Updated on 2024/05/13

写真a

 
EGUCHI Takanori
 
Organization
Faculty of Medicine, Dentistry and Pharmaceutical Sciences Associate Professor
Position
Associate Professor
Contact information
メールアドレス
Other name(s)
Taka
Profile

Dr. Takanori Eguchi is a medical biologist, a faculty member at Okayama University, and a dentist. His motto is creativity, problem-solving, understanding of principles, and collaboration. Recently, he has been working on extracellular vesicle, including exosome medicine, dentistry, and biology.

Biography: Graduated Okayama University Dental School, DDS. Completed the graduate course at Okayama University, Ph. D. in Biochemistry, Molecular Biology, and Dentistry; Postdoctoral Research Fellow of JSPS (the Japan Society for the Promotion of Science); Section Chief at NCGG (National Center of Geriatrics and Gerontology), the National Institute for Longevity Sciences; Research Fellow and Instructor at Harvard Medical School (HMS) and Beth Israel Deconess Medical Center (BIDMC) before assuming his current position.

Awards: Japanese Society of Cartilage Metabolism Award, International CCN Society Springer Scholarship, Okayama Dental Society Outstanding Paper Award, etc.

He has published more than 90 papers and publications in cancer research and bone metabolism research with his expertise in exosomes and other vesicle particle science, intercellular communication, cellular molecular biology, regulation of gene expression, and cellular stress response. He has published papers in Seminars in Cancer Biology and the Journal of Extracellular Vesicles (see below for representative papers) and continues to pursue higher-quality research.

His representative works are;

Sheta M et al, Eguchi T*. Extracellular vesicles: new classification and tumor immunosuppression. Biology (Basel) 2023, 12(1):110. Top 1% Article.

Ono K et al, Eguchi T*. Triple Knockdown of CDC37, HSP90-alpha and HSP90-beta Diminishes Extracellular Vesicles-Driven Malignancy Events and Macrophage M2 Polarization in Oral Cancer. Journal of Extracellular Vesicles 2020, 9(1):1769373. Top 2% Article.

Taha EA, Ono K, Eguchi T*. Roles of Extracellular HSPs as Biomarkers in Immune Surveillance and Immune Evasion. International Journal of Molecular Sciences 2019, 20(18):4588. Top 2% Article, > 100 citations

Ono K, Eguchi T* et al. HSP-enriched properties of extracellular vesicles involve survival of metastatic oral cancer cells. Journal of Cellular Biochemistry 2018, 119(9):7363-7376. Top 2% Article,> 100 citations

Eguchi T* et al. Comprehensive Method for Exosome Isolation and Proteome Analysis for Detection of CCN Factors in/on Exosomes. In: Takigawa M (eds) CCN Proteins. Methods in Molecular Biology 2023, vol 2582. Humana, New York, NY. pp.59-76. Top 2% Article

Eguchi T* et al. Cancer Extracellular Vesicles, Tumoroid Models and Tumor Microenvironment. Seminars in Cancer Biology 2022,86(Pt 1):112-126. Top 3% Article.

Eguchi T* et al. Organoids with Cancer Stem Cell-like Properties Secrete Exosomes and HSP90 in a 3D Nanoenvironment. PLoS One 2018, 13(2):1-34. Top 3% Article,> 100 citations

Eguchi T* et al. OstemiR: A Novel Panel of microRNA Biomarkers in Osteoblastic and Osteocytic Differentiation from Mesenchymal Stem Cells. PLoS One 2013, 8(3):e58796. Top 3% Article, > 100 citations

Eguchi T et al. Novel Transcription-Factor-Like Function of Human Matrix Metalloproteinase 3 Regulating the CTGF/CCN2 Gene. Molecular and Cellular Biology 2008, 28(7): 2391-2413. Top 3% Article,  > 200 citations

Dr. Eguchi is also active in collaborative research and has a track record of various joint research projects within the university, in Japan, and overseas.

Oo MW, Kawai H, et al. Resident Stroma-secreted Chemokine CCL2 Governs Myeloid-derived Suppressor Cells in the Tumor Microenvironment. JCI Insight 2022, 7(1):e148960. Top 4% Article

Seyama M, Yoshida K, et al. Outer membrane vesicles of Porphyromonas gingivalis attenuate insulin sensitivity by delivering gingipains to the liver. Biochimica et Biophysica Acta - Molecular Basis of Disease 2020, 1866(6):165731. Top 4% Article

 

In terms of education, the department supervises graduate students, international students, dental students, and post-doctoral fellows, leading them to obtain degrees in medical and dental pharmacology, extracellular vesicles, molecular and cellular biology, biochemistry, pharmacology, etc. In addition to training many dentists, the department has also connected them to career paths at Harvard University, Kyoto University, Peking University, University of Texas, Fred Hutchinson Cancer Research Center, and others. He has also connected his career path to Harvard University, Kyoto University, Peking University, the University of Texas, Fred Hutchinson Cancer Research Center, etc.

Graduate students, technical staff, postdocs, joint research and development, and research funding are very welcome.

External link

Degree

  • PhD ( 2003 )

  • DDS ( 1999 )

Research Interests

  • Intercellular Communication

  • Extracellular Vesicles

  • Tumoroids

  • 3D culture systems

  • Organoid

  • Extracellular Particle

  • Exosome

  • Non-coding RNA

  • Gene expression control

  • Cancers

  • Omics

  • Cellular and Molecular Biology

  • Moonlighting Proteins

  • RNA

  • Protein

Research Areas

  • Life Science / Oral biological science

  • Life Science / Oral pathobiological science

  • Life Science / Tumor biology

  • Life Science / Cell biology

  • Life Science / Oral pathobiological science

  • Life Science / Tumor biology

  • Life Science / Nutrition science and health science

  • Life Science / Hygiene and public health (laboratory)

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Education

  • Okayama University   大学院医歯薬学総合研究科  

    1999.4 - 2003.3

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    Notes: 博士課程

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  • Okayama University   歯学部  

    1993.4 - 1999.3

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  • 広島大学附属福山高校    

    1990.4 - 1993.3

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Research History

  • Harvard Medical School   Instructor

    2011.9 - 2015.12

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  • Japan Society for Promotion of Science

    2003.4 - 2006.3

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  • Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences   Associate Professor   D.D.S., Ph.D.

    2021.11 - 2024.3

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  • Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences   Assistant Professor   D.D.S., Ph.D.

    2016.4 - 2021.10

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  • Harvard Medical School   Research fellow

    2011.7 - 2011.8

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  • National Center for Geriatrics and Gerontrogy   Section Chief

    2009.1 - 2011.6

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  • 国立長寿医療センター研究所   流動研究員

    2007.9 - 2008.12

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Professional Memberships

  • Japanese Society for Extracellular Vesicles (JSEV)

    2017

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  • THE JAPANESE CANCER ASSOCIATION

    2016

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  • International Society for Extracellular Vesicles (ISEV)

    2019 - 2022

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  • THE JAPANESE PHARMACOLOGICAL SOCIETY

    2017.1 - 2020.12

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  • BioMedical Society of Stress Response (BSSR) Japan

    2016

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  • American Society for Biochemistry and Molecular Biology

    2013 - 2016

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  • 歯科基礎医学会

    2001

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  • Japanese Society of Cartilage Metabolism

    2001 - 2010

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  • Japanese Biochemical Society

    2001 - 2008

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  • アメリカ骨代謝学会

    2001 - 2007

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  • The Japanese Society for Bone and Mineral Research

    2001 - 2005

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  • Okayama Dental Society

    2000

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  • The Molecular Biology Society of Japan

    2000 - 2019

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  • International CCN Society

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  • エピジェネティクス研究会

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  • 日本CCN研究会

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  • 日本補綴歯科学会

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  • 日本歯周病学会

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Committee Memberships

  • 日本臨床ストレス応答学会   若手奨励賞審査委員  

    2022   

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    Committee type:Academic society

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  • 日本臨床ストレス応答学会   広報  

    2022   

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  • 岡山大学歯学部   学校推薦型選抜Ⅱ及びバカロレア選抜選考委員  

    2022   

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  • 岡山大学歯学部   公募問題(CBT)作成部会委員  

    2021.4   

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  • 岡山大学歯学部   国家試験対策委員  

    2021.3   

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  • 日本臨床ストレス応答学会   幹事  

    2020.4   

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    Committee type:Academic society

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  • 岡山大学歯学部   OSCE部会委員  

    2018.4   

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  • 岡山大学歯学部   ARCOCS委員  

    2017.4   

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  • 岡山大学歯学部   早期見学実習検討部会委員  

    2016.4   

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  • 岡山大学歯学部   アクティブラーニング検討部会委員  

    2016.4 - 2018.3   

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  • 岡山大学歯学部   国際担当部会委員  

    2016.4   

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  • 日本臨床ストレス応答学会   評議員  

    2016   

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    Committee type:Academic society

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Papers

  • Exosome Invited Reviewed

    Takanori Eguchi

    Journal of Okayama Medical Association   136 ( 1 )   33 - 36   2024.1

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    Authorship:Lead author, Last author, Corresponding author   Language:Japanese   Publishing type:Research paper (scientific journal)  

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  • Extracellular Vesicles: New Classification and Tumor Immunosuppression. Invited Reviewed International journal

    Mona Sheta, Eman A Taha, Yanyin Lu, Takanori Eguchi

    Biology   12 ( 1 )   2023.1

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

    Extracellular vesicles (EVs) are cell-derived membrane-surrounded vesicles carrying various types of molecules. These EV cargoes are often used as pathophysiological biomarkers and delivered to recipient cells whose fates are often altered in local and distant tissues. Classical EVs are exosomes, microvesicles, and apoptotic bodies, while recent studies discovered autophagic EVs, stressed EVs, and matrix vesicles. Here, we classify classical and new EVs and non-EV nanoparticles. We also review EVs-mediated intercellular communication between cancer cells and various types of tumor-associated cells, such as cancer-associated fibroblasts, adipocytes, blood vessels, lymphatic vessels, and immune cells. Of note, cancer EVs play crucial roles in immunosuppression, immune evasion, and immunotherapy resistance. Thus, cancer EVs change hot tumors into cold ones. Moreover, cancer EVs affect nonimmune cells to promote cellular transformation, including epithelial-to-mesenchymal transition (EMT), chemoresistance, tumor matrix production, destruction of biological barriers, angiogenesis, lymphangiogenesis, and metastatic niche formation.

    File: 2023 01 Sheta et al, Eguchi, Biology EVs Review ★.pdf

    DOI: 10.3390/biology12010110

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  • Minimal information for studies of extracellular vesicles (MISEV2023): from basic to advanced approaches Invited Reviewed

    Journal of Extracellular Vesicles   2023

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  • SCAND1 Reverses Epithelial-to-Mesenchymal Transition (EMT) and Suppresses Prostate Cancer Growth and Migration Reviewed International journal

    Takanori Eguchi, Eva Csizmadia, Hotaka Kawai, Mona Sheta, Kunihiro Yoshida, Thomas L. Prince, Barbara Wegiel, Stuart K. Calderwood

    Cells   11 ( 24 )   3993   2022.12

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    Authorship:Lead author, Corresponding author   Language:English   Publishing type:Research paper (scientific journal)  

    Epithelial-mesenchymal transition (EMT) is a reversible cellular program that transiently places epithelial (E) cells into pseudo-mesenchymal (M) cell states. The malignant progression and resistance of many carcinomas depend on EMT activation, partial EMT, or hybrid E/M status in neoplastic cells. EMT is activated by tumor microenvironmental TGFβ signal and EMT-inducing transcription factors, such as ZEB1/2, in tumor cells. However, reverse EMT factors are less studied. We demonstrate that prostate epithelial transcription factor SCAND1 can reverse the cancer cell mesenchymal and hybrid E/M phenotypes to a more epithelial, less invasive status and inhibit their proliferation and migration in DU-145 prostate cancer cells. SCAND1 is a SCAN domain-containing protein and hetero-oligomerizes with SCAN-zinc finger transcription factors, such as MZF1, for accessing DNA and the transcriptional co-repression of target genes. We found that SCAND1 expression correlated with maintaining epithelial features, whereas the loss of SCAND1 was associated with mesenchymal phenotypes of tumor cells. SCAND1 and MZF1 were mutually inducible and coordinately included in chromatin with hetero-chromatin protein HP1γ. The overexpression of SCAND1 reversed hybrid E/M status into an epithelial phenotype with E-cadherin and β-catenin relocation. Consistently, the co-expression analysis in TCGA PanCancer Atlas revealed that SCAND1 and MZF1 expression was negatively correlated with EMT driver genes, including CTNNB1, ZEB1, ZEB2 and TGFBRs, in prostate adenocarcinoma specimens. In addition, SCAND1 overexpression suppressed tumor cell proliferation by reducing the MAP3K-MEK-ERK signaling pathway. Of note, in a mouse tumor xenograft model, SCAND1 overexpression significantly reduced Ki-67(+) and Vimentin(+) tumor cells and inhibited migration and lymph node metastasis of prostate cancer. Kaplan-Meier analysis showed high expression of SCAND1 and MZF1 to correlate with better prognoses in pancreatic cancer and head and neck cancers, although with poorer prognosis in kidney cancer. Overall, these data suggest that SCAND1 induces expression and coordinated heterochromatin-binding of MZF1 to reverse the hybrid E/M status into an epithelial phenotype and, inhibits tumor cell proliferation, migration, and metastasis, potentially by repressing the gene expression of EMT drivers and the MAP3K-MEK-ERK signaling pathway.

    File: cells-11-03993.pdf

    DOI: 10.3390/cells11243993

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  • Cancer Extracellular Vesicles, Tumoroid Models and Tumor Microenvironment Invited Reviewed International journal

    Takanori Eguchi, Mona Sheta, Masanori Fujii, Stuart K. Calderwood

    Seminars in Cancer Biology   86 ( Pt 1 )   112 - 126   2022.1

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    Cancer extracellular vesicles (EVs), or exosomes, promote tumor progression through enhancing tumor growth, initiating epithelial-to-mesenchymal transition, remodeling the tumor microenvironment, and preparing metastatic niches. Three-dimensionally (3D) cultured tumoroids / spheroids aim to reproduce some aspects of tumor behavior in vitro and show increased cancer stem cell properties. These properties are transferred to their EVs that promote tumor growth. Moreover, recent tumoroid models can be furnished with aspects of the tumor microenvironment, such as vasculature, hypoxia, and extracellular matrix. This review summarizes tumor tissue culture and engineering platforms compatible with EV research. For example, the combination experiments of 3D-tumoroids and EVs have revealed multifunctional proteins loaded in EVs, such as metalloproteinases and heat shock proteins. EVs or exosomes are able to transfer their cargo molecules to recipient cells, whose fates are often largely altered. In addition, the review summarizes approaches to EV labeling technology using fluorescence and luciferase, useful for studies on EV-mediated intercellular communication, biodistribution, and metastatic niche formation.

    File: 2022 01 Eguchi Semin Cancer Biol.pdf

    DOI: 10.1016/j.semcancer.2022.01.003

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  • Triple knockdown of CDC37, HSP90-alpha and HSP90-beta diminishes extracellular vesicles-driven malignancy events and macrophage M2 polarization in oral cancer. Reviewed International journal

    Kisho Ono, Chiharu Sogawa, Hotaka Kawai, Manh Tien Tran, Eman A Taha, Yanyin Lu, May Wathone Oo, Yuka Okusha, Hirohiko Okamura, Soichiro Ibaragi, Masaharu Takigawa, Ken-Ichi Kozaki, Hitoshi Nagatsuka, Akira Sasaki, Kuniaki Okamoto, Stuart K Calderwood, Takanori Eguchi

    Journal of extracellular vesicles   9 ( 1 )   1769373 - 1769373   2020.5

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

    Evidence has been accumulating to indicate that extracellular vesicles (EVs), including exosomes, released by cancer cells can foster tumour progression. The molecular chaperones - CDC37, HSP90α and HSP90β play key roles in cancer progression including epithelial-mesenchymal transition (EMT), although their contribution to EVs-mediated cell-cell communication in tumour microenvironment has not been thoroughly examined. Here we show that triple depletion of the chaperone trio attenuates numerous cancer malignancy events exerted through EV release. Metastatic oral cancer-derived EVs (MEV) were enriched with HSP90α HSP90β and cancer-initiating cell marker CD326/EpCAM. Depletion of these chaperones individually induced compensatory increases in the other chaperones, whereas triple siRNA targeting of these molecules markedly diminished the levels of the chaperone trio and attenuated EMT. MEV were potent agents in initiating EMT in normal epithelial cells, a process that was attenuated by the triple chaperone depletion. The migration, invasion, and in vitro tumour initiation of oral cancer cells were significantly promoted by MEV, while triple depletion of CDC37/HSP90α/β reversed these MEV-driven malignancy events. In metastatic oral cancer patient-derived tumours, HSP90β was significantly accumulated in infiltrating tumour-associated macrophages (TAM) as compared to lower grade oral cancer cases. HSP90-enriched MEV-induced TAM polarization to an M2 phenotype, a transition known to support cancer progression, whereas the triple chaperone depletion attenuated this effect. Mechanistically, the triple chaperone depletion in metastatic oral cancer cells effectively reduced MEV transmission into macrophages. Hence, siRNA-mediated knockdown of the chaperone trio (CDC37/HSP90α/HSP90β) could potentially be a novel therapeutic strategy to attenuate several EV-driven malignancy events in the tumour microenvironment. Abbreviations: CDC37: cell division control 37; EMT: epithelial-mesenchymal transmission; EV: extracellular vesicles; HNSCC: head and neck squamous cell carcinoma; HSP90: heat shock protein 90; TAM: tumour-associated macrophage.

    File: 2020 06 Ono JEV ★.pdf

    DOI: 10.1080/20013078.2020.1769373

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  • Knockout of MMP3 Weakens Solid Tumor Organoids and Cancer Extracellular Vesicles. Reviewed International journal

    Eman A Taha, Chiharu Sogawa, Yuka Okusha, Hotaka Kawai, May Wathone Oo, Abdellatif Elseoudi, Yanyin Lu, Hitoshi Nagatsuka, Satoshi Kubota, Ayano Satoh, Kuniaki Okamoto, Takanori Eguchi

    Cancers   12 ( 5 )   1 - 29   2020.5

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    The tumor organoid (tumoroid) model in three-dimensional (3D) culture systems has been developed to reflect more closely the in vivo tumors than 2D-cultured tumor cells. Notably, extracellular vesicles (EVs) are efficiently collectible from the culture supernatant of gel-free tumoroids. Matrix metalloproteinase (MMP) 3 is a multi-functional factor playing crucial roles in tumor progression. However, roles of MMP3 within tumor growth and EVs have not unveiled. Here, we investigated the protumorigenic roles of MMP3 on integrities of tumoroids and EVs. We generated MMP3-knockout (KO) cells using the CRISPR/Cas9 system from rapidly metastatic LuM1 tumor cells. Moreover, we established fluorescent cell lines with palmitoylation signal-fused fluorescent proteins (tdTomato and enhanced GFP). Then we confirmed the exchange of EVs between cellular populations and tumoroids. LuM1-tumoroids released large EVs (200-1000 nm) and small EVs (50-200 nm) while the knockout of MMP3 resulted in the additional release of broken EVs from tumoroids. The loss of MMP3 led to a significant reduction in tumoroid size and the development of the necrotic area within tumoroids. MMP3 and CD9 (a category-1 EV marker tetraspanin protein) were significantly down-regulated in MMP3-KO cells and their EV fraction. Moreover, CD63, another member of the tetraspanin family, was significantly reduced only in the EVs fractions of the MMP3-KO cells compared to their counterpart. These weakened phenotypes of MMP3-KO were markedly rescued by the addition of MMP3-rich EVs or conditioned medium (CM) collected from LuM1-tumoroids, which caused a dramatic rise in the expression of MMP3, CD9, and Ki-67 (a marker of proliferating cells) in the MMP3-null/CD9-low tumoroids. Notably, MMP3 enriched in tumoroids-derived EVs and CM deeply penetrated recipient MMP3-KO tumoroids, resulting in a remarkable enlargement of solid tumoroids, while MMP3-null EVs did not. These data demonstrate that EVs can mediate molecular transfer of MMP3, resulting in increasing the proliferation and tumorigenesis, indicating crucial roles of MMP3 in tumor progression.

    File: 2020 05 Taha cancers.pdf

    DOI: 10.3390/cancers12051260

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  • Extracellular Vesicles Enriched with Moonlighting Metalloproteinase Are Highly Transmissive, Pro-Tumorigenic, and Trans-Activates Cellular Communication Network Factor (CCN2/CTGF): CRISPR against Cancer. Reviewed International journal

    Yuka Okusha, Takanori Eguchi, Manh T Tran, Chiharu Sogawa, Kaya Yoshida, Mami Itagaki, Eman A Taha, Kisho Ono, Eriko Aoyama, Hirohiko Okamura, Ken-Ichi Kozaki, Stuart K Calderwood, Masaharu Takigawa, Kuniaki Okamoto

    Cancers   12 ( 4 )   1 - 27   2020.4

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)  

    Matrix metalloproteinase 3 (MMP3) plays multiple roles in extracellular proteolysis as well as intracellular transcription, prompting a new definition of moonlighting metalloproteinase (MMP), according to a definition of protein moonlighting (or gene sharing), a phenomenon by which a protein can perform more than one function. Indeed, connective tissue growth factor (CTGF, aka cellular communication network factor 2 (CCN2)) is transcriptionally induced as well as cleaved by MMP3. Moreover, several members of the MMP family have been found within tumor-derived extracellular vesicles (EVs). We here investigated the roles of MMP3-rich EVs in tumor progression, molecular transmission, and gene regulation. EVs derived from a rapidly metastatic cancer cell line (LuM1) were enriched in MMP3 and a C-terminal half fragment of CCN2/CTGF. MMP3-rich, LuM1-derived EVs were disseminated to multiple organs through body fluid and were pro-tumorigenic in an allograft mouse model, which prompted us to define LuM1-EVs as oncosomes in the present study. Oncosome-derived MMP3 was transferred into recipient cell nuclei and thereby trans-activated the CCN2/CTGF promoter, and induced CCN2/CTGF production in vitro. TRENDIC and other cis-elements in the CCN2/CTGF promoter were essential for the oncosomal responsivity. The CRISPR/Cas9-mediated knockout of MMP3 showed significant anti-tumor effects such as the inhibition of migration and invasion of tumor cells, and a reduction in CCN2/CTGF promoter activity and fragmentations in vitro. A high expression level of MMP3 or CCN2/CTGF mRNA was prognostic and unfavorable in particular types of cancers including head and neck, lung, pancreatic, cervical, stomach, and urothelial cancers. These data newly demonstrate that oncogenic EVs-derived MMP is a transmissive trans-activator for the cellular communication network gene and promotes tumorigenesis at distant sites.

    File: 2020 04 Okusha Cancers MMP3 KO.pdf

    DOI: 10.3390/cancers12040881

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  • Cell Stress Induced Stressome Release Including Damaged Membrane Vesicles and Extracellular HSP90 by Prostate Cancer Cells. Reviewed International journal

    Takanori Eguchi, Chiharu Sogawa, Kisho Ono, Masaki Matsumoto, Manh Tien Tran, Yuka Okusha, Benjamin J Lang, Kuniaki Okamoto, Stuart K Calderwood

    Cells   9 ( 3 )   1 - 24   2020.3

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    Tumor cells exhibit therapeutic stress resistance-associated secretory phenotype involving extracellular vesicles (EVs) such as oncosomes and heat shock proteins (HSPs). Such a secretory phenotype occurs in response to cell stress and cancer therapeutics. HSPs are stress-responsive molecular chaperones promoting proper protein folding, while also being released from cells with EVs as well as a soluble form known as alarmins. We have here investigated the secretory phenotype of castration-resistant prostate cancer (CRPC) cells using proteome analysis. We have also examined the roles of the key co-chaperone CDC37 in the release of EV proteins including CD9 and epithelial-to-mesenchymal transition (EMT), a key event in tumor progression. EVs derived from CRPC cells promoted EMT in normal prostate epithelial cells. Some HSP family members and their potential receptor CD91/LRP1 were enriched at high levels in CRPC cell-derived EVs among over 700 other protein types found by mass spectrometry. The small EVs (30-200 nm in size) were released even in a non-heated condition from the prostate cancer cells, whereas the EMT-coupled release of EVs (200-500 nm) and damaged membrane vesicles with associated HSP90α was increased after heat shock stress (HSS). GAPDH and lactate dehydrogenase, a marker of membrane leakage/damage, were also found in conditioned media upon HSS. During this stress response, the intracellular chaperone CDC37 was transcriptionally induced by heat shock factor 1 (HSF1), which activated the CDC37 core promoter, containing an interspecies conserved heat shock element. In contrast, knockdown of CDC37 decreased EMT-coupled release of CD9-containing vesicles. Triple siRNA targeting CDC37, HSP90α, and HSP90β was required for efficient reduction of this chaperone trio and to reduce tumorigenicity of the CRPC cells in vivo. Taken together, we define "stressome" as cellular stress-induced all secretion products, including EVs (200-500 nm), membrane-damaged vesicles and remnants, and extracellular HSP90 and GAPDH. Our data also indicated that CDC37 is crucial for the release of vesicular proteins and tumor progression in prostate cancer.

    File: 2020 03 Eguchi Cells Stressome★.pdf

    DOI: 10.3390/cells9030755

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  • Roles of Extracellular HSPs as Biomarkers in Immune Surveillance and Immune Evasion. Reviewed International journal

    Eman A Taha, Kisho Ono, Takanori Eguchi

    International journal of molecular sciences   20 ( 18 )   1 - 32   2019.9

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    Extracellular heat shock proteins (ex-HSPs) have been found in exosomes, oncosomes, membrane surfaces, as well as free HSP in cancer and various pathological conditions, also known as alarmins. Such ex-HSPs include HSP90 (α, β, Gp96, Trap1), HSP70, and large and small HSPs. Production of HSPs is coordinately induced by heat shock factor 1 (HSF1) and hypoxia-inducible factor 1 (HIF-1), while matrix metalloproteinase 3 (MMP-3) and heterochromatin protein 1 are novel inducers of HSPs. Oncosomes released by tumor cells are a major aspect of the resistance-associated secretory phenotype (RASP) by which immune evasion can be established. The concepts of RASP are: (i) releases of ex-HSP and HSP-rich oncosomes are essential in RASP, by which molecular co-transfer of HSPs with oncogenic factors to recipient cells can promote cancer progression and resistance against stresses such as hypoxia, radiation, drugs, and immune systems; (ii) RASP of tumor cells can eject anticancer drugs, targeted therapeutics, and immune checkpoint inhibitors with oncosomes; (iii) cytotoxic lipids can be also released from tumor cells as RASP. ex-HSP and membrane-surface HSP (mHSP) play immunostimulatory roles recognized by CD91+ scavenger receptor expressed by endothelial cells-1 (SREC-1)+ Toll-like receptors (TLRs)+ antigen-presenting cells, leading to antigen cross-presentation and T cell cross-priming, as well as by CD94+ natural killer cells, leading to tumor cytolysis. On the other hand, ex-HSP/CD91 signaling in cancer cells promotes cancer progression. HSPs in body fluids are potential biomarkers detectable by liquid biopsies in cancers and tissue-damaged diseases. HSP-based vaccines, inhibitors, and RNAi therapeutics are also reviewed.

