces growth arrest and apoptosis in human prostate carcinoma tumor cell lines without affecting normal cells by catalyzing the proteolytic release of soluble tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) from the cell surface. The antitumor activity of cathepsin E was corroborated by in vivo studies with mice bearing human and mouse tumor transplants. Administration of purified cathepsin E into human tumor xenografts in nude mice dose-dependently induced apoptosis in the tumor cells to inhibit tumor growth. The growth, viability, and metastasis of mouse B16 melanoma cells were also more profound in cathepsin E-deficient mice compared with those in the syngeneic wild-type and transgenic mice overexpressing cathepsin E. Taken together, the number of apoptotic tumor cells, as well as tumor-infiltrating activated macrophages, was apparently reduced in cathepsin E-deficient mice compared with those in the other two groups, implying the positive correlation of endogenous cathepsin E levels with the extent of tumor suppression in vivo. These results thus indicate thatcathepsinEplaysasubstantialroleinhostdefense against tumor cells through TRAIL-dependent apoptosis and/or tumor-associated macrophage-mediated cytotoxicity.

DOI： 10.1158/0008-5472.CAN-07-2048

Construction of recombinant hemagglutinin derived from the gingipain-encoding gene of Porphyromonas gingivalis, identification of its target protein on erythrocytes, and inhibition of hemagglutination by an interdomain regional peptide. International journal

Eiko Sakai, Mariko Naito, Keiko Sato, Hitoshi Hotokezaka, Tomoko Kadowaki, Arihide Kamaguchi, Kenji Yamamoto, Kuniaki Okamoto, Koji Nakayama

Journal of bacteriology 189 ( 11 ) 3977 - 86 2007.6

Language：English

Porphyromonas gingivalis, an anaerobic gram-negative bacterium associated with chronic periodontitis, can agglutinate human erythrocytes. In general, hemagglutination can be considered the ability to adhere to host cells; however, P. gingivalis-mediated hemagglutination has special significance because heme markedly accelerates growth of this bacterium. Although a number of studies have indicated that a major hemagglutinin of P. gingivalis is intragenically encoded by rgpA, kgp, and hagA, direct evidence has not been obtained. We demonstrated in this study that recombinant HGP44(720-1081), a fully processed HGP44 domain protein, had hemagglutinating activity but that an unprocessed form, HGP44(720-1138), did not. A peptide corresponding to residues 1083 to 1102, which was included in HGP44(720-1138) but not in HGP44(720-1081), could bind HGP44(720-1081) in a dose-dependent manner and effectively inhibited HGP44(720-1081)-mediated hemagglutination, indicating that the interdomain regional amino acid sequence may function as an intramolecular suppressor of hemagglutinating activity. Analyses by solid-phase binding and chemical cross-linking suggested that HGP44 interacted with glycophorin A on the erythrocyte membrane. Glycophorin A and, more effectively, asialoglycophorin, which were added exogenously, inhibited HGP44(720-1081)-mediated hemagglutination. Treatment of erythrocytes with RgpB proteinase resulted in degradation of glycophorin A on the membrane and a decrease in HGP44(720-1081)-mediated hemagglutination. Surface plasmon resonance detection analysis revealed that HGP44(720-1081) could bind to asialoglycophorin with a dissociation constant of 3.0 x 10(-7) M. These results indicate that the target of HGP44 on the erythrocyte membrane appears to be glycophorin A.

DOI： 10.1128/JB.01691-06

Cathepsin E deficiency induces a novel form of lysosomal storage disorder showing the accumulation of lysosomal membrane sialoglycoproteins and the elevation of lysosomal pH in macrophages

Michiyo Yanagawa, Takayuki Tsukuba, Tsuyoshi Nishioku, Yoshiko Okamoto, Kuniaki Okamoto, Ryosuke Takii, Yoshihiro Terada, Keiichi I. Nakayama, Tomoko Kadowaki, Kenji Yamamoto

JOURNAL OF BIOLOGICAL CHEMISTRY 282 ( 3 ) 1851 - 1862 2007.1

Language：English Publisher：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Cathepsin E, an endolysosomal aspartic proteinase predominantly expressed in cells of the immune system, has an important role in immune responses. However, little is known about the precise roles of cathepsin E in this system. Here we report that cathepsin E deficiency (CatE(-/-)) leads to a novel form of lysosome storage disorder in macrophages, exhibiting the accumulation of the two major lysosomal membrane sialoglycoproteins LAMP-1 and LAMP-2 and the elevation of lysosomal pH. These striking features were also found in wild-type macrophages treated with pepstatin A and Ascaris inhibitor. Whereas there were no obvious differences in their expression, biosynthesis, and trafficking between wild-type and CatE(-/-) macrophages, the degradation rates of these two membrane proteins were apparently decreased as a result of cathepsin E deficiency. Because there was no difference in the vacuolar-type H+ -ATPase activity in both cell types, the elevated lysosomal pH in CatE(-/-) macrophages is most likely due to the accumulation of these lysosomal membrane glycoproteins highly modified with acidic monosaccharides, thereby leading to the disruption of non-proton factors controlling lysosomal pH. Furthermore, the selective degradation of LAMP-1 and LAMP-2, as well as LIMP-2, was also observed by treatment of the lysosomal membrane fraction isolated from wild-type macrophages with purified cathepsin E at pH 5. Our results thus suggest that cathepsin E is important for preventing the accumulation of these lysosomal membrane sialoglycoproteins that can induce a new form of lysosomal storage disorder.