    DOI: 10.3390/ijms20184588

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  • HSP-enriched properties of extracellular vesicles involve survival of metastatic oral cancer cells. Reviewed International journal

    Kisho Ono, Takanori Eguchi, Chiharu Sogawa, Stuart K Calderwood, Junya Futagawa, Tomonari Kasai, Masaharu Seno, Kuniaki Okamoto, Akira Sasaki, Ken-Ichi Kozaki

    Journal of cellular biochemistry   119 ( 9 )   7350 - 7362   2018.9

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)  

    Cancer cells often secrete extracellular vesicles (EVs) that carry heat shock proteins (HSPs) with roles in tumor progression. Oral squamous cell carcinoma (OSCC) belongs to head and neck cancers (HNC) whose lymph-node-metastases often lead to poor prognosis. We have examined the EV proteome of OSCC cells and found abundant secretion of HSP90-enriched EVs in lymph-node-metastatic OSCC cells. Double knockdown of HSP90α and HSP90β, using small interfering RNA significantly reduced the survival of the metastatic OSCC cells, although single knockdown of each HSP90 was ineffective. Elevated expression of these HSP90 family members was found to correlate with poor prognosis of HNC cases. Thus, elevated HSP90 levels in secreted vesicles are potential prognostic biomarkers and therapeutic targets in metastatic OSCC.

    DOI: 10.1002/jcb.27039

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  • Organoids with cancer stem cell-like properties secrete exosomes and HSP90 in a 3D nanoenvironment. Reviewed International journal

    Takanori Eguchi, Chiharu Sogawa, Yuka Okusha, Kenta Uchibe, Ryosuke Iinuma, Kisho Ono, Keisuke Nakano, Jun Murakami, Manabu Itoh, Kazuya Arai, Toshifumi Fujiwara, Yuri Namba, Yoshiki Murata, Kazumi Ohyama, Manami Shimomura, Hirohiko Okamura, Masaharu Takigawa, Tetsuya Nakatsura, Ken-Ichi Kozaki, Kuniaki Okamoto, Stuart K Calderwood

    PloS one   13 ( 2 )   e0191109 - 2332   2018

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    Ability to form cellular aggregations such as tumorspheres and spheroids have been used as a morphological marker of malignant cancer cells and in particular cancer stem cells (CSC). However, the common definition of the types of cellular aggregation formed by cancer cells has not been available. We examined morphologies of 67 cell lines cultured on three dimensional morphology enhancing NanoCulture Plates (NCP) and classified the types of cellular aggregates that form. Among the 67 cell lines, 49 cell lines formed spheres or spheroids, 8 cell lines formed grape-like aggregation (GLA), 8 cell lines formed other types of aggregation, and 3 cell lines formed monolayer sheets. Seven GLA-forming cell lines were derived from adenocarcinoma among the 8 lines. A neuroendocrine adenocarcinoma cell line PC-3 formed asymmetric GLA with ductal structures on the NCPs and rapidly growing asymmetric tumors that metastasized to lymph nodes in immunocompromised mice. In contrast, another adenocarcinoma cell line DU-145 formed spheroids in vitro and spheroid-like tumors in vivo that did not metastasize to lymph nodes until day 50 after transplantation. Culture in the 3D nanoenvironment and in a defined stem cell medium enabled the neuroendocrine adenocarcinoma cells to form slowly growing large organoids that expressed multiple stem cell markers, neuroendocrine markers, intercellular adhesion molecules, and oncogenes in vitro. In contrast, the more commonly used 2D serum-contained environment reduced intercellular adhesion and induced mesenchymal transition and promoted rapid growth of the cells. In addition, the 3D stemness nanoenvironment promoted secretion of HSP90 and EpCAM-exosomes, a marker of CSC phenotype, from the neuroendocrine organoids. These findings indicate that the NCP-based 3D environment enables cells to form stem cell tumoroids with multipotency and model more accurately the in vivo tumor status at the levels of morphology and gene expression.

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  • Intracellular MMP3 Promotes HSP Gene Expression in Collaboration With Chromobox Proteins. Reviewed International journal

    Takanori Eguchi, Stuart K Calderwood, Masaharu Takigawa, Satoshi Kubota, Ken-Ichi Kozaki

    Journal of cellular biochemistry   118 ( 1 )   43 - 51   2017.1

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    Matrix metalloproteinases (MMPs) are crucial factors in tumor progression, inflammatory/immune responses and tissue development/regeneration. Of note, it has been known that MMPs promote genome instability, epithelial-mesenchymal transition, invasion, and metastasis in tumor progression. We previously reported that human MMP3 could translocate into cellular nuclei and control transcription in human chondrosarcoma-derived cells and in articular cartilage (Eguchi et al. [2008] Mol Cell Biol 28(7):2391-2413); however, further transcriptional target genes and cofactors of intranuclear MMP3 have not been uncovered. In this paper, we used transcriptomics analysis in order to examine novel transcriptional target genes regulated by intracellular MMP3. We found that mRNA levels of HSP family members (HSP70B', HSP72, HSP40/DNAJ, and HSP20/CRYAB) are upregulated by the intracellular MMP3 overload. Bioinformatic analysis predicted several transcription factors that possibly interact with MMP3. Among these factors, heat shock factor 1 (HSF1) cooperated with the MMP3 to activate the HSP70B' gene promoter in reporter gene assays, while a dominant negative HSF1 blocked the role for MMP3 in the trans-activation. The hemopexin-like repeat (PEX) domain of the human MMP3 was essential for transcriptional induction of the HSP70B' gene. In addition, chromobox proteins CBX5/HP1α and CBX3/HP1γ cooperated with the PEX domain in induction of HSP70B' mRNA. Taken together, this study newly clarified that intracellular MMP3 cooperate with CBXs/HP1s in transcriptional promotion of HSP genes. J. Cell. Biochem. 118: 43-51, 2017. © 2016 Wiley Periodicals, Inc.

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  • OstemiR: a novel panel of microRNA biomarkers in osteoblastic and osteocytic differentiation from mesencymal stem cells. Reviewed International journal

    Takanori Eguchi, Ken Watanabe, Emilio Satoshi Hara, Mitsuaki Ono, Takuo Kuboki, Stuart K Calderwood

    PloS one   8 ( 3 )   e58796   2013

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    MicroRNAs (miRNAs) are small RNA molecules of 21-25 nucleotides that regulate cell behavior through inhibition of translation from mRNA to protein, promotion of mRNA degradation and control of gene transcription. In this study, we investigated the miRNA expression signatures of cell cultures undergoing osteoblastic and osteocytic differentiation from mesenchymal stem cells (MSC) using mouse MSC line KUSA-A1 and human MSCs. Ninety types of miRNA were quantified during osteoblastic/osteocytic differentiation in KUSA-A1 cells utilizing miRNA PCR arrays. Coincidently with mRNA induction of the osteoblastic and osteocytic markers, the expression levels of several dozen miRNAs including miR-30 family, let-7 family, miR-21, miR-16, miR-155, miR-322 and Snord85 were changed during the differentiation process. These miRNAs were predicted to recognize osteogenic differentiation-, stemness-, epinegetics-, and cell cycle-related mRNAs, and were thus designated OstemiR. Among those OstemiR, the miR-30 family was classified into miR-30b/c and miR-30a/d/e groups on the basis of expression patterns during osteogenesis as well as mature miRNA structures. In silico prediction and subsequent qRT-PCR in stable miR-30d transfectants clarified that context-dependent targeting of miR-30d on known regulators of bone formation including osteopontin/spp1, lifr, ccn2/ctgf, ccn1/cyr61, runx2, sox9 as well as novel key factors including lin28a, hnrnpa3, hspa5/grp78, eed and pcgf5. In addition, knockdown of human OstemiR miR-541 increased Osteopontin/SPP1 expression and calcification in hMSC osteoblastic differentiation, indicating that miR-541 is a negative regulator of osteoblastic differentiation. These observations indicate stage-specific roles of OstemiR especially miR-541 and the miR-30 family on novel targets in osteogenesis.

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  • Novel transcription-factor-like function of human matrix metalloproteinase 3 regulating the CTGF/CCN2 gene. Reviewed International journal

    Takanori Eguchi, Satoshi Kubota, Kazumi Kawata, Yoshiki Mukudai, Junji Uehara, Toshihiro Ohgawara, Soichiro Ibaragi, Akira Sasaki, Takuo Kuboki, Masaharu Takigawa

    Molecular and cellular biology   28 ( 7 )   2391 - 413   2008.4

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    Matrix metalloproteinase 3 (MMP3) is well known as a secretory endopeptidase that degrades extracellular matrices. Recent reports indicated the presence of MMPs in the nucleus (A. J. Kwon et al., FASEB J. 18:690-692, 2004); however, its function has not been well investigated. Here, we report a novel function of human nuclear MMP3 as a trans regulator of connective tissue growth factor (CCN2/CTGF). Initially, we cloned MMP3 cDNA as a DNA-binding factor for the CCN2/CTGF gene. An interaction between MMP3 and transcription enhancer dominant in chondrocytes (TRENDIC) in the CCN2/CTGF promoter was confirmed by a gel shift assay and chromatin immunoprecipitation. The CCN2/CTGF promoter was activated by overexpressed MMP3, whereas a TRENDIC mutant promoter lost the response. Also, the knocking down of MMP3 suppressed CCN2/CTGF expression. By cytochemical and histochemical analyses, MMP3 was detected in the nuclei of chondrocytic cells in culture and also in the nuclei of normal and osteoarthritic chondrocytes in vivo. The nuclear translocation of externally added recombinant MMP3 and six putative nuclear localization signals in MMP3 also were shown. Furthermore, we determined that heterochromatin protein gamma coordinately regulates CCN2/CTGF by interacting with MMP3. The involvement of this novel role of MMP3 in the development, tissue remodeling, and pathology of arthritic diseases through CCN2/CTGF regulation thus is suggested.

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  • Stress biology: Complexity and multifariousness in health and disease. International journal

    Matthias P Mayer, Laura Blair, Gregory L Blatch, Thiago J Borges, Ahmed Chadli, Gabriela Chiosis, Aurélie de Thonel, Albena Dinkova-Kostova, Heath Ecroyd, Adrienne L Edkins, Takanori Eguchi, Monika Fleshner, Kevin P Foley, Sotirios Fragkostefanakis, Jason Gestwicki, Pierre Goloubinoff, Jennifer A Heritz, Christine M Heske, Jonathan D Hibshman, Jenny Joutsen, Wei Li, Michael Lynes, Marc L Mendillo, Nahid Mivechi, Fortunate Mokoena, Yuka Okusha, Veena Prahlad, Elizabeth Repasky, Sara Sannino, Federica Scalia, Reut Shalgi, Lea Sistonen, Emily Sontag, Patricija van Oosten-Hawle, Anniina Vihervaara, Anushka Wickramaratne, Shawn Xiang Yang Wang, Tawanda Zininga

    Cell stress & chaperones   29 ( 1 )   143 - 157   2024.2

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    Preserving and regulating cellular homeostasis in the light of changing environmental conditions or developmental processes is of pivotal importance for single cellular and multicellular organisms alike. To counteract an imbalance in cellular homeostasis transcriptional programs evolved, called the heat shock response, unfolded protein response, and integrated stress response, that act cell-autonomously in most cells but in multicellular organisms are subjected to cell-nonautonomous regulation. These transcriptional programs downregulate the expression of most genes but increase the expression of heat shock genes, including genes encoding molecular chaperones and proteases, proteins involved in the repair of stress-induced damage to macromolecules and cellular structures. Sixty-one years after the discovery of the heat shock response by Ferruccio Ritossa, many aspects of stress biology are still enigmatic. Recent progress in the understanding of stress responses and molecular chaperones was reported at the 12th International Symposium on Heat Shock Proteins in Biology, Medicine and the Environment in the Old Town Alexandria, VA, USA from 28th to 31st of October 2023.

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  • Rab11 suppresses head and neck carcinoma by regulating EGFR and EpCAM exosome secretion. Reviewed International journal

    Kunihiro Yoshida, Kaung Htike, Takanori Eguchi, Hotaka Kawai, Htoo Shwe Eain, Manh Tien Tran, Chiharu Sogawa, Koki Umemori, Tatsuo Ogawa, Hideka Kanemoto, Kisho Ono, Hitoshi Nagatsuka, Akira Sasaki, Soichiro Ibaragi, Kuniaki Okamoto

    Journal of oral biosciences   66 ( 1 )   205 - 216   2023.12

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    OBJECTIVES: Rab11(Rab11a and Rab11b) localizes primarily along recycling endosomes in cells and is involved in various intracellular trafficking processes, including membrane receptor recycling and secretion of exosomes or small extracellular vesicles (EVs). Although Rab11 is closely associated with the progression and metastasis of various cancer types, little is known about Rab11' role in head and neck squamous cell carcinoma (HNSCC). In this study, we investigated the roles of Rab11a and Rab11b in HNSCC. METHODS: The clinical significance of Rab11 expression in HNSCC was investigated using a public database and tissue microarray analysis. Stable cell lines with loss and gain of Rab11a or Rab11b were originally established to investigate their roles in the proliferative, migratory, and invasive capabilities of HNSCC cells. RESULTS: Database analysis revealed a significant association between Rab11b mRNA expression and a favorable patient survival rate in HNSCC. Tissue microarray analysis revealed that Rab11b expression was the highest in normal tissues and gradually decreased across the stages of HNSCC progression. Overexpression of Rab11a or Rab11b resulted in a decrease in epidermal growth factor receptor (EGFR), Epithelial cell adhesion molecule (EpCAM) exosome secretion, and the migratory and invasive potential of HNSCC cells. The knockdown of Rab11a or Rab11b increased EpCAM/CD9 exosome secretion in addition to the migratory and invasive potential of HNSCC cells. CONCLUSIONS: Rab11 suppresses HNSCC by regulating EGFR recycling and EpCAM exosome secretion in HNSCC cells. Our results indicate that Rab11b is a superior prognostic indicator of HNSCC and holds promise for developing novel therapeutic strategies.

    File: 2023 12 Yoshida K Rab11 HNSC Exosome.pdf

    DOI: 10.1016/j.job.2023.11.007

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  • EpEX, the soluble extracellular domain of EpCAM, resists cetuximab treatment of EGFR-high head and neck squamous cell carcinoma. Reviewed International journal

    Koki Umemori, Kisho Ono, Takanori Eguchi, Hotaka Kawai, Tomoya Nakamura, Tatsuo Ogawa, Kunihiro Yoshida, Hideka Kanemoto, Kohei Sato, Kyoichi Obata, Shoji Ryumon, Hirokazu Yutori, Naoki Katase, Tatsuo Okui, Hitoshi Nagatsuka, Soichiro Ibaragi

    Oral oncology   142   106433 - 106433   2023.5

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    OBJECTIVES: Cetuximab (Cmab) is a molecularly targeted monoclonal antibody drug for head and neck squamous cell carcinoma (HNSC), although cetuximab resistance is a serious challenge. Epithelial cell adhesion molecule (EpCAM) is an established marker for many epithelial tumors, while the soluble EpCAM extracellular domain (EpEX) functions as a ligand for epidermal growth factor receptor (EGFR). We investigated the expression of EpCAM in HNSC, its involvement in Cmab action, and the mechanism by which soluble EpEX activated EGFR and played key roles in Cmab resistance. MATERIALS AND METHODS: We first examined EPCAM expression in HNSCs and its clinical significance by searching gene expression array databases. We then examined the effects of soluble EpEX and Cmab on intracellular signaling and Cmab efficacy in HNSC cell lines (HSC-3 and SAS). RESULTS: EPCAM expression was found to be enhanced in HNSC tumor tissues compared to normal tissues, and the enhancement was correlated with stage progression and prognosis. Soluble EpEX activated the EGFR-ERK signaling pathway and nuclear translocation of EpCAM intracellular domains (EpICDs) in HNSC cells. EpEX resisted the antitumor effect of Cmab in an EGFR expression-dependent manner. CONCLUSION: Soluble EpEX activates EGFR to increase Cmab resistance in HNSC cells. The EpEX-activated Cmab resistance in HNSC is potentially mediated by the EGFR-ERK signaling pathway and the EpCAM cleavage-induced nuclear translocation of EpICD. High expression and cleavage of EpCAM are potential biomarkers for predicting the clinical efficacy and resistance to Cmab.

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  • Stress-Inducible SCAND Factors Suppress the Stress Response and Are Biomarkers for Enhanced Prognosis in Cancers. Invited Reviewed International journal

    Mona Sheta, Kunihiro Yoshida, Hideka Kanemoto, Stuart K Calderwood, Takanori Eguchi

    International journal of molecular sciences   24 ( 6 )   2023.3

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    The cell stress response is an essential system present in every cell for responding and adapting to environmental stimulations. A major program for stress response is the heat shock factor (HSF)-heat shock protein (HSP) system that maintains proteostasis in cells and promotes cancer progression. However, less is known about how the cell stress response is regulated by alternative transcription factors. Here, we show that the SCAN domain (SCAND)-containing transcription factors (SCAN-TFs) are involved in repressing the stress response in cancer. SCAND1 and SCAND2 are SCAND-only proteins that can hetero-oligomerize with SCAN-zinc finger transcription factors, such as MZF1(ZSCAN6), for accessing DNA and transcriptionally co-repressing target genes. We found that heat stress induced the expression of SCAND1, SCAND2, and MZF1 bound to HSP90 gene promoter regions in prostate cancer cells. Moreover, heat stress switched the transcript variants' expression from long noncoding RNA (lncRNA-SCAND2P) to protein-coding mRNA of SCAND2, potentially by regulating alternative splicing. High expression of HSP90AA1 correlated with poorer prognoses in several cancer types, although SCAND1 and MZF1 blocked the heat shock responsiveness of HSP90AA1 in prostate cancer cells. Consistent with this, gene expression of SCAND2, SCAND1, and MZF1 was negatively correlated with HSP90 gene expression in prostate adenocarcinoma. By searching databases of patient-derived tumor samples, we found that MZF1 and SCAND2 RNA were more highly expressed in normal tissues than in tumor tissues in several cancer types. Of note, high RNA expression of SCAND2, SCAND1, and MZF1 correlated with enhanced prognoses of pancreatic cancer and head and neck cancers. Additionally, high expression of SCAND2 RNA was correlated with better prognoses of lung adenocarcinoma and sarcoma. These data suggest that the stress-inducible SCAN-TFs can function as a feedback system, suppressing excessive stress response and inhibiting cancers.

    File: 2023 03 Sheta IJMS Stress +supple.pdf

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  • Multiple Targeting of HSP Isoforms to Challenge Isoform Specificity and Compensatory Expression. International journal

    Kisho Ono, Takanori Eguchi

    Methods in molecular biology (Clifton, N.J.)   2693   141 - 161   2023

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    Heat shock proteins (HSPs) are molecular chaperones that assist in protein folding, trafficking, and metabolism. Intracellular chaperone functions of HSPs had been well-investigated, but extracellular and exosomal HSPs have been recently found. Exosomal HSPs are intercellularly transferred, while extracellular HSPs play cytokine-like roles called chaperokines. We have shown that exosomal HSPs play key roles in intercellular communication between tongue carcinoma and tumor-associated macrophages in the tumor microenvironment. Notably, HSP90 isoforms consist of HSP90alpha, HSP90beta, mitochondrial TRAP1, and GRP94 in the endoplasmic reticulum. Moreover, many pseudogenes of HSP90 can be transcribed into RNA. Besides, the function of HSP90 is defined by their cochaperones, such as CDC37 or AHA1. Therefore, isoform-specific small interfering RNA (siRNA) is necessary for precisely targeting each HSP90 isoform and cochaperone. Nevertheless, we often encountered compensatory expression of HSP90 isoforms in the knockdown studies. Here, we provide dual and triple knockdown methods to target multiple RNA for challenging isoform-specific roles and compensatory expression of intracellular, extracellular, and exosomal HSPs.

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  • Proteomic Profiling of the Extracellular Vesicle Chaperone in Cancer. International journal

    Kisho Ono, Takanori Eguchi

    Methods in molecular biology (Clifton, N.J.)   2693   233 - 249   2023

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    Molecular chaperones are widely distributed intracellular proteins that play essential roles in maintaining proteome function by assisting in the folding of client proteins. Molecular chaperones, such as heat shock proteins (HSPs), are found intracellularly and extracellularly. Extracellular vesicles (EVs), such as exosomes, contain HSPs and horizontally transfer the functional chaperones into various recipient cells. Besides, mass spectrometry has enabled a comprehensive analysis of exosomal and EV proteins, which is useful in basic biomedical research to clinical biomarker search. We have performed deep proteome analysis of EVs, including exosomes, from metastatic tongue and prostate cancers and detected >700 protein types, including cytoplasmic, ER, mitochondrial, small, and large HSPs. Here, we provide protocols for isolating exosomes/EVs and deep proteome analysis to detect the EV chaperone.

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  • Large-Scale Databases and Portals on Cancer Genome to Analyze Chaperone Genes Correlated to Patient Prognosis. International journal

    Kisho Ono, Takanori Eguchi

    Methods in molecular biology (Clifton, N.J.)   2693   293 - 306   2023

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    Molecular chaperones, such as heat shock proteins (HSPs), have attracted attention as molecules involved in malignant events in cancers and are potential therapeutic targets and biomarkers for tumor therapy. Furthermore, mutations in chaperones can significantly impact cancer risk and prognosis. Bioinformatics is a particularly useful method for developing biomarkers as a practical consideration for the immediate clinical application of data. Many large-scale databases and portals on cancer genome are nowadays publicly available, including the International Cancer Genome Consortium (ICGC); The Cancer Genome Atlas (TCGA), renamed as Genomic Data Commons (GDC); Catalogue of Somatic Mutations in Cancer (COSMIC); and Cancer Cell Line Encyclopedia (CCLE). Referring to these databases, advanced web portals are publicized, including cBioPortal, Human Protein Atlas (HPA), Kaplan-Meier (KM) plotter, Gene Expression Profiling Interactive Analysis 2 (GEPIA2), Genomics of Drug Sensitivity in Cancer (GDSC), and Dependency Map (DepMap). Here, we assemble these databases and portals to clarify what is available and useful for current cancer research and provide protocols to utilize the HPA, KM plotter, and GEPIA2 for studies on chaperone genes in cancer patients. Utilizing these portals will reveal the correlation between tumor subtype-specific high expression of chaperone genes and patient prognosis. Our protocols are useful to increase systematic awareness of chaperones and find new biomarkers for diagnosis and prognosis and new targets for anticancer drugs.

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  • Monitoring of the Heat Shock Response with a Real-Time Luciferase Reporter. International journal

    Andrew Ackerman, Toshiki Kijima, Takanori Eguchi, Thomas L Prince

    Methods in molecular biology (Clifton, N.J.)   2693   1 - 11   2023

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    The heat shock response (HSR) is a cellular mechanism for counteracting acute proteotoxic stress. In eukaryotes, transcriptional activation of the HSR is regulated by heat shock factor 1 (HSF1). Activation of HSF1 induces the expression of heat shock proteins (HSPs) that function as molecular chaperones to fold and maintain the three-dimensional structure of misfolded proteins. The regulation of the degree and duration of the HSR is controlled by multiple biochemical mechanisms that include posttranslational modification of HSF1 and numerous protein-protein interactions. In this chapter, we describe a method to evaluate the activation and deactivation of the HSR at the transcriptional level using a short half-life luciferase reporter assay. This assay can be used to further characterize the HSR or as a screen for small molecule inducers, amplifiers, or repressors.

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  • Protocol for CRISPR/Cas Genome Editing for Investigating Cell Communication Network. Invited Reviewed International journal

    Yuka Okusha, Takanori Eguchi

    Methods in molecular biology (Clifton, N.J.)   2582   157 - 167   2023

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    The Cellular Communication Network Factor (CCN) family is composed of six members: CCN1/CYR61, CCN2/CTGF, CCN3/NOV, CCN4/WISP1, CCN5/WISP2, and CCN6/WISP3. The second member, CCN2/CTGF is a matricellular protein that promotes extracellular matrix (ECM) synthesis and controls angiogenesis. On the other hand, moonlighting/matrix metalloproteinase 3 (MMP3) is an ECM-degrading enzyme that also functions as an intracellular transcription factor. Importantly, extracellular MMP3 is uptaken into cells, translocating into nuclei, and transcriptionally activating CCN2/CTGF gene in cancer and chondrocytes. Thus, the MMP3-CTGF axis balances the matrix metabolism and turnover in the tissue and tumor microenvironments. We established an MMP3 knockout cell line using the CRISPR/Cas9 system, demonstrating the sequential regulatory events of the MMP3-CCN2 axis in the microenvironment. Notably, our protocol is useful for generation of CCN knockout cells as well. Here we serve a protocol of the CRISPR/Cas9-based gene targeting in cultured cells for investigating cellular communication network.

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  • Transfection, Spinfection, Exofection, and Luciferase Assays for Analysis of CCN Genes Expression Mechanism. Invited Reviewed International journal

    Takanori Eguchi, Yanyin Lu, Eman A Taha, Yuka Okusha

    Methods in molecular biology (Clifton, N.J.)   2582   103 - 126   2023

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    Cell communication network factor 2 (CCN2), also known as connective tissue growth factor (CTGF), is protein inducible in response to TGFβ/Smad signal or the transcriptional activity of matrix metalloproteinase 3 (MMP3). We discovered that MMP3 in exosomes is transferable to recipient cells and then translocates into cell nuclei to transactivate the CCN2/CTGF gene. Exosomes and liposomes enable molecular transfection to recipient cells in vitro and in vivo. These small vesicles are surrounded by lipid membranes and carry proteins, RNA, DNA, and small chemicals. Here we define the exosome-based transfection as "exofection." In addition, spinfection increases the efficiencies of transfection, exofection, and viral infection, thus being compatible with various molecular transfer protocols. Here, we provide protocols, tips, and practical examples of transfection, spinfection, exofection, fluorescence microscopy, and luciferase assays to analyze the CCNs gene expression mechanisms.

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  • Comprehensive Method for Exosome Isolation and Proteome Analysis for Detection of CCN Factors in/on Exosomes. Invited Reviewed International journal

    Takanori Eguchi, Yuka Okusha, Yanyin Lu, Kisho Ono, Eman A Taha, Shiro Fukuoka

    Methods in molecular biology (Clifton, N.J.)   2582   59 - 76   2023

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    Cellular Communication Network (CCN) proteins are secretory growth factors often associated with extracellular matrix (ECM) and extracellular vesicles (EVs) such as exosomes or matrix-coated vesicles. CCN factors and fragments loaded on/in EVs may play key roles in cell communication networks in cancer biology, bone and cartilage metabolism, wound healing, and tissue regeneration. CCN proteins and EVs/exosomes are found in body fluids, such as blood, urine, milk, and supernatants of the two-dimensionally (2D) cultured cells and three-dimensionally (3D) cultured tissues, such as spheroids or organoids. More than ten methods to isolate exosomes or EVs have been developed with different properties. Here, we introduce comprehensive protocols for polymer-based precipitation, affinity purification, ultracentrifugation methods combined with the ultrafiltration method for isolating CCN-loaded exosomes/EVs from 2D and 3D cultured tissues, and proteome analysis using mass spectrometry for comprehensive analysis of CCN proteins.

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  • Western Blot Protocols for Analysis of CCN Proteins and Fragments in Exosomes, Vesicle-Free Fractions, and Cells. Reviewed International coauthorship International journal

    Kisho Ono, Yuka Okusha, Manh Tien Tran, Koki Umemori, Takanori Eguchi

    Methods in molecular biology (Clifton, N.J.)   2582   39 - 57   2023

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    Cellular Communication Network (CCN) proteins are growth factors that play key roles in many pathophysiological events, including bone formation, wound healing, and cancer. CCN factors and fragments generated by metalloproteinases-dependent cleavage are often associated with extracellular matrix (ECM) or small extracellular vesicles (sEVs) such as exosomes or matrix-coated vesicles. We provide reliable methods and protocols for Western blotting to analyze CCN factors and fragments in cells, sEVs, and vesicle-free fractions.

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  • Cdc37 as a Co-chaperone to Hsp90 Invited Reviewed International journal

    Thomas L Prince, Benjamin J Lang, Yuka Okusha, Takanori Eguchi, Stuart K Calderwood

    Sub-cellular biochemistry   101   141 - 158   2023

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    The co-chaperone p50/Cdc37 is an important partner for Hsp90, assisting in molecular chaperone activities, particularly with regard to the regulation of protein kinases. Analysis of the structure of Hsp90-Cdc37-kinase complexes demonstrates the way in which Cdc37 interacts with and controls the folding of a large proportion of intracellular protein kinases. This co-chaperone thus stands at the hub of a multitude of intracellular signaling networks. Indeed, the influence of Cdc37 reaches beyond the housekeeping pathways of protein folding into the regulation of a wide range of cellular processes. This co-chaperone has attracted attention as a potential intermediate in carcinogenesis. Cdc37 is an attractive potential target in cancer due to (1) high expression in a number of tumor types and (2) control of multiple signaling pathways. These properties indicate (3) a potential for selectivity due to its elevated expression in malignant cells and (4) robustness, as the co-chaperone may control multiple growth signaling pathways and thus be less prone to evolution of resistance than less versatile oncoproteins. Cdc37 may also be involved in other aspects of pathophysiology and has been shown to be secreted in exosomes. Protein aggregation disorders have been linked to age-related declines in molecular chaperones and co-chaperones. Cdc37 also appears to be a potential agent in longevity due to its links to protein folding and autophagy, and it will be informative to study the role of Cdc37 maintenance/decline in aging organisms.

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  • Multiplex Immunostaining Method to Distinguish HSP Isoforms in Cancer Tissue Specimens. Invited Reviewed International journal

    Hotaka Kawai, Kisho Ono, Takanori Eguchi

    Methods in molecular biology (Clifton, N.J.)   2693   281 - 291   2023

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    Heat shock proteins (HSPs) are often expressed in all nucleated cells, but their expression profiles differ. In particular, HSP90α and HSP90β have high sequence identity and have not been fully examined for their individual and compensatory functions as molecular chaperones, differences in client proteins, and extracellular distributions with exosomes. Immunohistochemical staining is a technique to visualize the presence and localization of target antigens using specific antibodies, of which the multiplex immunostaining method can reveal differences in protein expression in the same tumor tissue and the localization of proteins of interest within tumor tissue or single cells. The common multiplex immunostaining method uses multiple secondary antibodies of different reacting animal species to identify and detect different antigens, thus requiring different animals to be immunized with each primary antibody. Furthermore, the fluorescent-antibody method is the predominant multiplex staining method but has the critical disadvantage that permanent specimens cannot be prepared. Here, we outline a multiplex staining method for HSP90α and HSP90β based on the enzyme-antibody method that allows permanent specimens to be prepared without the restriction of immunized animal species.

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  • Roles of Cancer Exosomes in Immunosuppression and Immune Evasion

    Mona Sheta, Eman Ahmed Taha, Yanyin Lu, Takanori Eguchi

    Preprints   2022110004   2022.11

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    Extracellular vesicles (EV), including exosomes and microvesicles, are released from various cells and alter recipient cell phenotypes and fates by their biomolecules. Here we review current knowledge about tumor EVs and how they prompt malignant cell communication with tumor-associated cells, such as cancer-associated fibroblasts, tumor endothelial cells, and immune cells. We delineate the major pathways and molecular players that influence each step of cancer initiation, progression, and resistance. Of note, cancer exosomes involve immunosuppression by tumor-associated macrophages, myeloid-derived suppressor cells, and regulatory T cells. Moreover, tumor exosomes can induce the apoptosis of killer T cells and immune checkpoint of dendritic cells and attenuate natural killer cells. An in-depth understanding of EV biology is essential to ensure the clinical development of exosome/EV-based therapeutic products, which will be of benefit to exosome manipulation in cancer management.

    DOI: 10.20944/preprints202211.0004.v1

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  • SCAND1 Reverts EMT and Suppresses Tumor Growth and Collective Migration

    Takanori Eguchi, Eva Csizmadia, Thomas L. Prince, Barbara Wegiel, Stuart K. Calderwood

    Preprints   2022.10

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    Epithelial-mesenchymal transition (EMT) is a reversible cellular program that transiently places epithelial (E) cells into pseudo-mesenchymal (M) cell states. The malignant progression and resistance of many types of carcinomas depends on EMT activation, partial EMT and hybrid E/M status in neoplastic cells. EMT is activated by tumor microenvironmental TGFβ signal and EMT-inducing transcription factors, such as ZEB1/2 in tumor cells. However, reverse EMT factors are less studied. We demonstrate that transcription factor SCAND1 can revert mesenchymal and hybrid E/M phenotype of cancer cells to a more epithelial, less invasive status and inhibit their proliferation and migration. SCAND1 is a SCAN domain-containing protein and hetero-oligomerizes with SCAN-zinc finger transcription factors, such as MZF1, for accessing DNA and transcriptional co-repression of target genes. We found that SCAND1-MZF1 co-expression and interaction correlated with maintaining epithelial features, whereas the simultaneous loss of SCAND1 and MZF1 correlated with mesenchymal features of tumor cells. Overexpression of SCAND1 over endogenous MZF1 in DU-145 prostate cancer cells reverted their hybrid E/M status into cobblestone morphology with increased epithelial adhesion by E-cadherin and β-catenin relocation. Consistently, co-expression analysis in TCGA PanCancer Atlas revealed that both SCAND1 and MZF1 co-express and are negatively correlated with EMT driver genes, including CTNNB1, ZEB1, ZEB2 and TGFBR, in prostate tumor specimens. In addition, SCAND1 overexpression suppressed tumor cell proliferation by reducing the MAP3K-MEK-ERK signaling pathway. Of note, SCAND1-overexpressing DU-145 cells migrated slower than control cells with decreased lymph node metastasis of prostate cancer in a mouse tumor xenograft model. Kaplan-Meyer analysis showed high expression of MZF1 and SCAND1 to correlate with better prognoses in pancreatic cancer and head and neck cancers, although with poorer prognosis in kidney cancer. Overall, these data suggest that the combination of SCAND1-MZF1 complexes may revert the EMT mechanism in cancer to establish an epithelial phenotype. These effects seem to include co-repression of EMT-driver genes and suppression of tumor cell proliferation via inhibition of the MAP3K-MEK-ERK signaling pathway.

    DOI: 10.20944/preprints202210.0475.v1

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  • HSP90 drives the Rab11a-mediated vesicular transport of the cell surface receptors in osteoclasts. Reviewed International journal

    Manh Tien Tran, Yuka Okusha, Kaung Htike, Chiharu Sogawa, Takanori Eguchi, Tomoko Kadowaki, Eiko Sakai, Takayuki Tsukuba, Kuniaki Okamoto

    Cell biochemistry and function   40 ( 8 )   838 - 855   2022.9

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    Rab11a, which ubiquitously localizes to early and recycling endosomes, is required for regulating the vesicular transport of cellular cargos. Interestingly, our previous study revealed that Rab11a served as a negative regulator of osteoclastogenesis by facilitating the lysosomal proteolysis of (1) colony-stimulating factor-1 (c-fms) receptor and (2) receptor activator of nuclear factor-κB (RANK) receptor, thereby resulting in inhibition of osteoclast (OC) differentiation, maturation, and bone-resorbing activity. However, the molecular mechanisms of how Rab11a negatively affected osteoclastogenesis were largely unknown. Heat shock protein (HSP90), including two isoforms HSP90α and HSP90β, necessitates the stability, maturation, and activity of a broad range of its clients, and is essentially required for a vast array of signal transduction pathways in nonstressful conditions. Furthermore, cumulative evidence suggests that HSP90 is a vital element of the vesicular transport network. Indeed, our recent study revealed that HSP90, a novel effector protein of Rab11b, modulated Rab11b-mediated osteoclastogenesis. In this study, we also found that Rab11a interacted with both HSP90α and HSP90β in OCs. Upon blockade of HSP90 ATPase activity by a specific inhibitor(17-allylamino-demethoxygeldanamycin), we showed that (1) the ATPase domain of HSP90 was a prerequisite for the interaction between HSP90 and Rab11a, and (2) the interaction of HSP90 to Rab11a sufficiently maintained the inhibitory effects of Rab11a on osteoclastogenesis. Altogether, our findings undoubtedly indicate a novel role of HSP90 in regulating Rab11a-mediated osteoclastogenesis.

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  • 口腔癌間質細胞由来CCL2による骨髄由来細胞動員への関与(Resident stroma-secreted chemokine CCL2 governs myeloid-derived suppressor cells in the tumor microenvironment)

    河合 穂高, 冨田 秀太, 江口 傑徳, 大原 利章, 小野 喜章, 長塚 仁

    日本癌学会総会記事   81回   P - 3186   2022.9

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  • 口腔癌間質由来のCCL2はCCR2陽性骨髄由来免疫抑制細胞の腫瘍間質への動員に関与する

    河合 穂高, メイ・ワトウ, 高畠 清文, 冨田 秀太, 小野 喜章, 江口 傑徳, 大原 利章, 中野 敬介, 長塚 仁

    日本がん免疫学会総会プログラム・抄録集   26回   91 - 91   2022.6

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  • Rab34 plays a critical role as a bi-directional regulator of osteoclastogenesis Reviewed International journal

    Yunxia Feng, Manh Tien Tran, co-first author, Yanyin Lu, Kaung Htike, Yuka Okusha, Chiharu Sogawa, Takanori Eguchi, Tomoko Kadowaki, Eiko Sakai, Takayuki Tsukuba, Kuniaki Okamoto

    Cell Biochemistry and Function   40 ( 3 )   263 - 277   2022.3

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    Accumulating evidence suggests that Rab GTPases representing the largest branch of Ras superfamily have recently emerged as the core factors for the regulation of osteoclastogenesis through modulating vesicular transport amongst specific subcellular compartments. Among these, Rab34 GTPase has been identified to be important for the post-Golgi secretory pathway and for phagocytosis; nevertheless, its specific role in osteoclastogenesis has been completely obscure. Here, upon the in vitro model of osteoclast formation derived from murine macrophages like RAW-D cells or bone marrow-derived macrophages, we reveal that Rab34 regulates osteoclastogenesis bidirectionally. More specifically, Rab34 serves as a negative regulator of osteoclast differentiation by promoting the lysosome-induced proteolysis of two osteoclastogenic surface receptors, c-fms and RANK, via the axis of early endosomes-late endosomes-lysosomes, leading to alleviate the transcriptional activity of two of the master regulator of osteoclast differentiation, c-fos and NFATc-1, eventually attenuating osteoclast differentiation and bone resorption. Besides, Rab34 plays a crucial role in modulating the secretory network of lysosome-related proteases including matrix metalloprotease 9 and Cathepsin K across the ruffled borders of osteoclasts, contributing to the regulation of bone resorption.

    DOI: 10.1002/cbf.3691

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    Other Link: https://onlinelibrary.wiley.com/doi/full-xml/10.1002/cbf.3691

  • Resident stroma-secreted chemokine CCL2 governs myeloid-derived suppressor cells in the tumor microenvironment. Reviewed International journal

    May Wathone Oo, Hotaka Kawai, Kiyofumi Takabatake, Shuta Tomida, Takanori Eguchi, Kisho Ono, Qiusheng Shan, Toshiaki Ohara, Saori Yoshida, Haruka Omori, Shintaro Sukegawa, Keisuke Nakano, Kuniaki Okamoto, Akira Sasaki, Hitoshi Nagatsuka

    JCI insight   7 ( 1 )   3186   2022.1

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    Accumulating evidence has shown that cancer stroma and bone marrow-derived cells (BMDCs) in the tumor microenvironment (TME) play vital roles in tumor progression. However, the mechanism by which oral cancer stroma recruits any particular subset of BMDCs remains largely unknown. Here we sought to identify the subset of BMDCs that is recruited by cancer stroma. We established a sequential transplantation model in BALB/c nude mice, including (i) bone marrow transplantation of GFP-expressing cells and (ii) co-xenografting of patient-derived stroma (two cases, designated PDS1 and PDS2) with oral cancer cells (HSC-2). As controls, xenografting was performed with HSC-2 alone or in combination with normal human dermal fibroblasts (HDF). PDS1, PDS2, and HDF all promoted BMDCs migration in vitro and recruitment in vivo. Multicolor immunofluorescence revealed that the PDS co-xenografts recruited Arginase-1/CD11b/GR1/GFP quadruple-positive cells, which are myeloid-derived suppressor cells (MDSCs), to the TME, whereas the HDF co-xenograft did not. Screening using microarrays revealed that PDS1 and PDS2 expressed CCL2 mRNA (encoding C-C motif chemokine ligand 2) at higher levels than did HDF. Indeed, PDS xenografts contained significantly higher proportions of CCL2-positive stromal cells and CCR2/Arginase-1/CD11b/GR1 quadruple-positive MDSCs (as receiver cells) than the HDF co-xenograft. Consistently, a CCL2 synthesis inhibitor and a CCR2 antagonist significantly inhibited the PDS-driven migration of BM cells in vitro. Furthermore, intraperitoneal injection of the CCR2 antagonist to the PDS xenograft models significantly reduced the CCR2/Arginase-1/CD11b/GR1 quadruple-positive MDSCs infiltration to the TME. In conclusion, oral cancer stroma-secreted CCL2 is a key signal for recruiting CCR2-positive MDSCs from bone marrow to the TME.

    DOI: 10.1172/jci.insight.148960

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  • Porphyromonas gingivalisはマクロファージの細胞外小胞を介して胎盤・胎児の成長発育を阻害する(Porphyromonas gingivalis impairs placental and fetal development through macrophage-derived extracellular vesicles)

    棚井 あいり, 福原 瑶子, 江口 傑徳, 河合 穂高, 池亀 美華, 岡村 裕彦, 植田 幸嗣

    Journal of Oral Biosciences Supplement   2021   205 - 205   2021.10

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  • A novel role of HSP90 in regulating osteoclastogenesis by abrogating Rab11b-driven transport. Reviewed International journal

    Manh Tien Tran, Yuka Okusha, Yunxia Feng, Chiharu Sogawa, Takanori Eguchi, Tomoko Kadowaki, Eiko Sakai, Takayuki Tsukuba, Kuniaki Okamoto

    Biochimica et biophysica acta. Molecular cell research   1868 ( 10 )   119096 - 119096   2021.9

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    Heat shock protein 90 (HSP90) is a highly conserved molecular chaperone that plays a pivotal role in folding, activating and assembling a variety of client proteins. In addition, HSP90 has recently emerged as a crucial regulator of vesicular transport of cellular proteins. In our previous study, we revealed Rab11b negatively regulated osteoclastogenesis by promoting the lysosomal proteolysis of c-fms and RANK surface receptors via the axis of early endosome-late endosome-lysosomes. In this study, using an in vitro model of osteoclasts differentiated from murine macrophage-like RAW-D cells, we revealed that Rab11b interacted with both HSP90 isoforms, HSP90 alpha (HSP90α) and HSP90 beta (HSP90β), suggesting that Rab11b is an HSP90 client. Using at specific blocker for HSP90 ATPase activity, 17-allylamino-demethoxygeldanamycin (17-AAG), we found that the HSP90 ATPase domain is indispensable for maintaining the interaction between HSP90 and Rab11b in osteoclasts. Nonetheless, its ATPase activity is not required for regulating the turnover of endogenous Rab11b. Interestingly, blocking the interaction between HSP90 and Rab11b by either HSP90-targeting small interfering RNA (siHSP90) or 17-AAG abrogated the inhibitory effects of Rab11b on osteoclastogenesis by suppressing the Rab11b-mediated transport of c-fms and RANK surface receptors to lysosomes via the axis of early endosome-late endosome-lysosomes, alleviating the Rab11b-mediated proteolysis of these surface receptors in osteoclasts. Based on our observations, we propose a HSP90/Rab11b-mediated regulatory mechanism for osteoclastogenesis by directly modulating the c-fms and RANK surface receptors in osteoclasts, thereby contributing to the maintenance of bone homeostasis.

    DOI: 10.1016/j.bbamcr.2021.119096

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  • Extracellular Hsp90 and protection of neuronal cells through Nrf2 Reviewed International journal

    Stuart K Calderwood, Thiago J Borges, Takanori Eguchi, Benjamin J Lang, Ayesha Murshid, Yuka Okusha, Thomas L Prince

    Biochemical Society Transactions   49 ( 5 )   2299 - 2306   2021.8

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    Heat shock protein 90 (Hsp90), although one of the most essential intracellular chaperones, can also play key roles in the extracellular milieu. Here, we review the properties of extracellular Hsp90 in cellular homeostasis in the heat shock response (HSR), focusing on cells of the central nervous system. Hsp90 can be secreted by microglia as well as other cell types by non-canonical pathways of secretion. The chaperone may then influence the behavior of distant cells and can for instance protect neuronal cells from the oxidative burst accompanying phagocytosis by microglia of beta-amyloid fibrils. A mechanism involving activation of the transcription factor Nrf2, and induction of the antioxidant response is reported. We review the potential role of extracellular Hsp90, Nrf2 and transcellular chaperone signaling in the non-cell-intrinsic HSR.

    DOI: 10.1042/BST20210370

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  • Exosome-Based Molecular Transfer Activity of Macrophage-Like Cells Involves Viability of Oral Carcinoma Cells: Size Exclusion Chromatography and Concentration Filter Method. Reviewed International journal

    Yanyin Lu, Takanori Eguchi, Chiharu Sogawa, Eman A Taha, Manh Tien Tran, Toshiki Nara, Penggong Wei, Shiro Fukuoka, Takuya Miyawaki, Kuniaki Okamoto

    Cells   10 ( 6 )   1 - 23   2021.5

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    Extracellular vesicles (EV) heterogeneity is a crucial issue in biology and medicine. In addition, tumor-associated macrophages are key components in cancer microenvironment and immunology. We developed a combination method of size exclusion chromatography and concentration filters (SEC-CF) and aimed to characterize different EV types by their size, cargo types, and functions. A human monocytic leukemia cell line THP-1 was differentiated to CD14-positive macrophage-like cells by stimulation with PMA (phorbol 12-myristate 13-acetate) but not M1 or M2 types. Using the SEC-CF method, the following five EV types were fractionated from the culture supernatant of macrophage-like cells: (i) rare large EVs (500-3000 nm) reminiscent of apoptosomes, (ii) EVs (100-500 nm) reminiscent of microvesicles (or microparticles), (iii) EVs (80-300 nm) containing CD9-positive large exosomes (EXO-L), (iv) EVs (20-200 nm) containing unidentified vesicles/particles, and (v) EVs (10-70 nm) containing CD63/HSP90-positive small exosomes (EXO-S) and particles. For a molecular transfer assay, we developed a THP-1-based stable cell line producinga GFP-fused palmitoylation signal (palmGFP) associated with the membrane. The THP1/palmGFP cells were differentiated into macrophages producing palmGFP-contained EVs. The macrophage/palmGFP-secreted EXO-S and EXO-L efficiently transferred the palmGFP to receiver human oral carcinoma cells (HSC-3/palmTomato), as compared to other EV types. In addition, the macrophage-secreted EXO-S and EXO-L significantly reduced the cell viability (ATP content) in oral carcinoma cells. Taken together, the SEC-CF method is useful for the purification of large and small exosomes with higher molecular transfer activities, enabling efficient molecular delivery to target cells.

    File: 2021 05 Lu, Eguchi, et al, Cells.pdf

    DOI: 10.3390/cells10061328

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  • Macrophage-Derived Small Exosomes: Efficient Transmission and Cytotoxicity to Cancer Cells

    Yanyin Lu, Takanori Eguchi, Chiharu Sogawa, Eman A. Taha, Manh Tien Tran, Toshiki Nara, Penggong Wei, Shiro Fukuoka, Takuya Miyawaki, Kuniaki Okamoto

    Preprints   doi: 10.20944/preprints202104.0464.v1   2021.4

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    Tumor-associated macrophages are a key component in the tumor microenvironment, secreting extracellular vesicles (EVs) such as exosomes and other various factors for intercellular communication. However, macrophage-derived EVs heterogeneity and their cytotoxicity to cancer cells has not been well understood. Here, we aimed to separately isolate various types of macro-phage-EVs by size exclusion chromatography (SEC) method and investigate EV transmission and cytotoxicity to oral cancer cells. For fluorescence-labeling of cellular and EV membranes, palmitoylation signal-fused GFP and tdTomato were expressed in THP-1 monocytic cells and HSC-3 oral cancer cells, respectively. We found that fluorescence-labeled EVs secreted by macrophages were highly transmissive to oral cancer cells than those from parental monocytic cells. In a co-culture system and conditioned medium (CM), a macrophage-secreted unidentified factor was cytotoxic to oral cancer cells. We fractionated macrophage-derived EVs by the SEC method and performed western blotting to characterize various EV types. Three fractions were characterized: small exosomes (EXO-S: < 50 nm) fraction containing HSP90α, HSP90β, CD63 (EV marker) and β-actin; large exosomes (EXO-L: 50-200 nm) fraction containing CD9 (EV marker) and HSP90β; large EVs (100-500 nm) fraction. Notably, the macrophage-derived small exosomes fraction was cytotoxic to oral cancer cells, while large exosomes and large EVs were not. There-fore, it was implicated that macrophage-derived small exosomes are cytotoxic with high trans-mission potential to cancer cells.

    DOI: 10.20944/preprints202104.0464.v1

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  • Gel-Free 3D Tumoroids with Stem Cell Properties Modeling Drug Resistance to Cisplatin and Imatinib in Metastatic Colorectal Cancer. Reviewed International journal

    Chiharu Sogawa, Takanori Eguchi, Yuri Namba, Yuka Okusha, Eriko Aoyama, Kazumi Ohyama, Kuniaki Okamoto

    Cells   10 ( 2 )   1 - 17   2021.2

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    Researchers have developed several three-dimensional (3D) culture systems, including spheroids, organoids, and tumoroids with increased properties of cancer stem cells (CSCs), also called cancer-initiating cells (CICs). Drug resistance is a crucial issue involving recurrence in cancer patients. Many studies on anti-cancer drugs have been reported using 2D culture systems, whereas 3D cultured tumoroids have many advantages for assessing drug sensitivity and resistance. Here, we aimed to investigate whether Cisplatin (a DNA crosslinker), Imatinib (a multiple tyrosine kinase inhibitor), and 5-Fluorouracil (5-FU: an antimetabolite) alter the tumoroid growth of metastatic colorectal cancer (mCRC). Gene expression signatures of highly metastatic aggregative CRC (LuM1 cells) vs. low-metastatic, non-aggregative CRC (Colon26 and NM11 cells) were analyzed using microarray. To establish a 3D culture-based multiplexing reporter assay system, LuM1 was stably transfected with the Mmp9 promoter-driven ZsGreen fluorescence reporter gene, which was designated as LuM1/m9 cells and cultured in NanoCulture Plate®, a gel-free 3D culture device. LuM1 cells highly expressed mRNA encoding ABCG2 (a drug resistance pump, i.e., CSC/CIC marker), other CSC/CIC markers (DLL1, EpCAM, podoplanin, STAT3/5), pluripotent stem cell markers (Sox4/7, N-myc, GATA3, Nanog), and metastatic markers (MMPs, Integrins, EGFR), compared to the other two cell types. Hoechst efflux stem cell-like side population was increased in LuM1 (7.8%) compared with Colon26 (2.9%), both of which were markedly reduced by verapamil treatment, an ABCG2 inhibitor. Smaller cell aggregates of LuM1 were more sensitive to Cisplatin (at 10 μM), whereas larger tumoroids with increased ABCG2 expression were insensitive. Notably, Cisplatin (2 μM) and Imatinib (10 μM) at low concentrations significantly promoted tumoroid formation (cell aggregation) and increased Mmp9 promoter activity in mCRC LuM1/m9, while not cytotoxic to them. On the other hand, 5-FU significantly inhibited tumoroid growth, although not completely. Thus, drug resistance in cancer with increased stem cell properties was modeled using the gel-free 3D cultured tumoroid system. The tumoroid culture is useful and easily accessible for the assessment of drug sensitivity and resistance.