DOI： 10.1074/jbc.M604143200

カテプシンE欠損はマクロファージの膜蛋白質輸送異常とミトコンドリア機能異常を起こす

筑波隆幸, 柳川三千代, 岡元邦彰, 岡本美子, 門脇知子, 山本健二

生化学 2007

Cathepsin E mediates membrane trafficking in macrophages

Takayuki Tsukuba, Shinya Yamamoto, Michiyo Yanagawa, Yoshiko Okamoto, Kuniaki Okamoto, Kenji Yamamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 103 214P - 214P 2007

カテプシンE欠損によるミトコンドリア機能異常と酸化ストレス亢進

筑波隆幸, 柳川三千代, 柳川三千代, 岡元邦彰, 門脇知子, 山本健二

Journal of Oral Biosciences 49 ( Supplement ) 2007

P.gingivalis感染とインスリン抵抗性に関する実験的解析

門脇知子, 瀧井良祐, 筑波隆幸, 岡元邦彰, 山本健二

Journal of Oral Biosciences 49 ( Supplement ) 2007

Pepstatin A, an aspartic proteinase inhibitor, suppresses RANKL-induced osteoclast differentiation. International journal

Hajime Yoshida, Kuniaki Okamoto, Tsutomu Iwamoto, Eiko Sakai, Kazuhiro Kanaoka, Jin-Ping Hu, Mitsue Shibata, Hitoshi Hotokezaka, Kazuhisa Nishishita, Akio Mizuno, Yuzo Kato

Journal of biochemistry 139 ( 3 ) 583 - 90 2006.3

Language：English

Pepstatin A is well known to be an inhibitor of aspartic proteinases such as pepsin, cathepsins D and E. Except for its role as a proteinase inhibitor, however, the pharmacological action of pepstatin A upon cells remain unclear. In this study, we found that pepstatin A suppressed receptor activator of NF-kappaB ligand (RANKL)-induced osteoclast differentiation. Pepstatin A suppressed the formation of multinuclear osteoclasts dose-dependently. This inhibition of the formation only affected osteoclast cells, i.e., not osteoblast-like cells. Furthermore, pepstatin A also suppressed differentiation from pre-osteoclast cells to mononuclear osteoclast cells dose-dependently. This inhibition seems to be independent of the activities of proteinases such as cathepsin D, because the formation of osteoclasts was not suppressed with the concentration that inhibited the activity of cathepsin D. Cell signaling analysis indicated that the phosphorylation of ERK was inhibited in pepstatin A-treated cells, while the phosphorylation of IkappaB and Akt showed almost no change. Furthermore, pepstatin A decreased the expression of nuclear factor of activated T cells c1 (NFATc1). These results suggest that pepstatin A suppresses the differentiation of osteoclasts through the blockade of ERK signaling and the inhibition of NFATc1 expression.

DOI： 10.1093/jb/mvj066

Characterization of rat cathepsin E and mutants with changed active-site residues and lacking propeptides and N-glycosylation, expressed in human embryonic kidney 293T cells

T Tsukuba, S Ikeda, K Okamoto, Y Yasuda, E Sakai, T Kadowaki, H Sakai, K Yamamoto

FEBS JOURNAL 273 ( 1 ) 219 - 229 2006.1

Language：English Publisher：WILEY-BLACKWELL

To study the roles of the catalytic activity, propeptide, and N-glycosylation of the intracellular aspartic proteinase cathepsin E in biosynthesis, processing, and intracellular trafficking, we constructed various rat cathepsin E mutants in which active-site Asp residues were changed to Ala or which lacked propeptides and N-glycosylation. Wild-type cathepsin E expressed in human embryonic kidney 293T cells was mainly found in the LAMP-1-positive endosomal organelles, as determined by immunofluorescence microscopy. Consistently, pulse-chase analysis revealed that the initially synthesized pro-cathepsin E was processed to the mature enzyme within a 24 h chase. This process was completely inhibited by brefeldin A and bafilomycin A, indicating its transport from the endoplasmic reticulum (ER) to the endosomal acidic compartment. Mutants with Asp residues in the two active-site consensus motifs changed to Ala and lacking the propeptide (Leu23-Phe58) and the putative ER-retention sequence (Ser59-Asp98) were neither processed nor transported to the endosomal compartment. The mutant lacking the ER-retention sequence was rapidly degraded in the ER, indicating the importance of this sequence in correct folding. The single (N92Q or N324D) and double (N92Q/N324D) N-glycosylation-deficient mutants were neither processed into a mature form nor transported to the endosomal compartment, but were stably retained in the ER without degradation. These data indicate that the catalytic activity, propeptides, and N-glycosylation of this protein are all essential for its processing, maturation, and trafficking.