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  • Extracellular Vesicle-Associated Moonlighting Proteins: Heat Shock Proteins and Metalloproteinases Invited Reviewed

    Takanori Eguchi, Eman Ahmed Taha

    Heat Shock Proteins   22   1 - 18   2021

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    DOI: 10.1007/7515_2020_25

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  • Heat Shock Proteins and Periodontitis – Cross-Reaction Between Bacterial and Human HSP in Periodontal Infection Linking with Cardiovascular Diseases Invited Reviewed

    Tadashi Yamamoto, Takanori Eguchi

    Heat Shock Proteins   22   19 - 32   2021

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    DOI: 10.1007/7515_2020_24

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  • HSP Stimulation on Macrophages and Dendritic Cells Activates Innate Immune System Invited Reviewed

    Yanyin Lu, Takanori Eguchi

    Heat Shock Proteins   22   53 - 67   2021

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    DOI: 10.1007/7515_2020_26

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  • Liquid-based 3D Cultured Tumoroids Modeling Resistance to Cisplatin and Imatinib in Metastatic Colorectal Cancer

    Chiharu Sogawa, Takanori Eguchi, Yuri Namba, Eriko Aoyama, Kazumi Ohyama, Kuniaki Okamoto

    Preprints   doi: 10.20944/preprints202012.0582.v1   2020.12

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    Researchers have developed and used several three-dimensional (3D) culture systems, including spheroids, organoids, and tumoroids. Drug resistance is a crucial issue involving recurrence in cancer patients. Many studies on anticancer drugs have been done in 2D culture systems, where-as 3D cultured tumoroids have many advantages for assessing drug sensitivity and resistance. Here, we aim to investigate whether Cisplatin (a DNA crosslinker), Imatinib (a multiple tyro-sine kinase inhibitor), and 5-Fluorouracil (5-FU: an antimetabolite) alter tumoroid growth of metastatic colorectal cancer (mCRC). To establish a liquid-based 3D multiplexing reporter assay system, LuM1 (a murine mCRC cell line) was stably transfected with the Mmp9 promoter-driven ZsGreen reporter gene, which was designated as LuM1/m9 cells and cultured in NanoCulture Plate (NCP), a 3D culture device. The larger tumoroids were not sensitive to Cisplatin and ex-pressed ABCG2 (a marker of cancer stem cells, a.k.a. a drug efflux transporter), whereas smaller cell-aggregates were more sensitive to Cisplatin. Both Imatinib and Cisplatin significantly in-creased tumoroid growth (larger than 300 μm2) and Mmp9 promoter activity and were not cytotoxic to the mCRC tumoroids. On the other hand, 5-FU was cytotoxic to the tumoroids and significantly inhibited tumoroid growth, although not completely. Thus, platinum resistance and imatinib resistance in mCRC were modeled using the liquid-based 3D cultured tumoroid system. The tumoroid culture is useful and easily accessible for the assessment of drug sensitivity and resistance.

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  • The Inhibitory Role of Rab11b in Osteoclastogenesis through Triggering Lysosome-Induced Degradation of c-Fms and RANK Surface Receptors. Reviewed International journal

    Manh Tien Tran, Yuka Okusha, Yunxia Feng, Masatoshi Morimatsu, Penggong Wei, Chiharu Sogawa, Takanori Eguchi, Tomoko Kadowaki, Eiko Sakai, Hirohiko Okamura, Keiji Naruse, Takayuki Tsukuba, Kuniaki Okamoto

    International journal of molecular sciences   21 ( 24 )   9352 - 9352   2020.12

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    Rab11b, abundantly enriched in endocytic recycling compartments, is required for the establishment of the machinery of vesicle trafficking. Yet, no report has so far characterized the biological function of Rab11b in osteoclastogenesis. Using in vitro model of osteoclasts differentiated from murine macrophages like RAW-D cells or bone marrow-derived macrophages, we elucidated that Rab11b served as an inhibitory regulator of osteoclast differentiation sequentially via (i) abolishing surface abundance of RANK and c-Fms receptors; and (ii) attenuating nuclear factor of activated T-cells c1 (NFATc-1) upstream signaling cascades, following RANKL stimulation. Rab11b was localized in early and late endosomes, Golgi complex, and endoplasmic reticulum; moreover, its overexpression enlarged early and late endosomes. Upon inhibition of lysosomal function by a specific blocker, chloroquine (CLQ), we comprehensively clarified a novel function of lysosomes on mediating proteolytic degradation of c-Fms and RANK surface receptors, drastically ameliorated by Rab11b overexpression in RAW-D cell-derived osteoclasts. These findings highlight the key role of Rab11b as an inhibitor of osteoclastogenesis by directing the transport of c-Fms and RANK surface receptors to lysosomes for degradation via the axis of early endosomes-late endosomes-lysosomes, thereby contributing towards the systemic equilibrium of the bone resorption phase.

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  • 侵襲性歯周炎の血液診断バイオマーカーとしての細胞外小胞由来マイクロRNAの探索

    河本 美奈, 山本 直史, 河村 麻理, 森 彩乃, 山城 圭介, 大森 一弘, 小野 喜章, 江口 傑徳, 十川 千春, 高柴 正悟

    岡山歯学会雑誌   39 ( 2 )   35 - 36   2020.12

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  • Organoids and Liquid Biopsy in Oral Cancer Research. Invited Reviewed International journal

    Takanori Eguchi

    Journal of clinical medicine   9 ( 11 )   1 - 5   2020.11

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    To promote the newest discoveries in oral cancer research, a special issue "Frontiers in Oral Cancer-Basic and Clinical Sciences" in the Journal of Clinical Medicine (JCM) was opened from September 2019 to April 2020 [...].

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  • Rab11A Functions as a Negative Regulator of Osteoclastogenesis through Dictating Lysosome-Induced Proteolysis of c-fms and RANK Surface Receptors. Reviewed International journal

    Yuka Okusha, Manh Tien Tran, Mami Itagaki, Chiharu Sogawa, Takanori Eguchi, Tatsuo Okui, Tomoko Kadowaki, Eiko Sakai, Takayuki Tsukuba, Kuniaki Okamoto

    Cells   9 ( 11 )   2384 - 2384   2020.10

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    Osteoclast differentiation and activity are controlled by two essential cytokines, macrophage colony-stimulating factor (M-CSF) and the receptor activator of nuclear factor-κB ligand (RANKL). Rab11A GTPase, belonging to Rab11 subfamily representing the largest branch of Ras superfamily of small GTPases, has been identified as one of the crucial regulators of cell surface receptor recycling. Nevertheless, the regulatory role of Rab11A in osteoclast differentiation has been completely unknown. In this study, we found that Rab11A was strongly upregulated at a late stage of osteoclast differentiation derived from bone marrow-derived macrophages (BMMs) or RAW-D murine osteoclast precursor cells. Rab11A silencing promoted osteoclast formation and significantly increased the surface levels of c-fms and receptor activator of nuclear factor-κB (RANK) while its overexpression attenuated osteoclast formation and the surface levels of c-fms and RANK. Using immunocytochemical staining for tracking Rab11A vesicular localization, we observed that Rab11A was localized in early and late endosomes, but not lysosomes. Intriguingly, Rab11A overexpression caused the enhancement of fluorescent intensity and size-based enlargement of early endosomes. Besides, Rab11A overexpression promoted lysosomal activity via elevating the endogenous levels of a specific lysosomal protein, LAMP1, and two key lysosomal enzymes, cathepsins B and D in osteoclasts. More importantly, inhibition of the lysosomal activity by chloroquine, we found that the endogenous levels of c-fms and RANK proteins were enhanced in osteoclasts. From these observations, we suggest a novel function of Rab11A as a negative regulator of osteoclastogenesis mainly through (i) abolishing the surface abundance of c-fms and RANK receptors, and (ii) upregulating lysosomal activity, subsequently augmenting the degradation of c-fms and RANK receptors, probably via the axis of early endosomes-late endosomes-lysosomes in osteoclasts.

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  • Roles of HSP on Antigen Presentation Invited Reviewed

    Kazuyuki Furuta, Taka Eguchi

    Heat Shock Proteins in Human Diseases   275 - 280   2020.7

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    DOI: 10.1007/7515_2020_5

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  • Outer membrane vesicles of Porphyromonas gingivalis attenuate insulin sensitivity by delivering gingipains to the liver. Reviewed International journal

    Mariko Seyama, Kaya Yoshida, Kayo Yoshida, Natsumi Fujiwara, Kisho Ono, Takanori Eguchi, Hotaka Kawai, Jiajie Guo, Yao Weng, Yuan Haoze, Kenta Uchibe, Mika Ikegame, Akira Sasaki, Hitoshi Nagatsuka, Kuniaki Okamoto, Hirohiko Okamura, Kazumi Ozaki

    Biochimica et biophysica acta. Molecular basis of disease   1866 ( 6 )   165731 - 165731   2020.6

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    Outer membrane vesicles (OMVs) are nanosized particles derived from the outer membrane of gram-negative bacteria. Oral bacterium Porphyromonas gingivalis (Pg) is known to be a major pathogen of periodontitis that contributes to the progression of periodontal disease by releasing OMVs. The effect of Pg OMVs on systemic diseases is still unknown. To verify whether Pg OMVs affect the progress of diabetes mellitus, we analyzed the cargo proteins of vesicles and evaluated their effect on hepatic glucose metabolism. Here, we show that Pg OMVs were equipped with Pg-derived proteases gingipains and translocated to the liver in mice. In these mice, the hepatic glycogen synthesis in response to insulin was decreased, and thus high blood glucose levels were maintained. Pg OMVs also attenuated the insulin-induced Akt/glycogen synthase kinase-3 β (GSK-3β) signaling in a gingipain-dependent fashion in hepatic HepG2 cells. These results suggest that the delivery of gingipains mediated by Pg OMV elicits changes in glucose metabolisms in the liver and contributes to the progression of diabetes mellitus.

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  • A Novel Model of Cancer Drug Resistance: Oncosomal Release of Cytotoxic and Antibody-Based Drugs. Reviewed International journal

    Takanori Eguchi, Eman Ahmed Taha, Stuart K Calderwood, Kisho Ono

    Biology   9 ( 3 )   1 - 22   2020.3

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    Extracellular vesicles (EVs), such as exosomes or oncosomes, often carry oncogenic molecules derived from tumor cells. In addition, accumulating evidence indicates that tumor cells can eject anti-cancer drugs such as chemotherapeutics and targeted drugs within EVs, a novel mechanism of drug resistance. The EV-releasing drug resistance phenotype is often coupled with cellular dedifferentiation and transformation in cells undergoing epithelial-mesenchymal transition (EMT), and the adoption of a cancer stem cell phenotype. The release of EVs is also involved in immunosuppression. Herein, we address different aspects by which EVs modulate the tumor microenvironment to become resistant to anticancer and antibody-based drugs, as well as the concept of the resistance-associated secretory phenotype (RASP).

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    DOI: 10.3390/biology9030047

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  • Antiparkinson Drug Benztropine Suppresses Tumor Growth, Circulating Tumor Cells, and Metastasis by Acting on SLC6A3/DAT and Reducing STAT3. Reviewed International journal

    Chiharu Sogawa, Takanori Eguchi, Manh Tien Tran, Masayuki Ishige, Kilian Trin, Yuka Okusha, Eman Ahmed Taha, Yanyin Lu, Hotaka Kawai, Norio Sogawa, Masaharu Takigawa, Stuart K Calderwood, Kuniaki Okamoto, Ken-Ichi Kozaki

    Cancers   12 ( 2 )   1 - 22   2020.2

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    Tumor growth, progression, and therapy resistance are crucial factors in the prognosis of cancer. The properties of three-dimensional (3D) tumor-like organoids (tumoroids) more closely resemble in vivo tumors compared to two-dimensionally cultured cells and are therefore effectively used for assays and drug screening. We here established a repurposed drug for novel anticancer research and therapeutics using a 3D tumoroid-based screening system. We screened six pharmacologically active compounds by using an original tumoroid-based multiplex phenotypic screening system with a matrix metalloproteinase 9 (MMP9) promoter-driven fluorescence reporter for the evaluation of both tumoroid formation and progression. The antiparkinson drug benztropine was the most effective compound uncovered by the screen. Benztropine significantly inhibited in vitro tumoroid formation, cancer cell survival, and MMP9 promoter activity. Benztropine also reduced the activity of oncogenic signaling transducers and trans-activators for MMP9, including STAT3, NF-κB, and β-catenin, and the properties of cancer stem cells/cancer-initiating cells. Benztropine and GBR-12935 directly targeted the dopamine transporter DAT/SLC6A3, whose genetic alterations such as amplification were correlated with poor prognosis for cancer patients. Benztropine also inhibited the tumor growth, circulating tumor cell (CTC) number, and rate of metastasis in a tumor allograft model in mice. In conclusion, we propose the repurposing of benztropine for anticancer research and therapeutics that can suppress tumor progression, CTC, and metastasis of aggressive cancers by reducing key pro-tumorigenic factors.

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  • Extracellular Oncosomes Rich in Moonlighting Metalloproteinase (MMP3) Are Transmissive, Pro-Tumorigenic, and Induces Cellular Communication Network Factor 2 (CCN2/CTGF): CRISPR against Cancer

    Yuka Okusha, Takanori Eguchi, Manh Tien Tran, Chiharu Sogawa, Kaya Yoshida, Mami Itagaki, Eman Ahmed Taha, Kisho Ono, Eriko Aoyama, Hirohiko Okamura, Ken-ichi Kozaki, Stuart K. Calderwood, Masaharu Takigawa, Kuniaki Okamoto

    Preprints   doi: 10.20944/preprints202002.0281.v1   2020.2

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    Matrix metalloproteinase 3 (MMP3) plays multiple roles in pro-tumorigenic proteolysis and in intracellular transcription. These include inducing connective tissue growth factor [CTGF, also known as cellular communication network factor 2 (CCN2)] and prompting a new definition of MMP3 as a moonlighting metalloproteinase. Members of the MMP family have been found within tumor-derived extracellular vesicles (EVs) such as oncosomes or exosomes. We here investigated the roles of MMP3-rich oncosomes in tumor progression, molecular transmission, and gene regulation. MMP3 and CCN2/CTGF were significantly co-expressed in tumor samples derived from patients suffering from colorectal adenocarcinoma. We found that oncosomes derived from a rapidly metastatic colon cancer cells (LuM1) were enriched in MMP3 and a C-terminal half fragment of CCN2/CTGF. MMP3-rich oncosomes were highly transmissive into recipient cells and were pro-tumorigenic in an allograft mouse model. Oncosome-derived MMP3 was transmissive into recipient cell nuclei, trans-activated CCN2/CTGF promoter, and induced CCN2/CTGF production at 1 to 6 hours after the addition of oncosomes to culture media. In addition, CRISPR/Cas9-mediated knockout of MMP3 showed significant anti-tumor effects, including inhibition of migration and invasion of LuM1 cells in vitro, inhibition of tumor growth in vivo, and reduction of CCN2/CTGF and its promoter activity in vitro. These data newly demonstrate that the oncosome-derived moonlighting metalloproteinase promotes metastasis and is pro-tumorigenic at distant sites as well as a transmissive trans-activator for the cellular communication network gene.

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  • CDC37 and HSP90 are Essential for Stressome Release and Tumor Progression in Resistant Prostate Cancer

    Takanori Eguchi, Chiharu Sogawa, Kisho Ono, Masaki Matsumoto, Manh Tien Tran, Yuka Okusha, Benjamin Lang, Kuniaki Okamoto, Stuart Calderwood

    Preprints   doi: 10.20944/preprints202002.0148.v1   2020.2

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    Tumor cells exhibit a resistance-associated secretory phenotype involving extracellular vesicles (EVs) and heat shock proteins (HSPs). This response occurs in response to cell stress and cancer therapeutics. HSPs are stress-responsive molecular chaperones promoting proper protein folding, while also being released from cells with EVs as well as in free form as alarmins. We have here investigated the secretory phenotype of castration-resistant prostate cancer (CRPC) cells using proteome analysis. We have also examined the roles of the key co-chaperone CDC37 in stressome release, epithelial-to-mesenchymal transition (EMT), and tumor progression. A number of HSP family members and their common receptor CD91/LRP1 were enriched at high levels in CRPC cell-derived EVs among over 700 other protein species. The small EVs (30 to 200 nm in size, potentially exosomes) were released even in a non-heated condition from the prostate cancer cells, whereas EMT-coupled release of EVs (200 to 500 nm, likely ectosomes) with associated HSP90α was increased after heat shock stress (HSS). Lactate dehydrogenase, a marker of membrane leakage/damage of cells, was also released upon HSS from the prostate cancer cells. During this stress response, intracellular CDC37 was also transcriptionally inducible by heat shock factor 1, and knockdown of CDC37 decreased EMT-coupled release of EVs. Triple knockdown of CDC37, HSP90α, and HSP90β was required for efficient reduction of the chaperone trio and to reduce tumorigenicity of the CRPC cells in vivo. Taken together, the data indicated that CDC37 and HSP90 are essential for stressome release and for tumorigenesis in resistant cancer.

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  • Exosome Release of Drugs: Coupling with Epithelial-Mesenchymal Transition

    Takanori Eguchi, Kisho Ono, Stuart Calderwood, Kuniaki Okamoto

    Preprints   doi: 10.20944/preprints201912.0386.v1   2019.12

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    Extracellular vesicles (EVs), such as exosomes or oncosomes are released with molecules unfavorable for survival from cells. In addition, accumulating evidence has shown that tumor cells often eject anti-cancer drugs such as chemotherapeutics and targeted drugs within EVs, a novel mechanism of drug resistance. The EV-releasing, drug resistance phenotype is often coupled with cellular dedifferentiation and transformation, cells undergoing epithelial-mesenchymal transition (EMT) and taking on a cancer stem cell phenotype. Recent studies have shown that the release of EVs is also involved in immunosuppression. The concept of the resistance-associated secretory phenotype (RASP) is reviewed herein.

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  • Exosome Release of Drugs: Coupling with Epithelial-Mesenchymal Transition

    Takanori Eguchi, Kisho Ono, Stuart Calderwood, Kuniaki Okamoto

    Preprints   2019.12

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    Extracellular vesicles (EVs), such as exosomes or oncosomes are released with molecules unfavorable for survival from cells. In addition, accumulating evidence has shown that tumor cells often eject anti-cancer drugs such as chemotherapeutics and targeted drugs within EVs, a novel mechanism of drug resistance. The EV-releasing, drug resistance phenotype is often coupled with cellular dedifferentiation and transformation, cells undergoing epithelial-mesenchymal transition (EMT) and taking on a cancer stem cell phenotype. Recent studies have shown that the release of EVs is also involved in immunosuppression. The concept of the resistance-associated secretory phenotype (RASP) is reviewed herein.

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  • オルガノイドを応用したドラッグリポジショニング開発

    十川 千春, 江口 傑徳, Tran Tien Manh, 石毛 真行, 河合 穂高, 奥舎 有加, 中野 敬介, 十川 紀夫, 小崎 健一, 岡元 邦彰

    岡山歯学会雑誌   38 ( 2 )   89 - 89   2019.12

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  • Regulatory Roles of HSP90-Rich Extracellular Vesicles

    Takanori Eguchi, Kisho Ono, Kazumi Kawata, Kuniaki Okamoto, Stuart K. Calderwood

    Heat Shock Proteins   3 - 17   2019.11

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    DOI: 10.1007/978-3-030-23158-3_1

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  • A Reporter System Evaluates Tumorigenesis, Metastasis, β-catenin/MMP Regulation, and Druggability. Invited Reviewed International journal

    Chiharu Sogawa, Takanori Eguchi, Yuka Okusha, Kisho Ono, Kazumi Ohyama, Motoharu Iizuka, Ryu Kawasaki, Yusaku Hamada, Masaharu Takigawa, Norio Sogawa, Kuniaki Okamoto, Ken-Ichi Kozaki

    Tissue engineering. Part A   25 ( 19-20 )   1413 - 1425   2019.10

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    Cancer invasion, metastasis, and therapy resistance are the crucial phenomena in cancer malignancy. The high expression of matrix metalloproteinase 9 (MMP9) is a biomarker as well as a causal factor of cancer invasiveness and metastatic activity. However, a regulatory mechanism underlying MMP9 expression in cancer is not clarified yet. In addition, a new strategy for anticancer drug discovery is becoming an important clue. In the present study, we aimed (i) to develop a novel reporter system evaluating tumorigenesis, invasiveness, metastasis, and druggability with a combination of three-dimensional tumoroid model and Mmp9 promoter and (ii) to examine pharmacological actions of anticancer medications using this reporter system. High expression and genetic amplification of MMP9 were found in colon cancer cases. We found that proximal promoter sequences of MMP9 in murine and human contained conserved binding sites for transcription factors β-catenin/TCF/LEF, glucocorticoid receptor (GR), and nuclear factor kappa-B (NF-κB). The murine Mmp9 promoter (-569 to +19) was markedly activated in metastatic colon cancer cells and additionally activated by tumoroid formation and by β-catenin signaling stimulator lithium chloride. The Mmp9 promoter-driven fluorescent reporter cells enabled the monitoring of activities of MMP9/gelatinase, tumorigenesis, invasion, and metastasis in syngeneic transplantation experiments. We also demonstrated pharmacological actions as follows: dexamethasone and hydrocortisone, steroidal medications binding to GR, inhibited the Mmp9 promoter but did not inhibit tumorigenesis. On the contrary, antimetabolite 5-fluorouracil, a gold standard for colon cancer chemotherapy, inhibited tumoroid formation but did not inhibit Mmp9 promoter activity. Notably, antimalaria medication artesunate inhibited both tumorigenesis and the Mmp9 promoter in vitro, potentially through inhibition of β-catenin/TCF/LEF signaling. Thus, this novel reporter system enabled monitoring tumorigenesis, invasiveness, metastasis, key regulatory signalings such as β-catenin/MMP9 axis, and druggability. Impact Statement Cancer invasion and metastasis have been shown to be driven by matrix metalloproteinase 9 (MMP9), whose expression mechanism is not clarified yet. In addition, a new strategy for anticancer drug discovery is becoming important. We established a novel reporter system evaluating tumorigenesis, invasiveness, metastasis, and druggability with a combination of three-dimensional (3D) tumoroid model and Mmp9 promoter. Using this reporter system, we demonstrated pharmacological actions of anticancer medications such as antimetabolite 5-fluorouracil (5-FU) and antimalaria medication artesunate (ART), which inhibited both tumorigenesis and β-catenin/MMP regulatory signaling. Our study impacts the translational fields of oncology, drug discovery, and organoid model.

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  • Tumor Angiogenic Inhibition Triggered Necrosis (TAITN) in Oral Cancer. Reviewed International journal

    Saori Yoshida, Hotaka Kawai, Takanori Eguchi, Shintaro Sukegawa, May Wathone Oo, Chang Anqi, Kiyofumi Takabatake, Keisuke Nakano, Kuniaki Okamoto, Hitoshi Nagatsuka

    Cells   8 ( 7 )   761   2019.7

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    CXCR4 is a chemokine receptor crucial in tumor progression, although the angiogenic role of CXCR4 in oral squamous cell carcinoma (OSCC) has not been investigated. Here we show that CXCR4 is crucial for tumor angiogenesis, thereby supporting tumor survival in OSCC. Immunohistochemistry on human clinical specimens revealed that CXCR4 and a tumor vasculature marker CD34 were co-distributed in tumor vessels in human OSCC specimens. To uncover the effects of CXCR4 inhibition, we treated the OSCC-xenografted mice with AMD3100, so-called plerixafor, an antagonist of CXCR4. Notably, we found a unique pathophysiological structure defined as tumor angiogenic inhibition triggered necrosis (TAITN), which was induced by the CXCR4 antagonism. Treatment with AMD3100 increased necrotic areas with the induction of hypoxia-inducible factor-1α in the xenografted tumors, suggesting that AMD3100-induced TAITN was involved in hypoxia and ischemia. Taken together, we demonstrated that CXCR4 plays a crucial role in tumor angiogenesis required for OSCC progression, whereas TAITN induced by CXCR4 antagonism could be an effective anti-angiogenic therapeutic strategy in OSCC treatment.

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  • MZF1 and SCAND1 Reciprocally Regulate CDC37 Gene Expression in Prostate Cancer. Reviewed International journal

    Takanori Eguchi, Thomas L Prince, Manh Tien Tran, Chiharu Sogawa, Benjamin J Lang, Stuart K Calderwood

    Cancers   11 ( 6 )   792   2019.6

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    Cell division control 37 (CDC37) increases the stability of heat shock protein 90 (HSP90) client proteins and is thus essential for numerous intracellular oncogenic signaling pathways, playing a key role in prostate oncogenesis. Notably, elevated expression of CDC37 was found in prostate cancer cells, although the regulatory mechanisms through which CDC37 expression becomes increased are unknown. Here we show both positive and negative regulation of CDC37 gene transcription by two members of the SREZBP-CTfin51-AW1-Number 18 cDNA (SCAN) transcription factor family-MZF1 and SCAND1, respectively. Consensus DNA-binding motifs for myeloid zinc finger 1 (MZF1/ZSCAN6) were abundant in the CDC37 promoter region. MZF1 became bound to these regulatory sites and trans-activated the CDC37 gene whereas MZF1 depletion decreased CDC37 transcription and reduced the tumorigenesis of prostate cancer cells. On the other hand, SCAND1, a zinc fingerless SCAN box protein that potentially inhibits MZF1, accumulated at MZF1-binding sites in the CDC37 gene, negatively regulated the CDC37 gene and inhibited tumorigenesis. SCAND1 was abundantly expressed in normal prostate cells but was reduced in prostate cancer cells, suggesting a potential tumor suppressor role of SCAND1 in prostate cancer. These findings indicate that CDC37, a crucial protein in prostate cancer progression, is regulated reciprocally by MZF1 and SCAND1.

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  • Nicotine promotes lymph node metastasis and cetuximab resistance in head and neck squamous cell carcinoma. Reviewed International journal

    Rieko Shimizu, Soichiro Ibaragi, Takanori Eguchi, Daisuke Kuwajima, Shinichi Kodama, Takashi Nishioka, Tatsuo Okui, Kyoichi Obata, Kiyofumi Takabatake, Hotaka Kawai, Kisho Ono, Kuniaki Okamoto, Hitoshi Nagatsuka, Akira Sasaki

    International journal of oncology   54 ( 1 )   283 - 294   2019.1

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    Epidermal growth factor (EGF) is overexpressed in many cancers and is associated with worse prognosis. EGF binds to its cell surface receptor (EGFR), which induces EGFR phosphorylation. Phosphorylated EGFR (p‑EGFR) is translocated into the nucleus, which increases cancer cell activity. Nicotine, which is one of the main components of tobacco, is absorbed through pulmonary alveoli and mucosal epithelia in the head and neck region by smoking and moves into the blood. Nicotine in blood binds to nicotinic acetylcholine receptor (nAChR) in the central nervous system and serves a crucial role in tobacco addiction. Although nAChR localization is thought to be limited in the nervous system, nAChR is present in a wide variety of non‑neuronal cells, including cancer cells. Recent studies suggest that nicotine contributes to the metastasis and resistance to anti‑cancer drugs of various cancer cells. However, it remains unknown whether head and neck squamous cell carcinoma (HNSCC) cells can utilize nicotine‑nAChR signaling to metastasize and acquire resistance to anti‑cancer drugs, even though the mucosal epithelia of the head and neck region are the primary sites of exposure to tobacco smoke. To the best of our knowledge, the present study is the first to demonstrate the role of nicotine in metastasis and anti‑EGFR‑therapy resistance of HNSCC. The present findings demonstrated that nicotine increased proliferation, migration, invasion, p‑EGFR nuclear translocation and protein kinase B (Akt) phosphorylation in HNSCC cells. It was also demonstrated that nicotine restored cetuximab‑inhibited proliferation, migration and invasion of HNSCC cells. Finally, an in vivo experiment revealed that nicotine increased lymph node metastasis of xenografted tumors, whereas an nAChR inhibitor suppressed lymph node metastasis and p‑EGFR nuclear localization of xenografted tumors. Taken together, these results demonstrated that nicotine induced nuclear accumulation of p‑EGFR, and activation of Akt signaling. These signaling pathways elevated the activities of HNSCC cells, causing lymph node metastasis and serving a role in cetuximab resistance.

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  • Carcinogenic epithelial-mesenchymal transition initiated by oral cancer exosomes is inhibited by anti-EGFR antibody cetuximab. Reviewed International journal

    Toshifumi Fujiwara, Takanori Eguchi, Chiharu Sogawa, Kisho Ono, Jun Murakami, Soichiro Ibaragi, Jun-Ichi Asaumi, Stuart K Calderwood, Kuniaki Okamoto, Ken-Ichi Kozaki

    Oral oncology   86   251 - 257   2018.11

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    Overexpression and increased signaling from the epidermal growth factor receptor (EGFR) often changes oral squamous cell carcinoma (OSCC) and thus EGFR is frequently targeted molecularly by the therapeutic antibody cetuximab. We assessed the roles of OSCC-derived extracellular vesicles (EVs), including exosomes in the trafficking of cetuximab and in epithelial-mesenchymal transition (EMT) of epithelial cells. OSCC cells abundantly expressed EGFR, which was secreted from cells with OSCC-EVs upon EGF stimulations. The OSCC-EGFR-EVs were then able to enter into and transform epithelial cells leading to increased mesenchymal traits with increased vimentin and spindle-like shapes. EGF priming of OSCC cells further increased this EMT-initiating effect of the OSCC-EVs. The internalization and pro-EMT effects of the OSCC-EVs were largely blocked by cetuximab. Thus, OSCC-derived EVs transform normal epithelial cells into a mesenchymal phenotype and anti-EGFR therapeutic antibody cetuximab inhibits such a carcinogenic effect of the OSCC-EVs.

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  • Lipoprotein Cholesterols Are Stored in High-Resistant, Metastatic Cancer Cells and Released Upon Stress: Implication for a Mechanism Underlying Hypocholesterolemia in Cancer Patients

    Taka Eguchi, Chiharu Sogawa, Kisho Ono, Mami Itagaki, Masaki Matsumoto

    Preprints   2018.10

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    Resistant cancer often shows a particular secretory trait such as heat shock proteins (HSPs) and extracellular vesicles (EVs), including exosomes and oncosomes surrounded by lipid bilayers. Lipoproteins are biochemical assemblies that transport hydrophobic lipid (a.k.a. fat) molecules in body fluid and are composed of a single-layer phospholipid and cholesterol outer shell, lipids molecules within the particles, and apolipoproteins embedded in the membrane. However, lipoprotein storage and secretion by cancer cells have not well-investigated yet. We found lipoproteins were stored and abundantly secreted by neuroendocrine, castration-resistant prostate cancer (NEPC / CRPC) cells but barely secreted by colon cancer cells and oral squamous cell carcinoma (OSCC) cells. In addition, large EVs (approx. 300 nm diameter) and potential oncosomes were released by CRPC and OSCC cells. Proteomics revealed that CRPC cells secreted EVs enriched with tetraspanins and extracellular matrices which were reduced upon heat shock stress and alternatively lipoproteins and HSPs were secreted upon stress. Heat shock stress triggered secretion of lipoprotein-EV complexes that contained apolipoprotein A, B, C and E. These data suggested that vesicular assembly composed of EVs and lipoproteins enriched with cholesterols and phospholipids may be stored in resistant cancer cells but released upon cell stress that is increased in cancer therapies.

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  • 侵襲性歯周炎患者の血漿エクソソーム由来microRNAの発現解析

    高木 美奈, 山本 直史, 河村 麻理, 高知 信介, 山城 圭介, 大森 一弘, 江口 傑徳, 十川 千春, 高柴 正悟

    日本歯周病学会会誌   60 ( 秋季特別 )   134 - 134   2018.10

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  • Anti-EGFR antibody cetuximab is secreted by oral squamous cell carcinoma and alters EGF-driven mesenchymal transition. Reviewed International journal

    Toshifumi Fujiwara, Takanori Eguchi, Chiharu Sogawa, Kisho Ono, Jun Murakami, Soichiro Ibaragi, Jun-Ichi Asaumi, Kuniaki Okamoto, Stuart K Calderwood, Ken-Ichi Kozaki

    Biochemical and biophysical research communications   503 ( 3 )   1267 - 1272   2018.9

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    Genetic amplification, overexpression, and increased signaling from the epidermal growth factor receptor (EGFR) are often found in oral squamous cell carcinoma (OSCC) and thus EGFR is frequently targeted molecularly by the therapeutic antibody cetuximab. We assessed effects of cetuximab in control of EGF-driven malignant traits of OSCC cells. EGF stimulation promoted progression level of mesenchymal traits in OSCC cells, which were attenuated by cetuximab but incompletely. We pursued a potential mechanism underlying such incomplete attenuation of OSCC malignant traits. Cetuximab promoted secretion of EGFR-EVs by OSCC cells and failed to inhibit EGF-driven secretion of EGFR-EVs. Cetuximab was also found to be robustly secreted with the EGFR-EVs by the OSCC cells. Thus, EGF promotes the level of mesenchymal traits of OSCC cells and secretion of EGFR-EVs, which involve cetuximab resistance.

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  • The intranuclear PEX domain of MMP involves proliferation, migration, and metastasis of aggressive adenocarcinoma cells. Reviewed International journal

    Yuka Okusha, Takanori Eguchi, Chiharu Sogawa, Tatsuo Okui, Keisuke Nakano, Kuniaki Okamoto, Ken-Ichi Kozaki

    Journal of cellular biochemistry   119 ( 9 )   7363 - 7376   2018.9

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    Members of matrix metalloproteinase (MMP) family promote cancer cell migration, invasion, and metastasis through alteration of the tumor milieu, intracellular signaling pathways, and transcription. We examined gene expression signatures of colon adenocarcinoma cell lines with different metastatic potentials and found that rapidly metastatic cells powerfully expressed genes encoding MMP3 and MMP9. The non-proteolytic PEX isoform and proteolytic isoforms of MMPs were significantly expressed in the metastatic cells in vitro. Knockdown of MMP3 attenuated cancer cell migration and invasion in vitro and lung metastasis in vivo. Profound nuclear localization of MMP3/PEX was found in tumor-stroma marginal area. In contrast, MMP9 was localized in central area of subcutaneous tumors. Overexpression of the PEX isoform of MMP3 promoted proliferation and migration of the rapidly metastatic cells in vitro. Taken together, the non-proteolytic PEX isoform of MMPs locating in cell nuclei involves proliferation, migration, and subsequent metastasis of aggressive adenocarcinoma cells.

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  • エクソソーム研究の「がん」における新展開 細胞外小胞のHSPに富む特性は、転移性口腔癌細胞の生存を担う(HSP-enriched properties of extracellular vesicles involve survival of metastatic oral cancer cells)

    小野 喜章, 江口 傑徳, 十川 千春, 佐々木 朗

    日本癌学会総会記事   77回   897 - 897   2018.9

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  • 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, 小野 喜章, 岡元 邦彰

    Journal of Oral Biosciences Supplement   2018   347 - 347   2018.9

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  • 大腸癌の増殖,浸潤,転移を促進するPEXドメインの役割とその制御(The intranuclear PEX domain of MMP involves proliferation, migration, and metastasis of aggressive adenocarcinoma cells)

    江口 傑徳, 奥舎 有加, 小野 喜章, 十川 千春, Calderwood Stuart K., 小崎 健一

    日本癌学会総会記事   77回   1332 - 1332   2018.9

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  • Genotoxic stress induces Sca-1-expressing metastatic mammary cancer cells. Reviewed International journal

    Jianlin Gong, Benjamin J Lang, Desheng Weng, Takanori Eguchi, Ayesha Murshid, Thiago J Borges, Sachin Doshi, Baizheng Song, Mary A Stevenson, Stuart K Calderwood

    Molecular oncology   12 ( 8 )   1249 - 1263   2018.8

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    We describe a cell damage-induced phenotype in mammary carcinoma cells involving acquisition of enhanced migratory and metastatic properties. Induction of this state by radiation required increased activity of the Ptgs2 gene product cyclooxygenase 2 (Cox2), secretion of its bioactive lipid product prostaglandin E2 (PGE2), and the activity of the PGE2 receptor EP4. Although largely transient, decaying to low levels in a few days to a week, this phenotype was cumulative with damage and levels of cell markers Sca-1 and ALDH1 increased with treatment dose. The Sca-1+ , metastatic phenotype was inhibited by both Cox2 inhibitors and PGE2 receptor antagonists, suggesting novel approaches to radiosensitization.

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  • Depletion of Lipid Efflux Pump ABCG1 Triggers the Intracellular Accumulation of Extracellular Vesicles and Reduces Aggregation and Tumorigenesis of Metastatic Cancer Cells. Reviewed International journal

    Yuri Namba, Chiharu Sogawa, Yuka Okusha, Hotaka Kawai, Mami Itagaki, Kisho Ono, Jun Murakami, Eriko Aoyama, Kazumi Ohyama, Jun-Ichi Asaumi, Masaharu Takigawa, Kuniaki Okamoto, Stuart K Calderwood, Ken-Ichi Kozaki, Takanori Eguchi

    Frontiers in oncology   8 ( 376 )   376 - 376   2018

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    The ATP-binding cassette transporter G1 (ABCG1) is a cholesterol lipid efflux pump whose role in tumor growth has been largely unknown. Our transcriptomics revealed that ABCG1 was powerfully expressed in rapidly metastatic, aggregative colon cancer cells, in all the ABC transporter family members. Coincidently, genetic amplification of ABCG1 is found in 10-35% of clinical samples of metastatic cancer cases. Expression of ABCG1 was further elevated in three-dimensional tumoroids (tumor organoids) within stemness-enhancing tumor milieu, whereas depletion of ABCG1 lowered cellular aggregation and tumoroid growth in vitro as well as hypoxia-inducible factor 1α in cancer cells around the central necrotic areas in tumors in vivo. Notably, depletion of ABCG1 triggered the intracellular accumulation of extracellular vesicles (EVs) and regression of tumoroids. Collectively, these data suggest that ABCG1 plays a crucial role in tumorigenesis in metastatic cancer and that depletion of ABCG1 triggers tumor regression with the accumulation of EVs and their derivatives and cargos, implicating a novel ABCG1-targeting therapeutic strategy by which redundant and toxic substances may be accumulated in tumors leading to their regression.

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  • Monitoring of the Heat Shock Response with a Real-Time Luciferase Reporter

    Toshiki Kijima, Takanori Eguchi, Len Neckers, Thomas L. Prince

    Methods in Molecular Biology   35 - 45   2018

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  • 異なる転移能を有する口腔扁平上皮癌細胞由来エクソソームに含まれるプロテオームの特性

    小野 喜章, 江口 傑徳, 十川 千春, 村上 純, 藤原 敏史, 笠井 智成, 妹尾 昌治, 佐々木 朗, 小崎 健一, 岡元 邦彰

    生命科学系学会合同年次大会   2017年度   [1LBA - 052]   2017.12

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  • A Tumor Suppressor Gene Product, Platelet-Derived Growth Factor Receptor-Like Protein Controls Chondrocyte Proliferation and Differentiation. Reviewed International journal

    Kazumi Kawata, Satoshi Kubota, Takanori Eguchi, Eriko Aoyama, Norifumi H Moritani, Morihiko Oka, Harumi Kawaki, Masaharu Takigawa

    Journal of cellular biochemistry   118 ( 11 )   4033 - 4044   2017.11

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    The platelet-derived growth factor receptor-like (PDGFRL) gene is regarded as a tumor suppressor gene. However, nothing is known about the molecular function of PDGFRL. In this study, we initially clarified its function in chondrocytes. Among all cell lines examined, the PDGFRL mRNA level was the highest in chondrocytic HCS-2/8 cells. Interestingly, the proliferation of chondrocytic HCS-2/8 cells was promoted by PDGFRL overexpression, whereas that of the breast cancer-derived MDA-MB-231 cells was inhibited. Of note, in PDGFRL-overexpressing HCS-2/8 cells, the expression of chondrocyte differentiation marker genes, SOX9, ACAN, COL2A1, COL10A1, and ALP, was decreased. Moreover, we confirmed the expression of PDGFRL mRNA in normal cartilage tissue and chondrocytes. Eventually, the expression of PDGFRL mRNA in condrocytes except in the case of hypertrophic chondrocytes was demonstrated in vivo and in vitro. These findings suggest that PDGFRL plays the different roles, depending upon cell types. Particularly, in chondrocytes, PDGFRL may play a new and important role which is distinct from the function previously reported. J. Cell. Biochem. 118: 4033-4044, 2017. © 2017 Wiley Periodicals, Inc.

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  • Catabolic effects of FGF-1 on chondrocytes and its possible role in osteoarthritis. Reviewed International journal

    Abdellatif El-Seoudi, Tarek Abd El Kader, Takashi Nishida, Takanori Eguchi, Eriko Aoyama, Masaharu Takigawa, Satoshi Kubota

    Journal of cell communication and signaling   11 ( 3 )   255 - 263   2017.9

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    Fibroblast growth factor 1 (FGF-1) is a classical member of the FGF family and is produced by chondrocytes cultured from osteoarthritic patients. Also, this growth factor was shown to bind to CCN family protein 2 (CCN2), which regenerates damaged articular cartilage and counteracts osteoarthritis (OA) in an animal model. However, the pathophysiological role of FGF-1 in cartilage has not been well investigated. In this study, we evaluated the effects of FGF-1 in vitro and its production in vivo by use of an OA model. Treatment of human chondrocytic cells with FGF-1 resulted in marked repression of genes for cartilaginous extracellular matrix components, whereas it strongly induced matrix metalloproteinase 13 (MMP-13), representing its catabolic effects on cartilage. Interestingly, expression of the CCN2 gene was dramatically repressed by FGF-1, which repression eventually caused the reduced production of CCN2 protein from the chondrocytic cells. The results of a reporter gene assay revealed that this repression could be ascribed, at least in part, to transcriptional regulation. In contrast, the gene expression of FGF-1 was enhanced by exogenous FGF-1, indicating a positive feedback system in these cells. Of note, induction of FGF-1 was observed in the articular cartilage of a rat OA model. These results collectively indicate a pathological role of FGF-1 in OA development, which includes an insufficient cartilage regeneration response caused by CCN2 down regulation.

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  • 舌癌細胞株由来エクソソーム解析による頸部リンパ節転移マーカーの探索

    小野 喜章, 江口 傑徳, 十川 千春, 村上 純, 笠井 智成, 妹尾 昌治, 佐々木 朗, 小崎 健一

    日本癌学会総会記事   76回   J - 1062   2017.9

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  • Semaphorin 4D promotes bone invasion in head and neck squamous cell carcinoma. Reviewed International journal

    Hiroyuki Takada, Soichiro Ibaragi, Takanori Eguchi, Tatsuo Okui, Kyoichi Obata, Masanori Masui, Ayaka Morisawa, Kiyofumi Takabatake, Hotaka Kawai, Norie Yoshioka, Nur Mohammad Monsur Hassan, Tsuyoshi Shimo, Guo-Fu Hu, Hitoshi Nagatsuka, Akira Sasaki

    International journal of oncology   51 ( 2 )   625 - 632   2017.8

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    Head and neck squamous cell carcinomas (HNSCCs) frequently invade the bones of the facial skeleton. Semaphorin 4D (Sema4D) is an axon guidance molecule produced by oligodendrocytes. Sema4D was also identified in the bone microenvironment and many cancer tissues including HNSCC. To date, however, the role of Sema4D in cancer-associated bone disease is still unknown. This is the first study to demonstrate the role of Sema4D in bone invasion of cancer. In the clinical tissue samples of bone lesion of HNSCC, Sema4D was detected at high levels, and its expression was correlated with insulin-like growth factor-I (IGF-I) expression. In vitro experiments showed that IGF-I regulates Sema4D expression and Sema4D increased proliferation, migration and invasion in HNSCC cells. Sema4D also regulated the expression of receptor activator of nuclear factor κβ ligand (RANKL) in osteoblasts, and this stimulated osteoclastgenesis. Furthermore, knockdown of Sema4D in HNSCC cells inhibited tumor growth and decreased the number of osteoclasts in a mouse xenograft model. Taken together, IGF-I-driven production of Sema4D in HNSCCs promotes osteoclastogenesis and bone invasion.

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  • Promoter Analyses of CCN Genes

    Takanori Eguchi, Satoshi Kubota, Masaharu Takigawa

    Methods in Molecular Biology   177 - 185   2017

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  • Cellular Reprogramming Using Defined Factors and MicroRNAs. Reviewed International journal

    Takanori Eguchi, Takuo Kuboki

    Stem cells international   2016   7530942 - 7530942   2016

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    Development of human bodies, organs, and tissues contains numerous steps of cellular differentiation including an initial zygote, embryonic stem (ES) cells, three germ layers, and multiple expertized lineages of cells. Induced pluripotent stem (iPS) cells have been recently developed using defined reprogramming factors such as Nanog, Klf5, Oct3/4 (Pou5f1), Sox2, and Myc. This outstanding innovation is largely changing life science and medicine. Methods of direct reprogramming of cells into myocytes, neurons, chondrocytes, and osteoblasts have been further developed using modified combination of factors such as N-myc, L-myc, Sox9, and microRNAs in defined cell/tissue culture conditions. Mesenchymal stem cells (MSCs) and dental pulp stem cells (DPSCs) are also emerging multipotent stem cells with particular microRNA expression signatures. It was shown that miRNA-720 had a role in cellular reprogramming through targeting the pluripotency factor Nanog and induction of DNA methyltransferases (DNMTs). This review reports histories, topics, and idea of cellular reprogramming.

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  • A Novel High-Throughput 3D Screening System for EMT Inhibitors: A Pilot Screening Discovered the EMT Inhibitory Activity of CDK2 Inhibitor SU9516. Reviewed International journal

    Kazuya Arai, Takanori Eguchi, M Mamunur Rahman, Ruriko Sakamoto, Norio Masuda, Tetsuya Nakatsura, Stuart K Calderwood, Ken-Ichi Kozaki, Manabu Itoh

    PloS one   11 ( 9 )   e0162394   2016

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    Epithelial-mesenchymal transition (EMT) is a crucial pathological event in cancer, particularly in tumor cell budding and metastasis. Therefore, control of EMT can represent a novel therapeutic strategy in cancer. Here, we introduce an innovative three-dimensional (3D) high-throughput screening (HTS) system that leads to an identification of EMT inhibitors. For the establishment of the novel 3D-HTS system, we chose NanoCulture Plates (NCP) that provided a gel-free micro-patterned scaffold for cells and were independent of other spheroid formation systems using soft-agar. In the NCP-based 3D cell culture system, A549 lung cancer cells migrated, gathered, and then formed multiple spheroids within 7 days. Live cell imaging experiments showed that an established EMT-inducer TGF-β promoted peripheral cells around the core of spheroids to acquire mesenchymal spindle shapes, loss of intercellular adhesion, and migration from the spheroids. Along with such morphological change, EMT-related gene expression signatures were altered, particularly alteration of mRNA levels of ECAD/CDH1, NCAD/CDH2, VIM and ZEB1/TCF8. These EMT-related phenotypic changes were blocked by SB431542, a TGF-βreceptor I (TGFβR1) inhibitor. Inside of the spheroids were highly hypoxic; in contrast, spheroid-derived peripheral migrating cells were normoxic, revealed by visualization and quantification using Hypoxia Probe. Thus, TGF-β-triggered EMT caused spheroid hypoplasia and loss of hypoxia. Spheroid EMT inhibitory (SEMTIN) activity of SB431542 was calculated from fluorescence intensities of the Hypoxia Probe, and then was utilized in a drug screening of EMT-inhibitory small molecule compounds. In a pilot screening, 9 of 1,330 compounds were above the thresholds of the SEMTIN activity and cell viability. Finally, two compounds SB-525334 and SU9516 showed SEMTIN activities in a dose dependent manner. SB-525334 was a known TGFβR1 inhibitor. SU9516 was a cyclin-dependent kinase 2 (CDK2) inhibitor, which we showed also had an EMT-inhibitory activity. The half maximal inhibitory concentration (IC50) of SB-525334 and SU9516 were 0.31 μM and 1.21 μM, respectively, while IC50 of SB431542 was 2.38 μM. Taken together, it was shown that this 3D NCP-based HTS system was useful for screening of EMT-regulatory drugs.

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  • Targeting the hsp70 gene delays mammary tumor initiation and inhibits tumor cell metastasis Reviewed

    J. Gong, D. Weng, T. Eguchi, A. Murshid, M. Y. Sherman, B. Song, S. K. Calderwood

    ONCOGENE   34 ( 43 )   5460 - 5471   2015.10

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    Elevated levels of the inducible heat-shock protein 70 (Hsp72) have been implicated in mammary tumorigenesis in histological investigations of human breast cancer. We therefore examined the role of Hsp72 in mice, using animals in which the hsp70 gene was inactivated. We used a spontaneous tumor system with mice expressing the polyomavirus middle T (PyMT) oncogene under control of the mouse mammary tumor virus (MMTV) long-terminal repeat (MMT mice). These mice developed spontaneous, metastatic mammary cancer. We then showed Hsp72 to be upregulated in a fraction of mammary cancer initiating cells (CIC) within the MMT tumor cell population. These cells were characterized by elevated surface levels of stem cell markers CD44 and Sca1 and by rapid metastasis. Inactivation of the hsp70 gene delayed the initiation of mammary tumors. This delay in tumor initiation imposed by loss of hsp70 was correlated with a decreased pool of CIC. Interestingly, hsp70 knockout significantly reduced invasion and metastasis by mammary tumor cells and implicated its product Hsp72 in cell migration and formation of secondary neoplasms. Impaired tumorigenesis and metastasis in hsp70-knockout MMT mice was associated with downregulation of the met gene and reduced activition of the oncogenic c-Met protein. These experiments therefore showed Hsp72 to be involved in the growth and progression of mammary carcinoma and highlighted this protein as a potential target for anticancer drug development.

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  • Role and Regulation of Myeloid Zinc Finger Protein 1 in Cancer. Reviewed International journal

    Taka Eguchi, Thomas Prince, Barbara Wegiel, Stuart K Calderwood

    Journal of cellular biochemistry   116 ( 10 )   2146 - 54   2015.10

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    Myeloid zinc finger 1 (MZF1) belongs to the SCAN-Zinc Finger (SCAN-ZF) transcription factor family that has recently been implicated in a number of types of cancer. Although the initial studies concentrated on the role of MZF1 in myeloid differentiation and leukemia, the factor now appears to be involved in the etiology of major solid tumors such as lung, cervical, breast, and colorectal cancer. Here we discuss the regulation of MZF1 that mediated its recruitment and activation in cancer, concentrating on posttranslational modification by phosphorylation, and sumoylation, formation of promyelocytic leukemia nuclear bodies and its association with co-activators and co-repressors.

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  • HSF1 regulation of β-catenin in mammary cancer cells through control of HuR/elavL1 expression. Reviewed International journal

    Shiuh-Dih Chou, Ayesha Murshid, Takanori Eguchi, Jianlin Gong, Stuart K Calderwood

    Oncogene   34 ( 17 )   2178 - 2188   2015.4

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    There is now compelling evidence to indicate a place for heat shock factor 1 (HSF1) in mammary carcinogenesis, tumour progression and metastasis. Here we have investigated a role for HSF1 in regulating the expression of the stem cell renewal factor β-catenin in immortalized human mammary epithelial and carcinoma cells. We found HSF1 to be involved in regulating the translation of β-catenin, by investigating effects of gain and loss of HSF1 on this protein. Interestingly, although HSF1 is a potent transcription factor, it was not directly involved in regulating levels of β-catenin mRNA. Instead, our data suggest a complex role in translational regulation. HSF1 was shown to regulate levels of the RNA-binding protein HuR that controlled β-catenin translation. An extra complexity was added to this scenario when it was shown that the long non-coding RNA molecule lincRNA-p21, known to be involved in β-catenin mRNA (CTNNB1) translational regulation, was controlled by HSF1 repression. We have shown previously that HSF1 was positively regulated through phosphorylation by mammalian target of rapamycin (mTOR) kinase on a key residue, serine 326, essential for transcriptional activity. In this study, we found that mTOR knockdown not only decreased HSF1-S326 phosphorylation in mammary cells, but also decreased β-catenin expression through a mechanism requiring HuR. Our data point to a complex role for HSF1 in the regulation of HuR and β-catenin expression that may be significant in mammary carcinogenesis.

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  • OstemiR: A Novel Panel of MicroRNA Biomarkers in Osteoblastic and Osteocytic Differentiation from Mesencymal Stem Cells

    Eguchi, Takanori, Hara, Emilio Satoshi, Ono, Mitsuaki, Watanabe, Ken, Kuboki, Takuo, Calderwood, Stuart

    Faseb Journal   29 ( 3 )   e58796   2015

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  • miRNA-720 regulates the stem cell phenotype and differentiation of human dental pulp-derived mesenchymal stromal cells

    Hara, Emilio Satoshi, Ono, Mitsuaki, Eguchi, Takanori, Hai Thanh Pham, Tajima, Shoji, Calderwood, Stuart K., Matsumoto, Takuya, Kuboki, Takuo, IEEE

    2014 International Symposium on Micro-Nanomechatronics and Human Science (Mhs)   1 - 2   2014

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  • Stress proteins in aging and life span. Reviewed International journal

    Ayesha Murshid, Takanori Eguchi, Stuart K Calderwood

    International journal of hyperthermia : the official journal of European Society for Hyperthermic Oncology, North American Hyperthermia Group   29 ( 5 )   442 - 7   2013.8

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    Heat shock proteins (HSP) are molecular chaperones and have been implicated in longevity and aging in many species. Their major functions include chaperoning misfolded or newly synthesised polypeptides, protecting cells from proteotoxic stress, and processing of immunogenic agents. These proteins are expressed constitutively and can be induced by stresses such as heat, oxidative stress and many more. The induction of HSP in aging could potentially maintain protein homeostasis and longevity by refolding the damaged proteins which accumulate during aging and are toxic to cells. HSP are shown to increase life span in model organisms such as Caenorhabditis elegans and decrease aging-related proteotoxicity. Thus, decrease in HSP in aging is associated with disruption of cellular homeostasis which causes diseases such as cancer, cell senescence and neurodegeneration. HSP levels are decreased with aging in most organs including neurons. Aging also causes attenuation or alteration of many signalling pathways as well as the expression of transcription factors such as heat shock factor (HSF). The alteration in regulation and synthesis of Forkhead box O3a (FoxO3a) family of transcription factors as well as major antioxidant enzymes (manganese superoxide dismutase, catalase) are also seen in aging. Among many signalling mechanisms involved in altering longevity and aging, the insulin/IGF-1 pathway and the Sir2 deacetylase are highly significant. This review enquires into the role of some of these pathways in longevity/aging along with HSP.