DOI： 10.1111/j.1742-4658.2005.05062.x

Gingipains inactivate a cell surface ligand on Porphyromonas gingivalis that induces TLR2- and TLR4-independent signaling

M Kishimoto, A Yoshimura, M Naito, K Okamoto, K Yamamoto, DT Golenbock, Y Hara, K Nakayama

MICROBIOLOGY AND IMMUNOLOGY 50 ( 4 ) 315 - 325 2006

Language：English Publisher：CENTER ACADEMIC PUBL JAPAN

Arginine-specific gingipain and lysine-specific gingipain are two major cysteine proteinases produced by Porphyromonas gingivalis. To clarify the role of gingipains in the interaction between P gingivalis and the innate immune system, CHO reporter cells expressing TLR2 or TLR4 were stimulated with wildtype or gingipain-deficient P gingivalis cells and activation of nuclear factor-kappa B in these cells was examined. While CHO/CD14 cells and 7.19 cells, an MD-2-defective mutant derived from CHO/CD14 cells, failed to respond to wild-type P gingivalis, they responded to gingipain-deficient P gingivalis. On the other hand, CHO/CD14/TLR2 cells responded to both wild-type and gingipain-deficient P gingivalis. These results suggested that gingipains have no effects on TLR2-dependent signaling from P gingivalis but have inhibitory effects on TLR2- and TLR4-independent signaling in CHO cells. Indeed, the activity of gingipain-deficient P gingivalis to induce the activation of 7.19 cells was diminished after treatment of the bacterial cells with gingipains. We next partially purified bacterial cell components activating 7.19 cells from gingipain-deficient P gingivalis. The activity of the partially purified components was diminished by treatment with heat or gingipains. It is also noteworthy that anti-CD14 mAb inhibited the activation of 7.19 cells induced by the partially purified components. These results indicated that the components of R gingivalis that were able to induce TLR2- and TLR4-independent signaling were inactivated by gingipains before being recognized by CD14. The inactivation of the components would be helpful for P gingivalis to escape from the innate immune system.

Berberine inhibits RANKL-, TNF alpha-, LPS- and PGN-induced mature osteoclasts survival

JP Hu, K Nishishita, E Sakai, K Okamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 100 268P - 268P 2006

Involvement of glycosphingolipids in LPS mediated survival of mature osteoclasts

E Sakai, S Fukumoto, K Nishishita, JP Hu, K Okamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 100 65P - 65P 2006

カテプシンE欠損マクロファージにおける膜タンパク質の輸送異常

筑波隆幸, 山本晋也, 山本晋也, 柳川三千代, 柳川三千代, 岡元邦彰, 門脇知子, 山本健二

Journal of Oral Biosciences 48 ( Supplement ) 2006

P.gingivalis感染の生活習慣病増悪機序について

門脇知子, 筑波隆幸, 瀧井良祐, 重松直樹, 岡元邦彰, 山本健二

Journal of Oral Biosciences 48 ( Supplement ) 2006

The role of the cathepsin E propeptide in correct folding, maturation and sorting to the endosome

Y Yasuda, T Tsukuba, K Okamoto, T Kadowaki, K Yamamoto

JOURNAL OF BIOCHEMISTRY 138 ( 5 ) 621 - 630 2005.11

Language：English Publisher：OXFORD UNIV PRESS

Cathepsin E (CE) is an endosomal aspartic proteinase of the A1 family that is highly homologous to the lysosomal aspartic proteinase cathepsin D (CD). Newly synthesized CE undergoes several proteolytic processing events to yield mature CE, from which the N-terminal propeptide usually comprising 39 amino acids is removed. To define the role of the propeptide of CE in its biosynthesis and processing, we constructed two fusion proteins using chimeric DNAs encoding the CE propeptide fused to the mature CD tagged with HA at the COOH terminus (termed ED-HA) and encoding the CD propeptide fused to the mature CE (termed DE). Pulse-chase analysis revealed that wild-type CE expressed in human embryonic kidney cells is autoproteolytically processed into mature CE within a 12-h chase, whereas the chimeric DE failed to be converted into mature CE even after a 24-h chase. The DE chimera was nevertheless capable of acid-dependent autoactivation in vitro to yield a catalytically active form, although its specificity constants (k(cat)/K(m)) were considerably high but less (35%) than those of the wild-type CE. By contrast, the chimeric ED-HA expressed in HeLa cells underwent neither processing into a catalytically active enzyme nor acid-dependent autoactivation in vitro. The ED-HA protein was less stable than wt-CD-HA, as determined on pulse-chase analysis and on trypsin digestion. These data indicate that the propeptide of CE is essential for the correct folding, maturation, and targeting of this protein to its final destination.

DOI： 10.1093/jb/mvi159

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

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

Language：English Publisher：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

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.