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  • miRNA-720 controls stem cell phenotype, proliferation and differentiation of human dental pulp cells. Reviewed International journal

    Emilio Satoshi Hara, Mitsuaki Ono, Takanori Eguchi, Satoshi Kubota, Hai Thanh Pham, Wataru Sonoyama, Shoji Tajima, Masaharu Takigawa, Stuart K Calderwood, Takuo Kuboki

    PloS one   8 ( 12 )   e83545   2013

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    Dental pulp cells (DPCs) are known to be enriched in stem/progenitor cells but not well characterized yet. Small non-coding microRNAs (miRNAs) have been identified to control protein translation, mRNA stability and transcription, and have been reported to play important roles in stem cell biology, related to cell reprogramming, maintenance of stemness and regulation of cell differentiation. In order to characterize dental pulp stem/progenitor cells and its mechanism of differentiation, we herein sorted stem-cell-enriched side population (SP) cells from human DPCs and periodontal ligament cells (PDLCs), and performed a locked nucleic acid (LNA)-based miRNA array. As a result, miR-720 was highly expressed in the differentiated main population (MP) cells compared to that in SP cells. In silico analysis and a reporter assay showed that miR-720 targets the stem cell marker NANOG, indicating that miR-720 could promote differentiation of dental pulp stem/progenitor cells by repressing NANOG. Indeed, gain-and loss-of-function analyses showed that miR-720 controls NANOG transcript and protein levels. Moreover, transfection of miR-720 significantly decreased the number of cells positive for the early stem cell marker SSEA-4. Concomitantly, mRNA levels of DNA methyltransferases (DNMTs), which are known to play crucial factors during stem cell differentiation, were also increased by miR-720 through unknown mechanism. Finally, miR-720 decreased DPC proliferation as determined by immunocytochemical analysis against ki-67, and promoted odontogenic differentiation as demonstrated by alizarin red staining, as well as alkaline phosphatase and osteopontin mRNA levels. Our findings identify miR-720 as a novel miRNA regulating the differentiation of DPCs.

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  • Role of LRP1 in transport of CCN2 protein in chondrocytes. Reviewed International journal

    Kazumi Kawata, Satoshi Kubota, Takanori Eguchi, Eriko Aoyama, Norifumi H Moritani, Seiji Kondo, Takashi Nishida, Masaharu Takigawa

    Journal of cell science   125 ( Pt 12 )   2965 - 72   2012.6

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    Low-density lipoprotein receptor-related protein 1 (LRP1) is known to be a receptor for signal transmission and endocytosis. We have previously reported that LRP1 regulates WNT-β-catenin and protein kinase C signaling in chondrocytes, represses the hypertrophy of chondrocytes during endochondral ossification and that LRP1 is colocalized with a ligand, CCN family member 2 (CCN2; also known as connective tissue growth factor, CTGF), which conducts endochondral ossification, in chondrocytes. However, the role of LRP1 in the endocytic transport of CCN2 in chondrocytes is not yet understood. In the present study, we investigated the interaction between LRP1 and CCN2 during endocytic trafficking. Small interfering RNA (siRNA)-mediated knockdown of LRP1 in chondrocytic HCS-2/8 cells showed that the amount of exogenous CCN2 binding and/or incorporation was decreased in the LRP1 downregulated cells. Importantly, we observed that CCN2 internalization in chondrocytes was dependent on clathrin, and internalizated CCN2 was colocalized with an early or recycling endosome marker. Transcytosis of CCN2 through HCS-2/8 cells was confirmed by performing experiments with a trans-well apparatus, and the amount of transcytosed CCN2 was decreased by an LRP1 antagonist. These findings rule out possible leakage and confirm the crucial involvement of LRP1 during experimental transcytosis. Moreover, under hypoxic conditions that mimic the cartilaginous microenvironment, the level of LRP1 and the amount of transcytosed CCN2 increased, and these increases were neutralized by treatment with the LRP1 antagonist. The distribution of LRP1 and its antagonist in the growth plate in vivo was consistent with that of CCN2 in this tissue, which is produced by and transported by LRP1 from the chondrocytes in the prehypertrophic layer. These findings suggest that LRP1 mediates the transcytosis of CCN2, which might be a crucial event that determines the distribution of CCN2 in cartilage.

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  • Role of low-density lipoprotein receptor related protein 1 (LRP1) in CCN2/connective tissue growth factor (CTGF) protein transport in chondrocytes Reviewed

    Kawata., K, Kubota, S, Eguchi, T, Aoyama, E, Moritani, N, Kondo, S, Nishida, T, Takigawa, M

    J Cell Sci   15   2965 - 2972   2012

  • 低密度リポタンパク受容体関連タンパク1(LRP1)による軟骨細胞でのCCNファミリー2/結合組織成長因子(CCN2/CTGF)タンパク質輸送

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 近藤 誠二, 滝川 正春

    日本結合組織学会学術大会・マトリックス研究会大会合同学術集会プログラム・抄録集   43回・58回   76 - 76   2011.5

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  • A coding RNA segment that enhances the ribosomal recruitment of chicken ccn1 mRNA. Reviewed International journal

    Yoshiki Mukudai, Satoshi Kubota, Takanori Eguchi, Kumi Sumiyoshi, Danilo Janune, Seiji Kondo, Satoru Shintani, Masaharu Takigawa

    Journal of cellular biochemistry   111 ( 6 )   1607 - 18   2010.12

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    CCN1, a member of the CCN family of proteins, plays important physiological or pathological roles in a variety of tissues. In the present study, we initially found a highly guanine-cytosine (GC)-rich region of approximately 200 bp near the 5'-end of the open reading frame, which was always truncated by amplification of the corresponding cDNA region through the conventional polymerase chain reaction. An RNA in vitro folding assay and selective ribonuclease digestion of the corresponding segment of the ccn1 mRNA confirmed the involvement of a stable secondary structure. Subsequent RNA electromobility-shift assays demonstrated the specific binding of some cytoplasmic factor(s) in chicken embryo fibroblasts to the RNA segment. Moreover, the corresponding cDNA fragment strongly enhanced the expression of the reporter gene in cis at the 5'-end, but did not do so at the 3'-end. According to the results of a ribosomal assembly test, the effect of the mRNA segment can predominantly be ascribed to the enhancement of transport and/or entry of the mRNA into the ribosome. Finally, the minimal GC-rich mRNA segment that was predicted and demonstrated to form a secondary structure was confirmed to be a functional regulatory element. Thus, we here uncover a novel dual-functionality of the mRNA segment in the ccn1 open reading frame, which segment acts as a cis-element that mediates posttranscriptional gene regulation, while retaining the information for the amino acid sequence of the resultant protein.

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  • 軟骨細胞のCCN2蛋白質輸送における低比重リポ蛋白受容体関連蛋白質1(LRP1)の役割(Role of the low-density lipoprotein receptor related protein 1 (LRP1) in CCN2 protein transportation in chondrocytes)

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 近藤 誠二, 滝川 正春

    日本生化学会大会・日本分子生物学会年会合同大会講演要旨集   83回・33回   2T10 - 12   2010.12

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  • 低密度リポタンパク受容体関連タンパク1(LRP1)による軟骨細胞でのタンパク質輸送

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 近藤 誠二, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   28回   188 - 188   2010.7

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  • Role of the low-density lipoprotein receptor-related protein-1 in regulation of chondrocyte differentiation. Reviewed International journal

    Kazumi Kawata, Satoshi Kubota, Takanori Eguchi, Norifumi H Moritani, Tsuyoshi Shimo, Seiji Kondo, Takashi Nishida, Shogo Minagi, Masaharu Takigawa

    Journal of cellular physiology   222 ( 1 )   138 - 48   2010.1

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    The low-density lipoprotein receptor-related protein 1 (LRP1) is known as an endocytic and signal transmission receptor. We formerly reported the gene expression and the localization of LRP1 in cartilage tissue and chondrocytes, but its roles in the differentiation of chondrocytes remained to be investigated. Here, in order to address this issue, we employed RNAi strategy to knockdown lrp1 in chondrocytic cells and obtained findings indicating a critical role therein. As a result of lrp1 knockdown, aggrecan and col2a1 mRNA levels were decreased. However, that of col10a1 or mmp13 mRNA was rather increased. Under this condition, we performed a promoter assay for Axin2, which is known to be induced by activation of the WNT/beta-catenin (betacat) signaling pathway. Thereby, we found that Axin2 promoter activity was enhanced in the lrp1 knockdown cells. Furthermore, when the WNT/beta-catenin pathway was activated in chondrocytic cells by WNT3a or SB216763, which inhibits the phosphorylation of GSK3beta, the mRNA levels of aggrecan and col2a1 were decreased, whereas that of mmp13 was increased. Additionally, the level of phosphorylated protein kinase C (PKC) zeta was also decreased in the lrp1 knockdown cells. When the phosphorylation of PKCzeta was selectively inhibited, aggrecan and col2a1 mRNA levels decreased, whereas the mmp13 mRNA level increased. These data demonstrate that LRP1 exerts remarkable effects to retain the mature phenotype of chondrocytes as a critical mediator of cell signaling. Our findings also indicate that the onset of hypertrophy during endochondral ossification appears to be particularly dependent on the WNT and PKC signaling initiated by LRP1.

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  • Novel Transcriptional Regulation of CCN2/CTGF by Nuclear Translocation of MMP3

    Takanori Eguchi, Satoshi Kubota, Kazumi Kawata, Yoshiki Mukudai, Junji Uehara, Toshihiro Ohgawara, Soichiro Ibaragi, Akira Sasaki, Takuo Kuboki, Masaharu Takigawa

    CCN PROTEINS IN HEALTH AND DISEASE: AN OVERVIEW OF THE FIFTH INTERNATIONAL WORKSHOP ON THE CCN FAMILY OF GENES   255 - +   2010

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    CCN2/CTGF, previously known as Connective Tissue Growth Factor, is a crucial regulator of extra-cellular matrix (ECM), which promotes ECM synthesis and stabilization. As their family name clearly implies, matrix metalloproteases (MMPs) are also localized in the ECM, where they function as proteases, modulating cell signaling by cleaving proteins such as matrix proteins, growth factors and growth factor receptors. Strong expression of CCN2/CTGF in chondrocytic cells occurs through transcription enhancer dominant in chondrocytes (TRENDIC). Matrix metalloprotease-3 (MMP3) is a novel TRENDIC-binding transcription factor for CCN2/CTGF expression. First, MMP3 cDNA was cloned as a TRENDIC-binding factor by Southwestern screening. The interaction between MMP3 and TRENDIC was confirmed by a gel shift assay and chromatin immunoprecipitation. The CCN2/CTGF promoter was activated by transfected MMP3, whereas a TRENDIC mutant for the promoter lost the response. In addition, the knockdown of MMP3 suppressed CCN2/CTGF expression. Cytochemical and histochemical analyses demonstrated that MMP3 was detected in the nuclei of chondrocytic cells in culture and also in the nuclei of normal and osteoarthritic chondrocytes in vivo. The nuclear translocation of externally added recombinant MMP3 was observed in 30 min after the addition, and six putative nuclear localization signals were found in MMP3. These results indicated a novel trans-activation mechanism of CCN2/CTGF by the nuclear translocation of MMP3 through binding with TRENDIC in chondrocytes. Although MMPs historically had been recognized as a protease for extra-cellular proteins, this study indicated that it also stimulates ECM synthesis through CCN2/CTGF trans-activation. This novel regulatory role of the ECM may contribute to understanding the mechanism of not only the development, but also the pathogenesis of arthritis fibrosis and periodontitis.

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  • Nucleophosmin/B23: A Multifunctional Regulator that Determines the Fate of CCN2 mRNA

    Satoshi Kubota, Yoshiki Mukudai, Harumi Kawaki, Seiji Kondo, Takanori Eguchi, Kumi Sumiyoshi, Toshihiro Ohgawara, Tsuyoshi Shimo, Masaharu Takigawa

    CCN PROTEINS IN HEALTH AND DISEASE: AN OVERVIEW OF THE FIFTH INTERNATIONAL WORKSHOP ON THE CCN FAMILY OF GENES   41 - +   2010

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    CCN2/CTGF is a multifunctional molecule that has been shown to play a central role in chondrocyte differentiation. During this process, the expression of ccn2 is tightly regulated to confer a maximal level at prehypertrophic - hypertrophic stages, in which the 3'-untranslated region (UTR) of the mRNA is critically involved in mediating its post-transcriptional regulation. In our previous studies, we found that a 40-kDa protein binding specifically to an RNA cis-element, 3'-100/50, in the 3'-UTR of the chicken ccn2 mRNA regulated the intracellular stability of the mRNA. The interaction of this 40-kDa protein with 3'-100/50 was enhanced in proliferating chondrocytes, in which ccn2 mRNA is rapidly degraded; whereas a prolonged half life of ccn2 mRNA is observed in hypertrophic chondrocytes, where the interaction of the 40 kDa-protein and 3'-100/50 is diminished. Collectively, the data suggested that this 40-kDa protein acts as a ccn2-specific mRNA destabilizer during chondrocyte differentiation.
    In this present study we finally identified this 40-kDa protein as nucleophosmin (NPM)/B23. NPM is a nuclear-cytoplasmic shuttling protein that is characterized by its multiple functionality. This protein is known to be a histone chaperone, a regulator of ribosomal RNA transcription, as well as an RNA-binding post-transcriptional regulator of gene expression. In our hands, direct binding of NPM to 3'-100/50 was confirmed not only by RNA EMSA and UV crosslinking assays, but also by RNA immunoprecipitation analysis. By using recombinant chicken NPM, we could successfully reconstitute the post-transcriptional regulation of ccn2 by NPM in vitro and found that this regulation was more robust in chondrocytes than in fibroblasts. Furthermore, siRNA-mediated gene silencing of NPM in vivo clearly showed enhanced ccn2 gene expression and a prolonged half life of the ccn2 mRNA, confirming the functional property of NPM as a specific destabilizer of the ccn2 mRNA in living cells.
    The 5'-100/50 element, a target of NPM, is evolutionally conserved among vertebrate species. Therefore, we consider NPM to be a critical post-transcriptional regulator of ccn2 acting via 3'-UTR during endochondral ossification and possibly, in other physiological and pathological states as well.

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  • 低密度リポタンパク受容体関連タンパク-1(LRP1)の軟骨細胞分化における多面的作用機構

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 森谷 徳文, 近藤 誠二, 西田 崇, 皆木 省吾, 滝川 正春

    日本生化学会大会プログラム・講演要旨集   82回 ( Vol.1 )   4T4p - 9   2009.9

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  • 軟骨細胞におけるCCN2遺伝子の転写後調節機構におけるNucleophosmin/B23の機能的意義

    住吉 久美, 久保田 聡, 椋代 義樹, 近藤 誠二, 川木 晴美, 江口 傑徳, 大河原 敏博, 山城 隆, 滝川 正春

    日本生化学会大会プログラム・講演要旨集   82回   3T18a - 8   2009.9

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  • Nucleophosmin/B23によるChicken CCN2遺伝子の軟骨細胞特異的転写後調節

    住吉 久美, 久保田 聡, 椋代 義樹, 近藤 誠二, 川木 晴美, 江口 傑徳, 山城 隆, 滝川 正春

    Journal of Oral Biosciences   51 ( Suppl. )   105 - 105   2009.8

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  • 低密度リポタンパク受容体関連タンパク-1(LRP1)の軟骨細胞分化における機能とその作用機構

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 森谷 徳文, 近藤 誠二, 西田 崇, 皆木 省吾, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   27回   179 - 179   2009.7

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  • Regulation of chondrocytic phenotype by micro RNA 18a: involvement of Ccn2/Ctgf as a major target gene. Reviewed International journal

    Toshihiro Ohgawara, Satoshi Kubota, Harumi Kawaki, Seiji Kondo, Takanori Eguchi, Naito Kurio, Eriko Aoyama, Akira Sasaki, Masaharu Takigawa

    FEBS letters   583 ( 6 )   1006 - 10   2009.3

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    We searched for miRNAs that were down-regulated in chondrocytic cells and predicted to target CCN2/connective tissue growth factor (CCN2/CTGF) that promotes endochondral ossification. Among them, expression of miR-18a was most strongly repressed in chondrocytic cells. Reporter gene analysis confirmed the functionality of an miR-18a target in the 3'-untranslated region of Ccn2 mRNA, which was predicted in silico. Indeed, introduction of miR-18a efficiently repressed the CCN2 production from chondrocytic cells. Finally, transfected miR-18a significantly repressed the mature chondrocytic phenotype. Our present study revealed a regulatory role for miR-18a in chondrocytic differentiation through CCN2.

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  • 低密度リポタンパク受容体関連タンパク-1(LRP1)の軟骨細胞分化における多面的作用機構

    河田かずみ, 河田かずみ, 久保田聡, 江口傑徳, 青山絵理子, 森谷徳文, 近藤誠二, 西田崇, 皆木省吾, 滝川正春

    生化学   2009

  • Posttranscriptional regulation of chicken ccn2 gene expression by nucleophosmin/B23 during chondrocyte differentiation. Reviewed International journal

    Yoshiki Mukudai, Satoshi Kubota, Harumi Kawaki, Seiji Kondo, Takanori Eguchi, Kumi Sumiyoshi, Toshihiro Ohgawara, Tsuyoshi Shimo, Masaharu Takigawa

    Molecular and cellular biology   28 ( 19 )   6134 - 47   2008.10

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    CCN2/CTGF is a multifunctional factor that plays a crucial role in the growth and differentiation of chondrocytes. The chicken ccn2 gene is regulated not only at the transcriptional level but also by the interaction between a posttranscriptional element in the 3' untranslated region (3'-UTR) and a cofactor. In the present study, we identified a nucleophosmin (NPM) (also called B23) as this cofactor. Binding of NPM to the element was confirmed, and subsequent analysis revealed a significant correlation between the decrease in cytosolic NPM and the increased stability of the ccn2 mRNA during chondrocyte differentiation in vivo. Furthermore, recombinant chicken NPM enhanced the degradation of chimeric RNAs containing the posttranscriptional cis elements in a chicken embryonic fibroblast extract in vitro. It is noteworthy that the RNA destabilization effect by NPM was far more prominent in the cytosolic extract of chondrocytes than in that of fibroblasts, representing a chondrocyte-specific action of NPM. Stimulation by growth factors to promote differentiation changed the subcellular distribution of NPM in chondrocytes, which followed the expected patterns from the resultant change in the ccn2 mRNA stability. Therefore, the present study reveals a novel aspect of NPM as a key player in the posttranscriptional regulation of ccn2 mRNA during the differentiation of chondrocytes.

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  • マイクロRNA 18aによるCcn2/Ctgf遺伝子を介した軟骨細胞分化の制御機構の解明

    大河原 敏博, 久保田 聡, 川木 晴美, 近藤 誠二, 江口 傑徳, 佐々木 朗, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   26回   168 - 168   2008.10

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  • 軟骨細胞成熟における低密度リポ蛋白受容体関連タンパク-1(LRP1)の関与

    河田かずみ, 河田かずみ, 久保田聡, 江口傑徳, 青山絵理子, 皆木省吾, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   21st   2008

  • Gene expression and distribution of connective tissue growth factor (CCN2/CTGF) during secondary ossification center formation. Reviewed International journal

    Morihiko Oka, Satoshi Kubota, Seiji Kondo, Takanori Eguchi, Chisa Kuroda, Kazumi Kawata, Shogo Minagi, Masaharu Takigawa

    The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society   55 ( 12 )   1245 - 55   2007.12

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    CCN2/connective tissue growth factor (CCN2/CTGF) is a critical signaling modulator of mesenchymal tissue development. This study investigated the localization and expression of CCN2/CTGF as a factor supporting angiogenesis and chondrogenesis during development of secondary ossification centers in the mouse tibial epiphysis. Formation of the secondary ossification center was initiated by cartilage canal formation and blood vessel invasion at 7 days of age, and onset of ossification was observed at 14 days. In situ hybridization showed that CCN2/CTGF mRNA was distinctively expressed in the region of the cartilage canal and capsule-attached marginal tissues at 7 days of age, and distinct expression was also observed in proliferating chondrocytes around the marrow space at 14 days of age. Immunostaining showed that CCN2/CTGF was distributed broadly around the expressed cells located in the central region of the epiphysis, where the chondrocytes become hypertrophic and the cartilage canal enters into the hypertrophic mass. Furthermore, an overlapping distribution of metalloproteinase (MMP)9 and CCN2/CTGF was found in the secondary ossification center. These findings suggest that the CCN2/CTGF is involved in establishing epiphyseal vascularization and remodeling, which eventually determines the secondary ossification center in the developing epiphysial cartilage.

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  • Different transcriptional strategies for ccn2/ctgf gene induction between human chondrocytic and breast cancer cell lines. Reviewed International journal

    Takanori Eguchi, Satoshi Kubota, Kazumi Kawata, Yoshiki Mukudai, Toshihiro Ohgawara, Kohei Miyazono, Kyouji Nakao, Seiji Kondo, Masaharu Takigawa

    Biochimie   89 ( 3 )   278 - 88   2007.3

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    Connective tissue growth factor (CTGF/CCN2) plays a critical role in endochondral bone formation; however, CCN2 also promotes angiogenesis and bone metastasis in breast cancer. Chondrocytic HCS-2/8 cells and breast cancer MDA231 cells produce over 6 times more CCN2 than any other cell type. In this study, we demonstrate that these cell lines employ different transcriptional strategies for ccn2 gene induction. Four tandem copies of the dominant transcriptional enhancer in chondrocytes (4 x TRENDIC) were chimerically connected to an SV40 promoter-luciferase construct and subsequently analyzed. The enhancement of the promoter activity by 4 x TRENDIC was greater in the HCS-2/8 cells (7-fold) than in the other 4 cell lines (3-4 fold). The TRENDIC-binding protein complex was detected at a higher signal in the HCS-2/8 cells than in the other cell lines. In addition, the HCS-2/8 nuclear factors strongly targeted not only TRENDIC, but also the previously reported basal control element and a novel enhancer element in the ccn2 promoter. In contrast, high-level ccn2 gene induction in MDA231 cells was largely dependent on Smad signaling through the Smad-binding element in the ccn2 promoter. Based on these results, we propose a model of differential transcription of the ccn2 gene between the chondrocytic cell line and the breast cancer cell line, and therefore imply that these cells utilize distinct transcriptional strategies to obtain the enhanced CCN2 production that is not observed in other types of cells.

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  • Pathogenic role of connective tissue growth factor (CTGF/CCN2) in osteolytic metastasis of breast cancer. Reviewed International journal

    Tsuyoshi Shimo, Satoshi Kubota, Norie Yoshioka, Soichiro Ibaragi, Sachiko Isowa, Takanori Eguchi, Akira Sasaki, Masaharu Takigawa

    Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research   21 ( 7 )   1045 - 59   2006.7

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    UNLABELLED: The role of CTGF/CCN2 in osteolytic metastasis by breast cancer cells and its mechanism of action were studied. Osteolytic metastasis accompanied by CCN2 and PTHrP overproduction was efficiently inhibited by an anti-CCN2 antibody. Furthermore, we found that CCN2 was induced by PTHrP through PKA-, PKC-, and ERK-mediated pathways therein. INTRODUCTION: Connective tissue growth factor (CTGF/CCN2) is a mediator of local angiogenesis induced by breast cancer, but its role in osteolytic metastasis has not been evaluated. PTH-related peptide (PTHrP) is another critical factor in the development of the osteolytic metastasis. Using both in vivo and in vitro approaches, we studied whether/how neutralization of CCN2 prevented bone metastasis and how PTHrP signaling is related. MATERIALS AND METHODS: A mouse model of bone metastasis by human breast cancer cell line MDA231 was treated with a CCN2-neutralizing antibody, and osteolytic bone metastases were assessed on radiographs and immunohistochemistry. Ccn2 gene expression and transcription were examined by Northern blot and luciferase analysis. Immunoblot analysis and kinase inhibitors were used to identify the signaling pathways implicated. Anti-angiogenic/osteoclastogenic effects of ccn2 downregulation were also evaluated. RESULTS: Treatment of mice with a CCN2-neutralizing antibody greatly decreased osteolytic bone metastasis, microvasculature, and osteoclasts involved. The antibody also suppressed the growth of subcutaneous tumor in vivo and proliferation and migration of human umbilical vein endothelial cells (HUVECs) in vitro. Downregulation of ccn2 also repressed osteoclastogenesis. CCN2 expression was specifically observed in cancer cells producing PTHrP and type I PTH/PTHrP receptor (PTH1R) invaded the bone marrow, and PTHrP strongly upregulated ccn2 in MDA231 cells in vitro. Activation of protein kinase C (PKC) and protein kinase A (PKA) was necessary and sufficient for the stimulation of ccn2 by PTHrP. Indeed, inhibition of the extracellular signal-regulated kinase (ERK1/2), PKC, or PKA by specific inhibitors counteracted the stimulation of ccn2 expression. Incubation of MDA231 cells with PTHrP induced the activation of ERK1/2. Consistent with these findings, inhibition of PKC prevented PTHrP-induced ERK1/2 activation, whereas 12-O-tetradecanoylphorbol13-acetate (TPA), a stimulator of PKC, upregulated it. CONCLUSIONS: CCN2 was critically involved in osteolytic metastasis and was induced by PKA- and PKC-dependent activation of ERK1/2 signaling by PTHrP. Thus, CCN2 may be a new molecular target for anti-osteolytic therapy to shut off the PTHrP-CCN2 signaling pathway.

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  • Possible role of LRP1, a CCN2 receptor, in chondrocytes. Reviewed International journal

    Kazumi Kawata, Takanori Eguchi, Satoshi Kubota, Harumi Kawaki, Morihiko Oka, Shogo Minagi, Masaharu Takigawa

    Biochemical and biophysical research communications   345 ( 2 )   552 - 9   2006.6

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    Low density lipoprotein receptor (LDLR)-related protein 1 (LRP1/CD91) is one of the receptors of CCN2 that conducts endochondral ossification and cartilage repair. LRP1 is a well-known endocytic receptor, but its distribution among chondrocytes remains to be elucidated. We herein demonstrate for the first time that the distribution of LRP1 in chondrocytes except for hypertrophic chondrocytes in vivo and in vitro. Interestingly, the LRP1 levels were higher in mature chondrocytic HCS-2/8 and osteoblastic SaOS-2 than in other cells, whereas the other LDLR family members involved in ossification were detected at lower levels in HCS-2/8. It was interesting to note that in HCS-2/8, LRP1 was observed not only on the cell surface and in the cytoplasm, but also in the nucleus. Exogenously added CCN2 was incorporated into HCS-2/8, which was partially co-localized with LRP1, and targeted to the recycling endosomes and nucleus as well as the lysosomes. These findings suggest specific roles of LRP1 in cartilage biology.

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  • 二次骨化中心形成過程に発現する結合組織成長因子CTGF/CCN2のパールカン陽性軟骨細胞への特異的集積

    岡 森彦, 久保田 聡, 近藤 誠二, 江口 傑徳, 河田 かずみ, 黒田 知沙, 皆木 省吾, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   23回   229 - 229   2005.6

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  • Collaborative action of M-CSF and CTGF/CCN2 in articular chondrocytes: possible regenerative roles in articular cartilage metabolism. Reviewed International journal

    Kyouji Nakao, Satoshi Kubota, Hideyuki Doi, Takanori Eguchi, Morihiko Oka, Takuo Fujisawa, Takashi Nishida, Masaharu Takigawa

    Bone   36 ( 5 )   884 - 92   2005.5

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    It is known that expression of the macrophage colony-stimulating factor (M-CSF) gene is induced in articular chondrocytes upon inflammation. However, the functional role of M-CSF in cartilage has been unclear. In this study, we describe possible roles of M-CSF in the protection and maintenance of the articular cartilage based on the results of experiments using human chondrocytic cells and rat primary chondrocytes. Connective tissue growth factor (CTGF/CCN2) is known to be a potent molecule to regenerate damaged cartilage by promoting the growth and differentiation of articular chondrocytes. Here, we uncovered the fact that M-CSF induced the mRNA expression of the ctgf/ccn2 gene in those cells. Enhanced production of CTGF/CCN2 protein by M-CSF was also confirmed. Furthermore, M-CSF could autoactivate the m-csf gene, forming a positive feed-back network to amplify and prolong the observed effects. Finally, promotion of proteoglycan synthesis was observed by the addition of M-CSF. These findings taken together indicate novel roles of M-CSF in articular cartilage metabolism in collaboration with CTGF/CCN2, particularly during an inflammatory response. Such roles of M-CSF were further supported by the distribution of M-CSF producing chondrocytes in experimentally induced rat osteoarthritis cartilage in vivo.

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  • Regulation of chicken ccn2 gene by interaction between RNA cis-element and putative trans-factor during differentiation of chondrocytes. Reviewed International journal

    Yoshiki Mukudai, Satoshi Kubota, Takanori Eguchi, Seiji Kondo, Kyouji Nakao, Masaharu Takigawa

    The Journal of biological chemistry   280 ( 5 )   3166 - 77   2005.2

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    CCN2/CTGF is a multifunctional growth factor. Our previous studies have revealed that CCN2 plays important roles in both growth and differentiation of chondrocytes and that the 3'-untranslated region (3'-UTR) of ccn2 mRNA contains a cis-repressive element of gene expression. In the present study, we found that the stability of chicken ccn2 mRNA is regulated in a differentiation stage-dependent manner in chondrocytes. We also found that stimulation by bone morphogenetic protein 2, platelet-derived growth factor, and CCN2 stabilized ccn2 mRNA in proliferating chondrocytes but that it destabilized the mRNA in prehypertrophic-hypertrophic chondrocytes. The results of a reporter gene assay revealed that the minimal repressive cis-element of the 3'-UTR of chicken ccn2 mRNA was located within the area between 100 and 150 bases from the polyadenylation tail. Moreover, the stability of ccn2 mRNA was correlated with the interaction between this cis-element and a putative 40-kDa trans-factor in nuclei and cytoplasm. In fact, the binding between them was prominent in proliferating chondrocytes and attenuated in (pre)hypertrophic chondrocytes. Stimulation by the growth factors repressed the binding in proliferating chondrocytes; however, it enhanced it in (pre)hypertrophic chondrocytes. Therefore, gene expression of ccn2 mRNA during endochondral ossification is properly regulated, at least in part, by changing the stability of the mRNA, which arises from the interaction between the RNA cis-element and putative trans-factor.

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  • Interaction of AP-1 and the ctgf gene: a possible driver of chondrocyte hypertrophy in growth cartilage. Reviewed

    Norifumi H Moritani, Satoshi Kubota, Takanori Eguchi, Tomohiro Fukunaga, Takashi Yamashiro, Teruko Takano-Yamamoto, Hideki Tahara, Kazumi Ohyama, Toshio Sugahara, Masaharu Takigawa

    Journal of bone and mineral metabolism   21 ( 4 )   205 - 10   2003

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    The expression of the connective tissue growth factor ( ctgf) gene increases along with the differentiation of growth cartilage cells, and the highest expression is observed in the hypertrophic stage. Similarly, recent reports demonstrated c- fos expression in chondrocytes in the early hypertrophic zone of growth cartilage, and suggested that the c- fos gene may play a crucial role in the regulation of hypertrophic differentiation. A chondrocytic human cell line, HCS-2/8, is known to retain a variety of chondrocytic phenotypes. When such cells were kept overconfluent, they expressed increasing levels of c- fos transcripts along a time course phenotypically similar to that of hypertrophic differentiation. Moreover, by using a competitive electromobility-shift assay, we found that AP-1, a Fos/Jun heterodimer, in HCS-2/8 was capable of binding not only to a typical AP-1-binding DNA fragment but also to the enhancer fragment of the ctgf gene. Based on the findings above, we hypothesize that, prior to hypertrophic differentiation, AP-1-related oncogenes are activated and that their gene products subsequently activate ctgf gene expression, which might eventually induce hypertrophy.

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  • 軟骨由来成長因子CTGF/Hcs24の細胞種特異的遺伝子発現抑制機構の解析

    森谷 徳文, 久保田 聡, 江口 傑徳, 近藤 誠二, 菅原 利夫, 滝川 正春

    生化学   74 ( 8 )   913 - 913   2002.8

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  • A novel cis-element that enhances connective tissue growth factor gene expression in chondrocytic cells. Reviewed International journal

    Takanori Eguchi, Satoshi Kubota, Seiji Kondo, Takuo Kuboki, Hirofumi Yatani, Masaharu Takigawa

    Biochemical and biophysical research communications   295 ( 2 )   445 - 51   2002.7

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    To clarify the chondrocyte-specific regulatory mechanism of connective tissue growth factor (ctgf) gene expression, we analyzed the functionality and DNA-protein interaction of the CTGF promoter. Comparative luciferase assay of the CTGF promoter deletion mutants among HCS-2/8 chondrocytic cells and fibroblastic cells revealed that a 110-bp region in the promoter was crucial for the HCS-2/8-specific transcriptional enhancement. Subsequent competitive gel shift assay revealed that transcription factors in HCS-2/8 nuclei bound to a 60-bp portion in the corresponding region. Relative luciferase activity from a CTGF promoter with mutant TGF-beta response element (TbRE) was 16.9% lower than that from an intact promoter. On the other hand, relative luciferase activity from a CTGF promoter with 4bp point mutations at 30bp upstream of the TbRE was 47.7% lower than that from the intact one. The binding activity of HCS-2/8 nuclear factor(s) to the sequence over the 4-bp was remarkably higher than that of any nuclear extract from other types of cells. Therefore, we entitled the sequence 'TRENDIC', a transcription enhancer dominant in chondrocytes, which stands for a novel enhancer for chondrocyte-specific CTGF gene expression.

    DOI: 10.1016/S0006-291X(02)00700-3

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  • Connective tissue growth factor increased by hypoxia may initiate angiogenesis in collaboration with matrix metalloproteinases. Reviewed International journal

    Seiji Kondo, Satoshi Kubota, Tsuyoshi Shimo, Takashi Nishida, Gen Yosimichi, Takanori Eguchi, Toshio Sugahara, Masaharu Takigawa

    Carcinogenesis   23 ( 5 )   769 - 76   2002.5

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    Connective tissue growth factor (CTGF) is known to be a potent angiogenic factor. Here we investigated how CTGF and matrix metalloproteinases (MMPs) are involved in the early stage of hypoxia-induced angiogenesis using human breast cancer cell line, MDA231, and vascular endothelial cells. Hypoxic stimulation (5% O(2)) of MDA231 cells increased their steady-state level of ctgf mRNA by approximately 2-fold within 1.5 h, and the levels remained at a plateau up to 6 h, and then decreased by 12 h as compared with the cells cultured under the normoxic condition. Membrane-type 1 MMP (MT1-MMP) mRNA levels was also increased within a few hours of the exposure to hypoxia. Indeed, ELISA revealed that the CTGF protein/cell in medium conditioned by MDA231 cells exposed to hypoxia was maximally greater at 24 h than in the medium from normoxic cultures and that the secretion rate (supernatant CTGF/cell layer CTGF) increased in a time-dependent manner from 24 to 72 h of hypoxic exposure. Hypoxic induction of CTGF was also confirmed by immunohistochemical analyses. Furthermore, zymogram analysis revealed that the production of active MMP-9 was also induced in MDA231 cells incubated under hypoxic conditions. Finally, we found that recombinant CTGF also increased the expression of a number of metalloproteinases that play a role in the vascular invasive processes and decreased the expression of tissue inhibitors of metalloproteinases by vascular endothelial cells. These findings suggest that hypoxia stimulates MDA231 cells to release CTGF as an angiogenic modulator, which initiates the invasive angiogenesis cascade by modulating the balance of extracellular matrix synthesis and degradation via MMPs secreted by endothelial cells in response to CTGF. This cascade may play critical roles in the hypoxia-induced neovascularization that accompanies tumor invasion in vivo.

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  • 遺伝子発現の抑圧的調節を仲介するマウスctgf3'-UTR segmentの性質

    近藤 誠二, 久保田 聡, 江口 傑徳, 服部 高子, 中西 徹, 菅原 利夫, 滝川 正春

    日本口腔科学会雑誌   50 ( 6 )   568 - 568   2001.11

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  • Cell-type-specific trans-activation of herpes simplex virus thymidine kinase promoter by the human T-cell leukemia virus type I Tax protein. Reviewed

    Kubota S, Mukudai Y, Hattori T, Eguchi T, Kondo S, Takigawa M

    DNA and cell biology   20 ( 9 )   563 - 568   2001.9

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    The human T-cell leukemia virus type I Tax protein (HTLV-I Tax) is known as a trans-activating factor for a variety of genes, including those of cytokines. Here, we show that Tax is capable of activating the herpes simplex virus thymidine kinase (HSV-TK) promoter in certain mammalian cell lines. In murine NIH 3T3 fibroblasts and human HeLa cells, trans-activation by Tax was remarkably strong, whereas in human chondrocytic HCS-2/8 and monkey kidney Cos-7 cells, the responsiveness of the TK promoter to Tax was poor. Deletion analysis revealed that one of the two previously described Sp1 sites is required for the Tax responsiveness, whereas the CTF binding site is not. The results suggest possible interactions between the oncogenic Tax protein and the viral TK in coinfected cells in vivo. Care should be taken in the context of HTLV-I research, as the HSV-TK promoter has been widely used in molecular biology and gene therapeutics.

    DOI: 10.1089/104454901317094972

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  • 多機能成長因子CTGF/Hcs24遺伝子の転写後制御エレメント,CAESARの作用機序

    久保田 聡, 近藤 誠二, 椋代 義樹, 江口 傑徳, 服部 高子, 中西 徹, 滝川 正春

    生化学   73 ( 8 )   710 - 710   2001.8

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  • Regulatory Mechanism of Human Connective Tissue Growth Factor (CTGF/Hcs24) Gene Expression in a Human Chondrocytic Cell Line, HCS-2/8 Reviewed

    T. Eguchi, S. Kubota, S. Kondo, T. Shimo, T. Hattori, T. Nakanishi, T. Kuboki, H. Yatani, M. Takigawa

    Journal of Biochemistry   130 ( 1 )   79 - 87   2001.7

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    DOI: 10.1093/oxfordjournals.jbchem.a002965

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  • Novel mode of processing and secretion of connective tissue growth factor/ecogenin(CTGF/Hcs24) in chondrocytic HCS-2/8 cells Reviewed

    S. Kubota, T. Eguchi, T. Shimo, T. Nishida, T. Hattori, S. Kondo, T. Nakanishi, M. Takigawa

    Bone   29 ( 2 )   155 - 161   2001

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    The synthesis, processing, and secretion of human connective tissue growth factor (CTGF/Hcs24) in a human chondrocytic cell line, HCS-2/8, were analyzed immunochemically. By metabolic pulse-labeling, chasing, and subsequent immunoprecipitation analyses, active synthesis of CTGF was observed not only in growing HCS-2/8 cells, but also in confluent cells. However, secretion and processing of CTGF were found to be regulated differentially, depending upon the growth status. During phases of growth, HCS-2/8 cells released CTGF molecules immediately without sequestering them within the cell layer. In contrast, after the cells reached confluence, the secretion slowed, resulting in an accumulation of CTGF in the cells or extracellular matrices (ECMs). Also, in confluent cell layers, a 10 kDa protein that was reactive to an anti-CTGF serum was observed. This CTGF-related small protein was not detected immediately after labeling, but gradually appeared within 6 h after chase, which suggests its entity as a processed subfragment of CTGF. Surprisingly, the 10 kDa protein was stable even 48 h after synthesis, and was not released by ECM digestion, suggesting an intracellular maintenance and function. Taken together, the behavior of CTGF in HCS-2/8 cells is remarkably different from that reported in fibroblasts, which may represent unique roles for CTGF in the growth and differentiation of chondrocytes. Copyright © 2001 Elsevier Science Inc.

    DOI: 10.1016/S8756-3282(01)00492-6

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  • Characterization of a mouse ctgf 3 '-UTR segment that mediates repressive regulation of gene expression

    S Kondo, S Kubota, T Eguchi, T Hattori, T Nakanishi, T Sugahara, M Takigawa

    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS   278 ( 1 )   119 - 124   2000.11

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    We isolated a small segment of the 3'-untranslated region (3'-UTR) in the mouse connective tissue growth factor (ctgf/fisp12) gene and evaluated its functionality. Comparison of the nucleotide sequences of human and mouse ctgf 3'-UTRs revealed a conserved small segment of 91 bases, The corresponding segments of the 3'-UTRs shared as much as 82.4% homology, whereas the overall homology between the 3'-UTRS was 71.8%. To study the functionality of the conserved segment, the corresponding region of mouse ctgf cDNA was amplified from NIH3T3 cells. When it was fused downstream of a marker gene, it showed remarkable repressive effects on gene expression. The repressive effect of the sense form was more prominent than that of the antisense form. Computer analyses of these sequence predicted stable secondary structures, suggesting that they act at the RNA level. The predicted structures of the sense and antisense forms appeared to be slightly different, which is consistent with the difference in repressive function. These findings defined the conserved small element in the mouse ctgf gene as a potent negative regulator of gene expression, which may act at a posttranscriptional level. (C) 2000 Academic Press.

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  • Characterization of a Mouse ctgf 3′-UTR Segment That Mediates Repressive Regulation of Gene Expression Reviewed

    Seiji Kondo, Satoshi Kubota, Takanori Eguchi, Takako Hattori, Tohru Nakanishi, Toshio Sugahara, Masaharu Takigawa

    Biochemical and Biophysical Research Communications   278 ( 1 )   119 - 124   2000.11

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    DOI: 10.1006/bbrc.2000.3780

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  • Identification of an RNA element that confers post-transcriptional repression of connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) gene: Similarities to retroviral RNA-protein interactions Reviewed

    S Kubota, S Kondo, T Eguchi, T Hattori, T Nakanishi, RJ Pomerantz, M Takigawa

    ONCOGENE   19 ( 41 )   4773 - 4786   2000.9

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    The repressive effect of the 3'-untranslated region (3'-UTR) in human connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) mRNA on gene expression had been demonstrated in our previous study. Here, we identified a minimal RNA element in the 3'-UTR, which acts as a cis-acting element of structure-anchored repression (CAESAR). Deletion analyses of the 3'-UTR led us to minimize the element of 84 bases at the junction of the coding region and the 3'-UTR. The minimized RNA segment is predicted, and actually capable of forming a stable secondary structure in vitro. Mutational analyses disclosed a significant relationship between the predicted structure and repressive effect. The utility of CAESAR as a post-transcriptional regulatory element was represented by the fact that steady-state mRNA levels were not affected by CAESAR linked in cis, while protein levels from such a chimeric gene were markedly reduced. Of note, the CAESAR sequence exerted no effect, when it was placed upstream of the promoter. Finally, RNA gel electromobility-shift analyses demonstrated a nuclear factor that interacts with the folded CAESAR. Taken together, it was uncovered that CAESAR of ctgf is a novel post-transcriptional structured RNA regulatory element, probably acting through direct interactions with a nuclear factor as observed in retroviral RNA elements with certain proteins.

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  • 軟骨由来の成長因子CTGF/Hcs24遺伝子の転写後制御エレメント,CAESARの構造機能連関

    久保田 聡, 近藤 誠二, 江口 傑徳, 服部 高子, 中西 徹, 滝川 正春

    生化学   72 ( 8 )   972 - 972   2000.8

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  • 軟骨由来の成長因子CTGF/Hcs24遺伝子に同定された新たな転写後制御エレメント,CAESAR

    久保田 聡, 近藤 誠二, 江口 傑徳, 服部 高子, 中西 徹, 滝川 正春

    日本骨代謝学会雑誌   18 ( 2 )   119 - 119   2000.6

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  • 軟骨細胞および線維芽細胞株におけるCTGF/Ecogenin遺伝子発現の3'-UTRによる調節

    近藤 誠二, 久保田 聡, 江口 傑徳, 服部 高子, 中西 徹, 菅原 利夫, 滝川 正春

    岡山歯学会雑誌   19 ( 1 )   205 - 206   2000.6

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  • アデノウイルスを用いた顎関節への遺伝子導入の試み

    縄稚 久美子, 窪木 拓男, 完山 学, 藤沢 拓生, 園山 亘, 江口 傑徳, 上原 淳二, 矢谷 博文, 山下 敦

    日本補綴歯科学会雑誌   43 ( 102回特別 )   180 - 180   1999.10

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Books

  • Multiplex immunostaining method to distinguish HSP isoforms. In: Methods Molecular Biology, Vol. , Stuart K. Calderwood and Thomas L. Prince (Eds): Chaperones.

    Hotaka Kawai, Kisho Ono, Takanori Eguchi( Role: Contributor ,  Corresponding author, Senior author)

    Springer Nature Switzerland AG, Cham, Switzerland.  2023.8 

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  • Large-scale databases and portals on cancer genome to analyze chaperone genes correlated to patient prognosis. In: Methods Molecular Biology, Vol. , Stuart K. Calderwood and Thomas L. Prince (Eds): Chaperones.

    Kisho Ono, Takanori Eguchi( Role: Contributor ,  Corresponding author, Senior author)

    Springer Nature Switzerland AG, Cham, Switzerland.  2023.8 

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  • Proteomic profiling of the extracellular vesicle-chaperome in cancer. In: Methods Molecular Biology, Vol. , Stuart K. Calderwood and Thomas L. Prince (Eds): Chaperones.

    Kisho Ono, Takanori Eguchi( Role: Contributor ,  Corresponding author, Senior author)

    Springer Nature Switzerland AG, Cham, Switzerland.  2023.8 

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  • Monitoring of the heat shock response with a real-time luciferase reporter. In: Methods Molecular Biology, Stuart K. Calderwood and Thomas L. Prince (Eds): Chaperones

    Andrew Ackerman, Toshiki Kijima, Takanori Eguchi, Thomas Prince( Role: Contributor ,  Co-author)

    Springer Nature Switzerland AG, Cham, Switzerland.  2023.8 

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  • Multiple targeting of HSP isoforms to challenge isoform specificity and compensatory expression. In: Methods Molecular Biology, Vol. , Stuart K. Calderwood and Thomas L. Prince (Eds): Chaperones.

    Kisho Ono, Takanori Eguchi( Role: Contributor ,  Corresponding author, Senior author)

    Springer Nature Switzerland AG, Cham, Switzerland.  2023.8 

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  • Western Blot Protocols for Analysis of CCN Proteins and Fragments in Exosomes, Vesicle-free Fractions, and Cells. In: Masaharu Takigawa (eds) CCN Proteins. Methods in Molecular Biology, vol 2582.

    Kisho Ono, Yuka Okusha, Manh Tien Tran, Koki Umemori, Takanori Eguchi( Role: Contributor ,  Corresponding author, senior author, pp.39-57)

    Humana, New York, NY.  2023 

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  • Comprehensive Method for Exosome Isolation and Proteome Analysis for Detection of CCN Factors in/on Exosomes. In: Masaharu Takigawa (eds) CCN Proteins. Methods in Molecular Biology, vol 2582.

    Takanori Eguchi, Yuka Okusha, Yanyin Lu, Kisho Ono, Eman A. Taha, Shiro Fukuoka( Role: Contributor ,  First and corresponding author, pp.59-76)

    Humana, New York, NY.  2023 

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  • Protocol for CRISPR/Cas Genome Editing for Investigating Cell Communication Network. In: Masaharu Takigawa (eds) CCN Proteins. Methods in Molecular Biology, vol 2582.

    Yuka Okusha, Takanori Eguchi( Role: Contributor ,  Corresponding author, pp.157-167.)

    Humana, New York, NY.  2023 

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  • Transfection, Spinfection, Exofection, and Luciferase Assays for Analysis of CCN Genes Expression Mechanism. In: Masaharu Takigawa. (eds) CCN Proteins. Methods in Molecular Biology, vol 2582.

    Takanori Eguchi, Yanyin Lu, Eman A. Taha, Yuka Okusha( Role: Contributor ,  First and corresponding author, pp.103-126.)

    Humana, New York, NY.  2023 

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  • Cdc37 as a Co-chaperone to Hsp90. In: Edkins, A.L., Blatch, G.L. (eds) The Networking of Chaperones by Co-Chaperones. Subcellular Biochemistry, vol 101.

    Thomas L. Prince, Benjamin J. Lang, Yuka Okusha, Takanori Eguchi, Stuart K. Calderwood( Role: Contributor ,  pp.141-158)

    Springer, Cham.  2023 

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  • HSP Stimulation on Macrophages and Dendritic Cells Activates Innate Immune System. In: Heat Shock Proteins in Inflammatory Diseases

    Yanyin Lu, Takanori Eguchi( Role: Contributor ,  Correspondence, Co-first author)

    Springer Nature Switzerland AG  2021 

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  • Heat Shock Proteins and Periodontitis – Cross-Reaction Between Bacterial and Human HSP in Periodontal Infection Linking with Cardiovascular Diseases. In: Heat Shock Proteins in Inflammatory Diseases, Heat Shock Proteins, vol. 22.

    Tadashi Yamamoto, Takanori Eguchi( Role: Contributor ,  Corresponding author, pp.19-32.)

    2021 

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  • Extracellular Vesicle-Associated Moonlighting Proteins: Heat Shock Proteins and Metalloproteinases. In: Heat Shock Proteins in Inflammatory Diseases, Heat Shock Proteins, vol. 22.

    Takanori Eguchi, Eman A. Taha( Role: Contributor ,  First and corresponding author, pp.1-18.)

    2021 

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  • Roles of HSP on Antigen Presentation. In: Heat Shock Proteins in Human Diseases, Heat Shock Proteins vol 21.

    Kazuyuki Furuta, Takanori Eguchi( Role: Contributor ,  Corresponding author, pp.275-280.)

    2020.7 

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  • Regulatory Roles of HSP90-Rich Extracellular Vesicles. In: Asea A., Kaur P. (eds) Heat Shock Protein 90 in Human Diseases and Disorders. Heat Shock Proteins, vol 19.

    Eguchi T, Ono K, Kawata K, Okamoto K, Calderwood S.K( Role: Contributor ,  First and corresponding author, pp.3-17.)

    2019 

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  • Monitoring of the Heat Shock Response with a Real-Time Luciferase Reporter. In: Stuart Calderwood and Thomas Prince. (eds) Chaperones. Methods in Molecular Biology, vol 1709.

    Toshiki Kijima, Takanori Eguchi, Len Neckers, Thomas L. Prince( Role: Contributor ,  pp.35-45.)

    2018 

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  • Regulatory Roles for Hsp70 in Cancer Incidence and Tumor Progression. In: Mario D. Galigniana (ed) Frontiers in Structural Biology, vol 1. Role of Molecular Chaperones in Structural Folding, Biological Functions, and Drug Interactions of Client Proteins.

    Taka Eguchi, Benjamin J. Lang, Ayesha Murshid, Thomas Prince, Jianlin Gong, Stuart K Calderwood( Role: Contributor ,  First author, pp.1-22.)

    2018 

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  • Promoter analyses of CCN genes. in Methods in Molecular Biology. CCN Proteins: Methods and Protocols

    Eguchi T, Kubota S, Takigawa M( Role: Contributor ,  First author, pp.177-185.)

    2017.1 

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  • CCN proteins in Health and Diseases: Novel Transcriptional Regulation of CCN2/CTGF by Nuclear Translocation of MMP3

    Eguchi T, Kubota S, Kawata K, Mukudai, Y, Takigawa M( Role: Contributor ,  First author, pp.255-264.)

    2010.1 

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  • CCN Proteins in Health and Disease: Nucleophosmin/B23: A Multifunctional Regulator that Determines the Fate of CCN2 mRNA.

    Kubota S, Mukudai Y, Kawaki H, Kondo S, Eguchi T, Sumiyoshi K, Ohgawara T, Shimo T, Takigawa M( Role: Contributor ,  pp.41-56.)