DOI： 10.1074/jbc.M413544200

Alteration of substance P levels in various regions of mouse brain caused by cathepsin E-deficiency

N Shigematsu, T Nishioku, T Tsukuba, T Fukuda, T Yamamoto, T Yamaguchi, S Kohsaka, K Okamoto, Y Okamoto, Y Tanaka, M Himeno, S Higuchi, K Yamamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 97 72P - 72P 2005

Up-regulation, Enhanced Maturation, and Secretion of Cathepsin E in Mouse Macrophages Treated with Interferon-γ or Lipopolysaccharide

Michiyo Yanagawa, Takayuki Tsukuba, Kuniaki Okamoto, Ryosuke Takii, Yoshihiro Terada, Tomoko Kadowaki, Kenji Yamamoto

journal of oral biosciences 48 ( 3 ) 218 - 225 2005

Language：English

Cathepsin E is an intracellular aspartic proteinase that is predominantly localized in the endosomal compartments of antigen presenting cells including macrophages. Here, we have investigated the expression of cathepsin E in mouse macrophages treated with interferon (IFN)-γ, lipopolysaccharide (LPS), interleukin (IL)-10, and IL-4. The mRNA levels of cathepsin E in macrophages stimulated with IFN-γ and LPS were increased, but conversely, that with IL-10 was decreased. However, upon stimulation with IFN-γ or LPS, the activity levels of cathepsin E in the activated cells were markedly decreased, but those in the culture media were increased. Immunoblot analysis revealed that procathepsin E in the cells was largely converted to the mature form in the cells upon stimulation with IFN-γ. In addition, the extracellular enzyme was reacted with antibodies to mature cathepsin E but not with antibodies for procathepsin E. By contrast, when macrophages were treated with IL-4, cathepsin E production was significantly decreased in both the cell and the medium. These results indicate that, upon stimulation with IFN-γ and LPS, cathepsin E is up-regulated in macrophages, which is accompanied by the enhanced maturation and secretion of the enzyme. Conversely, cathepsin E is down-regulated by treatment with IL-4 and IL-10. © 2006, Japanese Association for Oral Biology. All rights reserved.

DOI： 10.2330/joralbiosci.48.218

Porphyromonas gingivalisの産生するシステインプロテアーゼ(Kgp)の活性中心の決定

岡元邦彰, HU P J, 坂井詠子, 西下一久, 門脇知子, 山本健二, 中山浩次, 加藤有三

Journal of Oral Biosciences 47 ( Supplement ) 2005

Cathepsin E is important for endosomal/lysosomal systems in macrophages against a bacterial infection

T Tsukuba, M Yanagawa, Y Okamoto, K Okamoto, K Yanamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 97 78P - 78P 2005

Glycosphingolipids regulate osteoclastgenesis mediated by lipopolysaccharide

E Sakai, S Fukumoto, Y Narita, M Tanaka, M Tachi, H Hotokezaka, K Kanaoka, K Okamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 97 178P - 178P 2005

Roles of Arg- and Lys-gingipains in coaggregation of Porphyromonas gingivalis: identification of its responsible molecules in translation products of rgpA, kgp, and hagA genes

N Abe, A Baba, R Takii, K Nakayama, A Kamaguchi, Y Shibata, Y Abiko, K Okamoto, T Kadowaki, K Yamamoto

BIOLOGICAL CHEMISTRY 385 ( 11 ) 1041 - 1047 2004.11

Language：English Publisher：WALTER DE GRUYTER & CO

Arg- (Rgp) and Lys-gingipains (Kgp) are two individual cysteine proteinases produced by Porphyromonas gingivalis, an oral anaerobic bacterium, and are implicated as major virulence factors in a wide range of pathologies of adult periodontitis. Coaggregation of this bacterium with other oral bacteria is an initial and critical step in infectious processes, yet the factors and mechanisms responsible for this process remain elusive. Here we show that the initial translation products of the rgpA, kgp and hemagglutinin hagA genes are responsible for coaggregation of P gingivalis and that the proteolytic activity of Rgp and Kgp is indispensable in this process. The rgpA rgpB kgp- and rgpA kgp hagA-deficient triple mutants exhibited no coaggregation activity with Actinomyces viscosus, whereas the kgp-null and rgpA rgpB-deficient double mutants significantly retained this activity. Consistently, the combined action of Rgp- and Kgp-specific inhibitors strongly inhibited the coaggregation activity of the bacterium, although single use of Rgp- or Kgp-specific inhibitor significantly retained this activity. We also demonstrate that the 47- and 43-kDa proteins produced from the translation products of the rgpA, kgp, and hagA genes by proteolytic activity of both Rgp and Kgp are responsible for the coaggregation of P. gingivalis.

DOI： 10.1515/BC.2004.135

ベルベリンのマウス骨髄細胞からの破骨細胞形成への抑制作用

胡錦萍, 西下一久, 柴田光枝, 坂井詠子, 金岡和博, 吉田一, 岡元邦彰, 加藤有三

Journal of oral biosciences 46 ( 5 ) 463 - 463 2004.9

カテプシンEによる免疫アレルギー性疾患の制御機構

筑波隆幸, 柳川三千代, 甲村恵子, 梯裕恵, 岡元邦彰, 山本健二

Journal of oral biosciences 46 ( 5 ) 474 - 474 2004.9

KIAA0377遺伝子産物の破骨細胞様細胞形成におよぼす影響

西下一久, 石田豊, 東洋一, 岡元邦彰, 加藤有三

Journal of oral biosciences 46 ( 5 ) 465 - 465 2004.9

Laminin alpha 2 is essential for odontoblast differentiation regulating dentin sialoprotein expression

K Yuasa, S Fukumoto, Y Kamasaki, A Yamada, E Fukumoto, K Kanaoka, K Saito, H Harada, E Arikawa-Hirasawa, Y Miyagoe-Suzuki, S Takeda, K Okamoto, Y Kato, T Fujiwara

JOURNAL OF BIOLOGICAL CHEMISTRY 279 ( 11 ) 10286 - 10292 2004.3

Language：English Publisher：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Laminin alpha2 is subunit of laminin-2 (alpha2beta1gamma1), which is a major component of the muscle basement membrane. Although the laminin alpha2 chain is expressed in the early stage of dental mesenchyme development and localized in the tooth germ basement membrane, its expression pattern in the late stage of tooth germ development and molecular roles are not clearly understood. We analyzed the role of laminin alpha2 in tooth development by using targeted mice with a disrupted lama2 gene. Laminin alpha2 is expressed in dental mesenchymal cells, especially in odontoblasts and during the maturation stage of ameloblasts, but not in the pre-secretory or secretory stages of ameloblasts. Lama2 mutant mice have thin dentin and a widely opened dentinal tube, as compared with wildtype and heterozygote mice, which is similar to the phenotype of dentinogenesis imperfecta. During dentin formation, the expression of dentin sialoprotein, a marker of odontoblast differentiation, was found to be decreased in odontoblasts from mutant mice. Furthermore, in primary cultures of dental mesenchymal cells, dentin matrix protein, and dentin sialophosphoprotein, mRNA expression was increased in laminin-2 coated dishes but not in those coated with other matrices, fibronectin, or type I collagen. Our results suggest that laminin alpha2 is essential for odontoblast differentiation and regulates the expression of dentin matrix proteins.

DOI： 10.1074/jbc.M310013200

Cathepsin E deficiency and brain functions

N Shigematsu, T Nishioku, T Tsukuba, S Kohsaka, K Okamoto, Y Okamoto, T Yamamoto, M Himeno, K Yamamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 94 89P - 89P 2004

Glycosphingolipids regulate osteoclastgenesis mediated by receptor activator of NF-kB ligand and lipopolysaccharide

E Sakai, S Fukumoto, H Hotokezaka, E Fukumoto, M Shibata, K Kanaoka, T Iwamoto, K Okamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 94 184P - 184P 2004

Effect of Pepstatin A in osteoclast

H Yoshida, K Okamoto, M Shibata, E Sakai, K Kanaoka, K Yuasa, A Mizuno, K Nishishita, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 94 188P - 188P 2004

Association of cathepsin E deficiency with development of atopic dermatitis

T Tsukuba, K Okamoto, Y Okamoto, M Yanagawa, K Kohmura, Y Yasuda, H Uchi, T Nakahara, M Furue, K Nakayama, T Kadowaki, K Yamamoto, KI Nakayama

JOURNAL OF BIOCHEMISTRY 134 ( 6 ) 893 - 902 2003.12

Language：English Publisher：OXFORD UNIV PRESS

Atopic dermatitis (AD) is a pruritic inflammatory skin diseases associated with a family history of atropy. Here we show that mice lacking the endolysosomal. aspartic proteinase cathepsin E spontaneously develop skin lesions similar to those of humans with AD when reared under conventional conditions but not under specific pathogen-free conditions. These mice showed the increase in the ratio of CD4(+)/CD8(+) T cells, the strong polarization of naive T cells to T helper 2 cells, and the systemic accumulation of IL-18 and IL-1beta accompanied by a marked increase in IL-4, IL-5, and IgE. The relative rates of degradation of IL-18 and IL-1beta were significantly lower in cathepsin E-deficient mice than wild-type mice. These results strongly suggest that the development of AD in cathepsin E-deficient mice is initiated by systemic accumulation of IL-18 and IL-1beta, mainly due to their reduced turnover rates. In addition, the reduced expression of cathepsin E was also observed in erythrocytes of both humans with AD and the AD mouse model NC/Nga. Cathepsin E deficiency might thus be responsible for the induction of AD in humans and mice.

DOI： 10.1093/jh/mvg216

ベルベリンによる破骨細胞分化因子(RANKL)細胞内シグナリング抑制効果

胡錦萍, 西下一久, 柴田光枝, 岡元邦彰, 加藤有三

歯科基礎医学会雑誌 45 ( 5 ) 374 - 374 2003.9

スフィンゴ糖脂質はRANKやc-Srcのラフトへの局在と破骨細胞形成を調節する

坂井詠子, 福本敏, 金岡和博, 柴田光枝, 福本恵美子, 岩本勉, 岡元邦彰, 加藤有三

歯科基礎医学会雑誌 45 ( 5 ) 343 - 343 2003.9

ラミニンα2の歯の発生における役割

湯浅健司, 福本敏, 釜崎陽子, 山田亜矢, 福本恵美子, 斉藤幹, 岡元邦彰, 藤原卓, 加藤有三

歯科基礎医学会雑誌 45 ( 5 ) 290 - 290 2003.9

口腔癌における破骨細胞分化因子の役割

岩本勉, 福本敏, 柳本惣市, 吉田一, 岡元邦彰, 吉冨泉, 川崎五郎, 加藤有三, 水野明夫

歯科基礎医学会雑誌 45 ( 5 ) 319 - 319 2003.9

Regulation of osteoclast survival and activation by glycosphingolipids.

K. Kanaoka, S. Fukumoto, K. Yuasa, K. Okamoto, M. Shibata, E. Sakai, Y. Kato, T. Iwamoto, F. Hashimoto, N. Yoshida

JOURNAL OF DENTAL RESEARCH 82 B358 - B358 2003.6

Subcellular localization of the receptor activator of NF-KappaB and SRC kinase in detergent insoluble fractions of osteoclasts

E Sakai, S Fukumoto, K Kanaoka, M Shibata, T Iwamoto, E Fukumoto, K Okamoto, Y Kato

BONE 32 ( 5 ) S155 - S155 2003.5

The regulation of bone resorption in tooth formation and eruption processes in mouse alveolar crest devoid of cathepsin K

M Okaji, H Sakai, E Sakai, M Shibata, F Hashimoto, Y Kobayashi, N Yoshida, K Okamoto, K Yamamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 91 ( 4 ) 285 - 294 2003.4

Language：English Publisher：JAPANESE PHARMACOLOGICAL SOC

Osteoclastic bone resorption has recently been implicated in the tooth formation and eruption in alveolar bone. Cathepsin K (CK) is a cysteine proteinase expressed predominantly in osteoclasts and is believed to play a critical role in degradation of bone matrix proteins. Here we present evidence that the alveolar bone resorption is essential for the tooth formation and that eruption proceeds normally in CK-deficient (CK-/-) mice. Radiographic and histological analyses revealed that the alveolar bone from these animals had no significant abnormalities during the tooth development between 5 and 28 days after birth. The tooth crown was normally erupted through the alveolar bone layer at 28 days after birth. The number of tartrate-resistant acid phosphatase-positive multinuclear cells in the alveolar bone around the tooth germ was apparently increased in 5-day-old CK-/- mice compared with age-matched littermates. More important, however, the immunohistochemical localization of matrix metalloproteinase-9 (MMP-9) was clearly increased in the CK-/- osteoclasts. In contrast, no significant difference in the immunoreactivity for cathepsin D was observed between the CK-/- osteoclasts and the wild-type ones. These results indicate that CK-/- osteoclasts are fully differentiated and are capable of degrading the organic phase of alveolar bone during the tooth formation and eruption, which may result from the compensatory action by MMP-9 increasingly expressed in the osteoclasts.

DOI： 10.1254/jphs.91.285

Disruption of structural and functional integrity of alpha(2)-macroglobulin by cathepsin E

M Shibata, H Sakai, E Sakai, K Okamoto, K Nishishita, Y Yasuda, Y Kato, K Yamamoto

EUROPEAN JOURNAL OF BIOCHEMISTRY 270 ( 6 ) 1189 - 1198 2003.3

Language：English Publisher：BLACKWELL PUBLISHING LTD

alpha(2) -Macroglobulin (alpha2M) is an abundant glycoprotein with the intrinsic capacity for capturing diverse proteins for rapid delivery into cells. After internalization by the receptor- mediated endocytosis, alpha2M-protein complexes were rapidly degraded in the endolysosome system. Although this is an important pathway for clearance of both alpha2M and biological targets, little is known about the nature of alpha2M degradation in the endolysosome system. To investigate the possible involvement of intracellular aspartic proteinases in the disruption of structural and functional integrity of alpha2M in the endolysosome system, we examined the capacity of alpha2M for interacting with cathepsin E and cathepsin D under acidic conditions and the nature of its degradation. alpha2M was efficiently associated with cathepsin E under acidic conditions to form noncovalent complexes and rapidly degraded through the generation of three major proteins with apparent molecular masses of 90, 85 and 30 kDa. Parallel with this reaction, alpha2M resulted in the rapid loss of its antiproteolytic activity. Analysis of the N-terminal amino-acid sequences of these proteins revealed that alpha2M was selectively cleaved at the Phe811-Leu812 bond in about 100mer downstream of the bait region. In contrast, little change was observed for alpha2M treated by cathepsin D under the same conditions. Together, the synthetic SPAFLA peptide corresponding to the Ser808-Ala813 sequence of human alpha2M, which contains the cathepsin E-cleavage site, was selectively cleaved by cathepsin E, but not cathepsin D. These results suggest the possible involvement of cathepsin E in disruption of the structural and functional integrity of alpha2M in the endolysosome system.

DOI： 10.1046/j.1432-1033.2003.03479.x

Porphyromonas gingivalisの産生するリジン特異的システインプロテアーゼ発現系の確立

岡元邦彰, 胡錦へい, 柴田光枝, 坂井詠子, 門脇知子, 庄子幹郎, 山本健二, 中山浩次, 加藤有三

歯科基礎医学会雑誌 45 ( 5 ) 2003

Cathepsin E and atopic dermatitis.

T Tsukuba, K Okamoto, K Yamamoto

JOURNAL OF PHARMACOLOGICAL SCIENCES 91 13P - 13P 2003

Expression and subcellular localization of caveolin with Src kinase and RANK in osteoclasts.

E Sakai, S Fukumoto, K Kanaoka, M Shibata, T Iwamoto, E Fukumoto, K Okamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 91 71P - 71P 2003

Purification and characterization of mouse soluble RANK expressed in a baculovirus system.

T Imai, M Shibata, K Kanaoka, K Yuasa, A Mizuno, K Okamoto, Y Kato

JOURNAL OF PHARMACOLOGICAL SCIENCES 91 239P - 239P 2003

なぜ,カテプシンK欠損マウスにおいて歯は正常に萌出できるのか?

岡地雅代, 柴田光枝, 坂井詠子, 岡元邦彰, 山本健二

歯科基礎医学会雑誌 44 ( 5 ) 433 - 433 2002.9

Porphyromonas gingivalisによる赤血球凝集機構解明の試み

坂井詠子, 佛坂斉祉, 庄子幹郎, 鎌口有秀, 岡元邦彰, 加藤有三, 中山浩次

歯科基礎医学会雑誌 44 ( 5 ) 466 - 466 2002.9

The role of propeptide of cathepsin E in its biosynthetic pathway

Y Yasuda, K Kohmura, K Okamoto, T Tsukuba, K Yamamoto

JAPANESE JOURNAL OF PHARMACOLOGY 88 183P - 183P 2002

カテプシンD欠損マウスにおける長骨骨端線の消失

岡元邦彰, 前田英史, 加藤有三, 山本健二

歯科基礎医学会雑誌 43 ( 5 ) 581 - 581 2001.8

歯周病原性細菌の蛋白質分解酵素を介する生存戦略 (新世紀における蛋白質科学の進展) -- (第3部生命現象の理解と医療・創薬に向けて)

山本健二, 馬場貴代, 岡元邦彰

蛋白質核酸酵素 46 ( 11 ) 1426 - 1427,1781〜1788 2001.8

Pathophysiological roles of two types of gingipains in periodontal diseases

K. Yamamoto, A. Baba, K. Okamoto, T. Kadowaki

Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme 46 ( 11 Suppl ) 1781 - 1788 2001.8

New functional aspects of cathepsin D and cathepsin E

T Tsukuba, K Okamoto, Y Yasuda, W Morikawa, H Nakanishi, K Yamamoto

MOLECULES AND CELLS 10 ( 6 ) 601 - 611 2000.12

Language：English Publishing type：Book review, literature introduction, etc. Publisher：SPRINGER-VERLAG SINGAPORE PTE LTD

Cathepsin D (CD) and cathepsin E are representative lysosomal and nonlysosomal aspartic proteinases, respectively, and play an important role in the degradation of proteins, the generation of bioactive proteins, antigen processing, etc. Recently, several lines of evidence have suggested the involvement of these two enzymes in the execution of neuronal death pathways induced by aging, transient forebrain ischemia, and excessive stimulation of glutamate receptors with excitotoxins. CD has also been shown to mediate apoptosis induced by various stimuli and p53-dependent tumor suppression. To gain more insight into in vivo functions of CD, mice deficient in this enzyme were generated. The mutant animals showed a progressive atrophy of the intestinal mucosa, a massive destruction of lymphoid organs, and a profound accumulation of ceroid lipofuscin, and developed a phenotype resembling neuronal ceroid lipofucinosis, suggesting that CD is essential for proteolysis of proteins regulating cell growth and tissue homeostasis. It has also been shown that CD molecules secreted from human prostate carcinoma cells are responsible for the generation of angiostatin, a potent endogenous inhibitor of angiogenesis, suggesting its contribution to the prevention of tumor growth and angiogenesis-dependent growth of metastases. Interestingly, pro-CD from human breast carcinoma cells showed a significantly lower angiostatin-generating activity than that from prostate carcinoma cells. Since deglycosylated CD molecules from both carcinoma cells showed a low angiostatin-generating activity, this discrepancy appeared to be attributed to the difference in the carbohydrate structures of CD molecules between the two cell types and to contribute to their potency to prevent tumor growth and metastases.

Porphyromonas gingivalis proteinases as virulence determinants in progression of periodontal diseases

Tomoko Kadowaki, Koji Nakayama, Kuniaki Okamoto, Naoko Abe, Atsuyo Baba, Yixin Shi, Dinath B. Ratnayake, Kenji Yamamoto

Journal of Biochemistry 128 ( 2 ) 153 - 159 2000.8

Language：English Publishing type：Book review, literature introduction, etc.

Porphyromonas gingivalis, one of the major causative agents of periodontal diseases, produces large amounts of arginine- and lysine-specific cysteine proteinases in cell-associated and secretory forms, which are now referred to as Arg-gingipain (Rgp) and Lysgingipain (Kgp), respectively. A number of studies have revealed that these proteinases are closely associated with the periodontopathogenesis of this bacterium: destruction of periodontal connective tissues, disruption of host defense mechanisms, and development and maintenance of inflammation in periodontal pockets. With respect to the physiology of the bacterium, Rgp and Kgp are indispensable for it to obtain nutrients from the environment, since it cannot utilize saccharides as carbon/energy sources for growth and totally depends on peptides and amino acids that are provided from environmental proteins by Rgp and Kgp. Furthermore, proteolytic activities of Rgp and Kgp contribute to processing/maturation of various cell-surface proteins of P. gingivalis, such as fimA fimbrilin (a subunit of major fimbriae), 75-kDa protein (a subunit of minor fimbriae), hemagglutinins, and the hemoglobin receptor protein, which are important for the bacterium to colonize and proliferate in the gingival crevice and to invade the periodontium. These findings strongly indicate critical roles of Rgp and Kgp in the virulence of P. gingivalis.

DOI： 10.1093/oxfordjournals.jbchem.a022735

歯周病原性細菌の産生するリジン特異的システインプロテアーゼの分布と機能解析

安部直子, 門脇知子, 岡元邦彰, 岡崎真治, 浅尾哲次, 中山浩次, 山本健二

歯科基礎医学会雑誌 42 ( 5 ) 2000

歯周病原性細菌のヘムおよびアミノ酸獲得機構

山本健二, 岡元邦彰, 門脇知子, 安部直子, 馬場貴代, 中山浩次

生化学 72 ( 8 ) 2000

カテプシンDおよびEの性状解析に有用な新しい合成基質の作製

安田善之, 岡元邦彰, 岡本美子, 景山節, 内山安男, 木南英紀, 山本健二

生化学 71 ( 8 ) 867 - 867 1999.8

Arginine- and lysine-specific gingipains from Porphyromonas gingivalis as major virulence factors of progressive periodontal disease.

山本健二, 門脇知子, 岡元邦彰, 安部直子, 中山浩次

炎症 18 ( 4 ) 259 - 264 1998

Structural and functional characterization of periodontal pathogenic cysteine proteinases from Porphyromonas gingivalis

K. Yamamoto, T. Kadowaki, K. Okamoto, K. Nakayama

Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme 42 ( 14 Suppl ) 2425 - 2432 1997.10

Biological roles of a novel class of cysteine proteinases from Porphyromonas gingivalis in periodontal disease progression

K Yamamoto, T Kadowaki, K Okamoto, N Abe, K Nakayama

MEDICAL ASPECTS OF PROTEASES AND PROTEASE INHIBITORS 15 139 - 149 1997

Language：English Publisher：I O S PRESS

We have studied the role of a novel class of cysteine proteinases from the oral anaerobic bacterium Porphyromonas gingivalis in periodontal disease progression. Two unique cysteine proteinases, so termed Arg-gingipain (RGP) and Lys-gingipain (KGP) due to their peptide cleavage specificity at arginine and lysine residues, respectively, have been implicated as major virulence factors in periodontal disease. Both enzymes are shown to be involved in the direct destruction of periodontal tissues and the disruption of normal host defense mechanisms. These enzymes have also been implicated in the hemagglutinating activity of the organism. To determine the structuraI, genetic, and functional relationships between the two enzymes, we have cloned the RGP- and KGP-encoding genes and ultimately constructed the rgp- or kgp-deficient mutants from the organism. In the course of this study, RGP has been shown to be encoded by two separate genes (rgpA and rgpB) on the chromosome of the organism. Deduced amino acid sequences of both RGP- and KGP-encoding genes revealed that the precursors of both enzymes were composed of at least four domains: the signal peptide, the NH2-terminal prosequence, the proteinase domain, and the COOH-terminal hemagglutinin domain. Although both enzymes were apparently different in the amino acid sequences of the NH2-terminal prosequences and the proteinase domains, their COOH-terminal hemagglutinin domains exhibited significant similarity. Analysis of the rgpA rgpB double mutant confirmed that RGP was a major virulence factor of the organism. However, the disruption of the KGP gene did not affect either the bactericidal activity of polymorphonucler cells or the hemagglutinin activity of the organism.

ラットカテプシンE分子のN-結合型糖鎖の役割

池田仁子, 西下一久, 坂井英昭, 岡元邦彰, 筑波隆幸, 加藤有三, 山本健二

日本分子生物学会年会プログラム・講演要旨集 19 255 - 255 1996.8

Processing of exine protein and secretion protein by Porphyromonas gingivalis arginine peculiar cysteine proteinase (Arg-gingipain).

門脇知子, 中山浩次, 吉村文信, 岡元邦彰, 山本健二

日本分子生物学会年会プログラム・講演要旨集 19th 1996

Periodontal disease and pathogenic protease.

山本健二, 門脇知子, 岡元邦彰

モダンフィジシャン 16 ( 12 ) 1996

Structural comparison of the pathogenicity protease gene of Porphyromonas gingivalis.

岡元邦彰, 門脇知子, 中山浩次, 山本健二

日本分子生物学会年会プログラム・講演要旨集 19th 1996

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