    2010.1 

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  • エンドトキシン研究12:58-60. ヒストンアセチル化制御薬を用いたHMGB1の放出制御

    ( Role: Contributor ,  p.58-60)

    2009 

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MISC

  • 細胞外小胞に搭載されたムーンライティングMMPによるマトリックス制御と転移促進(Metalloproteinase on extracellular vesicles regulates ECM in tumor microenvironment and promotes invasion and metastasis)

    奥舎 有加, タハ・エマン, 陸 彦因, シータ・モナ, 河合 穂高, 十川 千春, 岡元 邦彰, 江口 傑徳

    日本癌学会総会記事   81回   P - 1118   2022.9

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  • 高転移性癌細胞由来の細胞外小胞に搭載されたMMP3によるCtgf/Ccn2発現調節機能と癌転移促進(Extracellular vesicles enriched with moonlighting metalloproteinase are highly transmissive, Pro-tumorigenic, and trans-activates cellular communication network factor(CCN2/CTGF): CRISPR against cancer)

    奥舎 有加, 江口 傑徳, Tran Manh T., 十川 千春, 吉田 賀弥, 板垣 まみ, Taha Eman A., 小野 喜章, 青山 絵理子, 岡村 裕彦, 小崎 健一, Calderwood Stuart K., 滝川 正春, 岡元 邦彰

    Journal of Oral Biosciences Supplement   2022   35 - 35   2022.9

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  • 転写因子MZF1とSCAND1による相反的なEMT制御(MZF1 and SCAND1 reciprocally regulate epithelial-to-mesenchymal transition)

    江口 傑徳, ラング・ベンジャミン, シータ・モナ, チャン・マン, ヴェンギェル・バーバラ, カルダーウッド・スチュアート

    日本癌学会総会記事   81回   E - 3046   2022.9

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  • 口腔癌間質由来のCCL2はCCR2陽性骨髄由来免疫抑制細胞の腫瘍間質への動員に関与する

    河合 穂高, メイ・ワトウ, 高畠 清文, 冨田 秀太, 小野 喜章, 江口 傑徳, 大原 利章, 中野 敬介, 長塚 仁

    日本がん免疫学会総会プログラム・抄録集   26回   91 - 91   2022.6

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  • 侵襲性歯周炎の血液診断マーカー候補となる細胞外小胞由来マイクロRNAとその炎症誘導機構の探索

    森 彩乃, 山本 直史, 井手口 英隆, 河村 麻理, 河本 美奈, 伊東 昌洋, 小野 喜章, 中山 真彰, 江口 傑徳, 大野 充昭, 大森 一弘, 高柴 正悟

    日本歯周病学会会誌   64 ( 春季特別 )   114 - 114   2022.5

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  • 口腔癌のエクソソームを介した腫瘍進展機序の解明と新規治療戦略の開発に向けて 分子シャペロン搭載エクソソームの可能性

    小野 喜章, 江口 傑徳, 十川 千春, 奥舎 有加, 岡元 邦彰, 佐々木 朗

    口腔組織培養学会誌   31 ( 1 )   33 - 34   2022.3

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  • 侵襲性歯周炎の血液診断マーカー候補となる細胞外小胞由来マイクロRNAとその炎症誘導機構の探索

    森彩乃, 山本直史, 井手口英隆, 河村麻理, 河本美奈, 伊東昌洋, 小野喜章, 中山真彰, 江口傑徳, 大野充昭, 大森一弘, 高柴正悟

    日本歯周病学会会誌(Web)   64   2022

  • Porphyromonas gingivalis impairs fetal development through macrophage extracellular vesicles

    棚井あいり, 福原瑶子, 河合穂高, 江口傑徳, 池亀美華, 吉田賀弥, 岡村裕彦

    硬組織再生生物学会学術大会・総会プログラム・抄録集   2021   205 - 205   2021.10

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  • がん研究の新たな潮流〜歯学基礎研究からの発信〜 癌促進性エクソソームとその機能抑制(Cancer-promoting exosomes and their functional suppression)

    江口 傑徳

    Journal of Oral Biosciences Supplement   2021   96 - 96   2021.10

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  • 難治性がん克服のための腫瘍オルガノイドを利用した創薬研究

    江口 傑徳

    医科学応用研究財団研究報告   38   273 - 277   2021.2

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  • エクソソームに搭載される多機能タンパク質:HSP90とMMP3

    江口傑徳

    日本臨床ストレス応答学会大会抄録集   15th   2021

  • マクロファージ由来のHSP90搭載エクソソームは口腔癌細胞への効率的な分子送達能と細胞毒性を有す

    陸彦因, 陸彦因, 江口傑徳

    日本臨床ストレス応答学会大会抄録集   15th   2021

  • Cancer-promoting exosomes and their functional suppression

    江口傑徳

    Journal of Oral Biosciences Supplement (Web)   2021   2021

  • Porphyromonas gingivalis impairs placental and fetal development through macrophage-derived extracellular vesicles

    棚井あいり, 福原瑶子, 江口傑徳, 河合穂高, 池亀美華, 岡村裕彦

    Journal of Oral Biosciences Supplement (Web)   2021   2021

  • 難治性がん克服のための腫瘍オルガノイドを利用した創薬研究

    江口傑徳

    医科学応用研究財団研究報告(CD-ROM)   38   2021

  • 侵襲性歯周炎の血液診断バイオマーカーとしての細胞外小胞由来マイクロRNAの探索

    河本 美奈, 山本 直史, 河村 麻理, 森 彩乃, 山城 圭介, 大森 一弘, 小野 喜章, 江口 傑徳, 十川 千春, 高柴 正悟

    岡山歯学会雑誌   39 ( 2 )   35 - 36   2020.12

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  • 発生と疾患にみる新たな細胞間コミュニケーション オルガノイドと細胞外小胞を癌研究に応用する

    江口 傑徳, 十川 千春, 岡元 邦彰

    Journal of Oral Biosciences Supplement   2020   118 - 118   2020.9

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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

  • 三次元腫瘍オルガノイドの開発と細胞外小胞の新機能

    江口傑徳, 江口傑徳

    日本解剖学会総会・全国学術集会講演プログラム・抄録集   125th   2020

  • がんの多様性と新規治療法への展望 口腔癌の進展と抵抗性における細胞外小胞の重要な役割

    江口 傑徳

    Journal of Oral Biosciences Supplement   2019   77 - 77   2019.10

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  • 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, 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   2019   165 - 165   2019.10

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  • Heat Shock Proteinとがん温熱療法の最前線 悪性腫瘍の微小環境におけるCDC37とHSP90の重要性

    江口 傑徳

    Thermal Medicine   35 ( Suppl. )   50 - 50   2019.9

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  • 口腔癌の進展と抵抗性における細胞外小胞の重要な役割

    江口傑徳

    Journal of Oral Biosciences Supplement (Web)   2019   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

  • 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

  • Drug repositioning using three-dimensional, MMP9 promoter activity-based anticancer drug screening

    Sogawa Chiharu, Eguchi Takanori, Okusha Yuka, Ohyama Kazumi, Ono Kisho, Sogawa Norio, Okamoto Kuniaki

    Proceedings for Annual Meeting of The Japanese Pharmacological Society   92   1-P-096   2019

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    Cancer is one of the most serious diseases all over the world, especially metastasis and drug resistance are leading causes of death. There is an urgent need to establish new strategies for drug discovery. Success in the drug discovery depends on the development of appropriate tumor models that correspond closely to native tumor situation. Matrix metalloproteinases (MMPs) represent the most prominent family of proteinases associated with tumorigenesis and are regulators of tumor milieu. The cancer stem cell model fits well with tumorigenesis, metastasis and drug resistance. We have shown that cancer cell aggregation led to hypoxic tumoroids with marked upregulation of reprogramming and stemness genes as increased cancer stem cell using a 3D culture system. In the present study, we established a novel MMP9 promoter-driven cell-based reporter system using a rapidly metastatic colon cancer cell in the 3D culture system that evaluates cancer stemness and invasiveness. We used a concept of drug repositioning-using known molecules for new indications. We selected several compounds with inhibition to both tumoroid formation and MMP9 promoter activity. One of the compounds inhibited primary tumor formation, invasion and metastasis.

    DOI: 10.1254/jpssuppl.92.0_1-p-096

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  • Depletion of choresterol lipid efflux pump ABCG1 triggers accumulation of exosomes and regression of tumors

    Eguchi Takanori, Sogawa Chiharu, Namba Yuri, Okusha Yuka, Kawai Hotaka, Ono Kisho, Itagaki Mami, Murakami Jun, Ohyama Kazumi, Asaumi Junichi, Okamoto Kuniaki

    Proceedings for Annual Meeting of The Japanese Pharmacological Society   92   2-YIA-26   2019

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    The ATP-binding cassette transporter G1 (ABCG1) is a cholesterol lipid efflux pump whose role in tumor growth has been largely unknown. Our transcriptomics revealed that ABCG1 was powerfully expressed in rapidly metastatic, aggregative colon cancer cells, in all the ABC transporter family members. Coincidently, genetic amplification of ABCG1 is found in 10% to 35% of clinical samples of metastatic cancer cases. Expression of ABCG1 was further elevated in three-dimensional tumoroids (tumor organoids) within stemness-enhancing tumor milieu, whereas depletion of ABCG1 lowered cellular aggregation and tumoroid growth in vitro as well as hypoxia-inducible factor 1α in cancer cells around the central necrotic areas in tumors in vivo. Notably, depletion of ABCG1 triggered the intracellular accumulation of extracellular vesicles (EVs) and regression of tumoroids. Collectively, these data suggest that ABCG1 plays a crucial role in tumorigenesis in metastatic cancer and that depletion of ABCG1 triggers tumor regression with the accumulation of EVs, their derivatives and cargos, implicating a novel ABCG1-targeting therapeutic strategy by which redundant and toxic substances may be accumulated in tumors leading to their regression.

    DOI: 10.1254/jpssuppl.92.0_2-yia-26

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  • 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

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  • 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

    奥舎 有加, 板垣 まみ, 十川 千春, 江口 傑徳, 岡元 邦彰

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

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  • 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

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  • 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

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  • 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

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  • 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, 坂井 詠子, 筑波 隆幸, 岡元 邦彰

    Journal of Oral Biosciences Supplement   2018   219 - 219   2018.9

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  • EGFR標的抗体薬セツキシマブによる口腔癌由来のEGFR陽性エクソソームの制御

    藤原 敏史, 江口 傑徳, 十川 千春, 村上 純, 浅海 淳一, 小崎 健一

    日本口腔科学会雑誌   67 ( 2 )   184 - 184   2018.7

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  • 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

  • RASP:難治性がんにみられるストレス抵抗性と小胞/HSP分泌特性

    江口傑徳

    臨床ストレス応答学会大会抄録集   13th   2018

  • 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

  • 侵襲性歯周炎患者の血漿エクソソーム由来microRNAの発現解析

    高木美奈, 山本直史, 河村麻理, 高知信介, 山城圭介, 大森一弘, 江口傑徳, 十川千春, 高柴正悟

    日本歯周病学会会誌(Web)   60   2018

  • 口腔扁平上皮癌細胞株が分泌するエクソソーム中に見つかった腫瘍進展・転移因子

    小野喜章, 江口傑徳, 佐々木朗

    日本口腔腫瘍学会総会・学術大会プログラム・抄録集   36th   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

  • 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

  • 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

  • Tumoroid Never Knows?腫瘍オルガノイドと細胞外小胞からみる難治性がん研究・治療のミライ

    江口傑徳, 小野喜章, 奥舎有加, 藤原敏史, 十川千春, CALDERWOOD Stuart

    日本分子生物学会年会プログラム・要旨集(Web)   41st   2018

  • EGFR標的抗体薬セツキシマブによる口腔癌由来のEGFR陽性エクソソームの制御

    藤原敏史, 藤原敏史, 江口傑徳, 江口傑徳, 十川千春, 村上純, 浅海淳一, 小崎健一

    日本口腔科学会雑誌(Web)   67 ( 2 )   2018

  • 薬剤耐性遺伝子を標的としたがん幹細胞制御

    難波 友里, 江口 傑徳, 十川 千春, 奥舎 有加, 村上 純, 浅海 淳一, 岡元 邦彰, 小崎 健一

    生命科学系学会合同年次大会   2017年度   [1LBA - 050]   2017.12

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  • プロテアーゼ活性を持たないプロテアーゼの炎症および癌における役割

    江口 傑徳, 奥舎 有加, Calderwood Stuart, 岡元 邦彰

    生命科学系学会合同年次大会   2017年度   [2LBA - 028]   2017.12

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  • 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   2017   378 - 378   2017.9

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  • 細胞内におけるマトリックスメタロプロテアーゼ(MMP)の役割

    江口 傑徳, 奥舎 有加, 中野 敬介, 久保田 聡, 滝川 正春, カルダーウッド・スチュアート, 小崎 健一

    Journal of Oral Biosciences Supplement   2017   249 - 249   2017.9

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  • 高転移性細胞株が有する浸潤能・転移能におけるMmpsの役割

    奥舎 有加, 江口 傑徳, 十川 千春, 小崎 健一

    日本癌学会総会記事   76回   P - 3106   2017.9

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  • CATABOLIC EFFECTS OF FGF-1 ON CHONDROCYTES WITH REDUCED CCN2 PRODUCTION THAT PROMOTES CARTILAGE REGENERATION: POSSIBLE ROLE IN OSTEOARTHRITIS

    A. Elseoudi, T. Abd El Kader, T. Nishida, E. Aoyama, T. Eguchi, M. Takigawa, S. Kubota

    OSTEOPOROSIS INTERNATIONAL   28   S225 - S225   2017.7

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  • 転移性癌細胞クローン薬剤耐性

    難波 友里, 江口 傑徳, 村上 純, 十川 千春, 浅海 淳一, 小崎 健一

    日本口腔科学会雑誌   66 ( 2 )   184 - 184   2017.7

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  • 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

  • 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

  • 細胞内におけるマトリックスメタロプロテアーゼ(MMP)の役割

    江口傑徳, 江口傑徳, 奥舎有加, 中野敬介, 久保田聡, 滝川正春, CALDERWOOD SK, 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

  • 転写因子MZF1とSCAND1は前立腺癌細胞においてEMT、癌促進性キナーゼシグナルおよび分子シャペロン発現を制御する

    江口 傑徳, ベン・ラング, トーマス・プリンス, 十川 千春, 奥舎 有加, フィリップ・グレイ, 小崎 健一, スチュアート・カルダーウッド

    日本癌学会総会記事   75回   E - 1041   2016.10

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  • 異なる転移性と独自の細胞標識システムを具備したcolon26亜株における発現プロファイリング

    奥舎 有加, 江口 傑徳, 大山 和美, 十川 千春, 小崎 健一

    日本癌学会総会記事   75回   P - 3155   2016.10

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  • マウス大腸がん細胞株高転移性亜株を用いたがんの浸潤・転移イメージングモデルの確立

    十川 千春, 奥舎 有加, 大山 和美, 江口 傑徳, 小崎 健一

    日本癌学会総会記事   75回   P - 3153   2016.10

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  • An old dog's new tricks: iMMP-3 Processes Nuclear Matrix and Converts Heterochromatin Proteins Leading to Transcriptional Promotion of HSP genes

    Takanori Eguchi, Akihisa Mino, Benjamin J. Lang, Stuart K. Calderwood

    FASEB JOURNAL   30   2016.4

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  • An old dog's new tricks: iMMP-3 Processes Nuclear Matrix and Converts Heterochromatin Proteins Leading to Transcriptional Promotion of HSP genes

    Takanori Eguchi, Akihisa Mino, Benjamin J. Lang, Stuart K. Calderwood

    FASEB JOURNAL   30   2016.4

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  • 軟骨細胞分化における癌抑制遺伝子PDGFR-like(PDGFRL)の役割

    河田かずみ, 久保田聡, 江口傑徳, 青山絵理子, 森谷徳文, 岡森彦, 川木晴美, 滝川正春, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   29th   2016

  • 転移性癌細胞株に含まれる癌幹細胞様細胞および薬剤耐性についての解析

    難波友里, 難波友里, 十川千春, 奥舎有加, 村上純, 浅海淳一, 小崎健一, 江口傑徳, 江口傑徳

    日本分子生物学会年会プログラム・要旨集(Web)   39th   2016

  • 腫瘍微小環境シグナルによる上皮間葉転換およびエクソソーム変化についての解析

    藤原敏史, 十川千春, 小野喜章, 村上純, 浅海淳一, 小崎健一, 江口傑徳

    日本分子生物学会年会プログラム・要旨集(Web)   39th   2016

  • 軟骨細胞における血小板由来増殖因子受容体様(PDGFRL)遺伝子の発現(Expression of platelet-derived growth factor receptor-like (PDGFRL) gene in chondrocytes)

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 森谷 徳文, 岡 森彦, 川木 晴美, 滝川 正春

    日本生化学会大会プログラム・講演要旨集   84回   3P - 0351   2011.9

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  • バルプロ酸(VPA)はERK1/2の活性化を介してHigh Mobility Group Box 1(HMGB1)の産生を誘導する

    杉浦 進介, 江口 傑徳, 萩原 真, 小松 寿明, 谷川 順美, 野口 俊英, 松下 健二

    日本血栓止血学会誌   21 ( 2 )   229 - 229   2010.4

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  • ヒトMMP3は新規の転写因子様機能によりCTGF/CCN2遺伝子を制御する

    江口傑徳

    岡山歯学会雑誌   29 ( 1 )   2010

  • ヒストンアセチル化制御薬を用いたHMGB1の放出制御

    杉浦 進介, 江口 傑徳, 小松 寿明, 松下 健二

    エンドトキシン研究   12   58 - 60   2009.11

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  • Micro RNA 18a regulates chondrocytic phenotype: Involvement of Ccn2/Ctgf as a major target gene

    T. Ohgawara, S. Kubota, H. Kawaki, S. Kondo, T. Eguchi, A. Sasaki, M. Takigawa

    BONE   44   S42 - S43   2009.5

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    DOI: 10.1016/j.bone.2009.01.109

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  • E-selectinの新機能 感染の制御

    小松 寿明, 江口 傑徳, 杉浦 進介, 猪俣 恵, 古市 保志, 松下 健二

    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   130回   219 - 219   2009.5

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  • 新規炎症性サイトカインHMGB1のアセチル化制御薬による放出制御

    杉浦 進介, 江口 傑徳, 猪俣 恵, 小松 寿明, 野口 俊英, 松下 健二

    日本歯周病学会会誌   51 ( 春季特別 )   133 - 133   2009.4

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  • 軟骨細胞においてMMP3は核移行しCCN2/CTGFの転写活性化因子として働く

    江口傑徳, 江口傑徳, 久保田聡, 河田かずみ, 椋代義樹, 上原淳二, 大河原敏博, 大河原敏博, 伊原木聰一郎, 佐々木朗, 窪木拓男, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   22nd   2009

  • Release of HMGB1 is regulated by histone deacetylase inhibitors and histone acetyltransferase inhibitors

    Sugiura Shinsuke, Eguchi Takanori, Inomata Megumi, Komatsu Toshinori, Noguchi Toshihide, Matsushita Kenji

    Program and Abstracts of Annual Meeting of the Japanese Society of Periodontology   2009s   122 - 122   2009

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    DOI: 10.14833/amjsp.2009s.0.122.0

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  • 低密度リポタンパク受容体関連タンパク-1(LRP1)の軟骨細胞分化における機能とその作用機構

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 皆木 省吾, 滝川 正春

    日本生化学会大会・日本分子生物学会年会合同大会講演要旨集   81回・31回   4T13 - 2   2008.11

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  • 軟骨細胞分化における低密度リポタンパク受容体関連タンパク-1(LRP1)の関与

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵理子, 皆木 省吾, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   26回   234 - 234   2008.10

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  • 可溶型セレクチンによる病原体排除

    江口傑徳, 松下健二

    生化学   2008

  • Th1およびTh2由来サイトカインはSTATの活性化を介してWeibel-Palade Bodyの構成因子の発現を変化させる

    猪俣恵, 猪俣恵, 江口傑徳, 杉浦進介, 杉浦進介, 野口俊英, 松下健二

    生化学   2008

  • マイクロRNA18aによるCcn2/Ctgf遺伝子の制御機構の解明とその軟骨分化における意義

    大河原敏博, 大河原敏博, 久保田聡, 川木晴美, 近藤誠二, 江口傑徳, 佐々木朗, 滝川正春

    生化学   2008

  • 軟骨細胞における低密度リポタンパク受容体関連タンパク1(LRP1)の機能解析(Functional analysis of the low density lipoprotein receptor-related protein (LRP1) in chondrocytes)

    河田 かずみ, 久保田 聡, 江口 傑徳, 青山 絵里子, 皆木 省吾, 滝川 正春

    日本生化学会大会・日本分子生物学会年会合同大会講演要旨集   80回・30回   2T19 - 6   2007.11

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  • Novel transcription factor-like function of MMP-3/stromelysin-1 that regulates connective tissue growth factor (CTGF/CCN2) gene transcription

    T. Eguchi, S. Kubota, K. Kawata, Y. Mukudai, T. Yanagita, T. Ohgawara, J. Uehara, S. Ibaragi, M. Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   22   S261 - S261   2007.9

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  • 軟骨細胞においてマトリックス金属プロテアーゼ-3(MMP3)は核移行し結合組織成長因子(CCN2/CTGF)の転写活性化因子として働く

    江口傑徳, 久保田聡, 河田かずみ, 椋代義樹, 大河原敏博, 上原淳二, 伊原木聰一郎, 佐々木朗, 窪木拓男, 滝川正春, 滝川正春

    生化学   2007

  • 軟骨細胞にみられる低密度リポタンパク受容体関連タンパク1(LRP1)の局在と機能の解析

    河田かずみ, 河田かずみ, 久保田聡, 江口傑徳, 青山絵里子, 川木晴美, 岡森彦, 皆木省吾, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   20th   2007

  • Purification and functional characterization of a protein that regulate ccn2 gene expression during chicken chondrocyte differentiation.

    Y. Mukudai, S. Kubota, S. Kondo, T. Eguchi, H. Kawaki, M. Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   21   S149 - S149   2006.9

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  • Post-transcriptional regulation of CCN2/CTGF gene expression during differentiation of chicken chondrocytes: involvement of a putative trans-factor which interacts with a cis-element in the 3 '-UTR of mRNA

    Y Mukudai, S Kubota, T Eguchi, S Kondo, K Nakao, M Takigawa

    FEBS JOURNAL   272   284 - 285   2005.7

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  • 軟骨細胞における低密度リポタンパク受容体関連タンパク1(LRP1)の遺伝子発現とタンパク質局在

    河田 かずみ, 江口 傑徳, 久保田 聡, 川木 晴美, 岡 森彦, 皆木 省吾, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   23回   260 - 260   2005.6

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  • 各種軟骨細胞におけるM-CSFの産生とその生理的役割

    中尾 匡志, 久保田 聡, 藤澤 拓生, 岡 森彦, 江口 傑徳, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   23回   177 - 177   2005.6

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  • 軟骨組織及び軟骨細胞における低密度リポタンパク受容体関連タンパク1(LRP1)の遺伝子発現とタンパク質局在

    河田かずみ, 河田かずみ, 江口傑徳, 江口傑徳, 久保田聡, 川木晴美, 岡森彦, 皆木省吾, 滝川正春

    日本分子生物学会年会講演要旨集   28th   2005

  • 成長因子 1 二次骨化中心形成過程におけるCTGF/CCN2およびMMP-9の発現と局在

    岡森彦, 久保田聡, 近藤誠二, 江口傑徳, 河田かずみ, 黒田知沙, 皆木省吾, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   18th   2005

  • 軟骨発生と分化 6 軟骨細胞における低密度リポタンパク受容体関連タンパク1(LRP1)の発現

    河田かずみ, 江口傑徳, 久保田聡, 川木晴美, 岡森彦, 皆木省吾, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   18th   2005

  • 関節炎と軟骨(2)2 変形性関節症(OA)モデルにおけるM-CSFの産生と修復における意義

    中尾匡志, 久保田聡, 西田崇, 岡森彦, 江口傑徳, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   18th   2005

  • 軟骨細胞の分化過程におけるCCN2/CTGF遺伝子転写後調節機構の解析

    椋代 義樹, 久保田 聡, 江口 傑徳, 近藤 誠二, 中尾 匡志, 滝川 正春

    Journal of oral biosciences   46 ( 5 )   396 - 396   2004.9

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  • 軟骨細胞におけるM-CSFとCTGFの協調的誘導とその効果

    中尾 匡志, 久保田 聡, 西田 崇, 江口 傑徳, 滝川 正春

    Journal of oral biosciences   46 ( 5 )   396 - 396   2004.9

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  • ニワトリ軟骨細胞の分化過程におけるCCN2/CTGF遺伝子の転写後発現調節機構の解析

    椋代 義樹, 久保田 聡, 江口 傑徳, 近藤 誠二, 中尾 匡志, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   22回   159 - 159   2004.8

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  • 二次骨化中心形成過程における結合組織成長因子CTGF/CCN2の発現 血管新生因子としての関与

    岡 森彦, 久保田 聡, 江口 傑徳, 河田 かずみ, 黒田 知沙, 皆木 省吾, 滝川 正春

    日本骨代謝学会学術集会プログラム抄録集   22回   199 - 199   2004.8

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  • Connective tissue growth factor(CTGF)の軟骨細胞特異的な転写調節機構の探索

    江口 傑徳, 久保田 聡, 椋代 義樹, 森谷 徳文, 中尾 匡志, 滝川 正春

    生化学   76 ( 3 )   303 - 303   2004.3

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  • Connective Tissue Growth Factor (CTGF/CCN2)プロモーター上の3つのシスエレメント〈軟骨細胞優位型エンハンサー(TRENDIC),スマッド結合配列(SBE),TGF-beta応答領域(TbRE)〉の機能比較-軟骨細胞様細胞株HCS-2/8と乳癌細胞株MDA231における違い-

    江口傑徳, 久保田聡, 河田かずみ, 中尾匡志, 大河原敏博, 滝川正春

    日本分子生物学会年会プログラム・講演要旨集   27th   2004

  • 軟骨細胞の分化過程におけるニワトリ結合組織成長因子(CTGF/Hcs24)遺伝子の転写後発現調節機構の解析

    椋代義樹, 久保田聡, 江口傑徳, 近藤誠二, 滝川正春

    日本軟骨代謝学会プログラム・抄録集   17th   2004

  • stromelysin-1の翻訳開始機構に関する検討-軟骨細胞様細胞株HCS-2/8を用いた解析-

    柳田剛志, 江口傑徳, 久保田聡, 山本照子, 滝川正春

    日本分子生物学会年会プログラム・講演要旨集   27th   2004

  • 軟骨細胞の分化課程におけるニワトリ結合組織成長因子(CTGF/Hcs24)遺伝子の転写後発現調節機構の解析

    椋代義樹, 久保田聡, 江口傑徳, 近藤誠二, 滝川正春

    日本分子生物学会年会プログラム・講演要旨集   26th   2003

  • Novel cis-element TRENDIC that enhance connective tissue growth factor (ctgf) gene expression in chondrocytic HCS-2/8.

    T Eguchi, S Kubota, S Kondo, Y Mukudai, T Kuboki, H Yatani, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   17   S224 - S224   2002.9

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  • Effects of IL-1beta and LPS on CTGF expression in mouse-derived odontoblast-like cells, MDPC-23.

    W Sonoyama, T Kuboki, T Fujisawa, T Eguchi, J Uehara, H Yatani, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   17   S327 - S327   2002.9

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  • Effects of proinflammatory factors on CTGF expression in odontoblast-like cells

    W Sonoyama, T Kuboki, T Fujisawa, T Eguchi, J Uehara, S Takashiba, H Yatani, M Takigawa

    JOURNAL OF DENTAL RESEARCH   81   A155 - A155   2002.3

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  • Immunohistochemical Localization and Transcriptional Regulation of Connective Tissue Growth Factor during Reparative Dentinogenesis

    SONOYAMA W, KUBOKI T, EGUCHI T, KOMORI C, FUJISAWA T, KANYAMA M, YATANI H

    45 ( 106 )   135 - 135   2001.10

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  • 低酸素によるヒト乳癌細胞における結合組織成長因子CTGF及びマトリクスメタロプロテアーゼの発現誘導

    近藤 誠二, 久保田 聡, 志茂 剛, 西田 崇, 吉道 玄, 江口 傑徳, 菅原 利夫, 滝川 正春

    日本癌学会総会記事   60回   183 - 183   2001.9

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  • Promoter activity determinant of human connective tissue growth factor (CTGF/Hcs24) gene in a human chondrocytic cell line, HCS-2/8.

    T Eguchi, S Kubota, S Kondo, T Shimo, T Nakanishi, T Kuboki, H Yatani, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   16   S326 - S326   2001.9

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  • 軟骨由来成長因子CTGF/Hcs24遺伝子の転写後制御エレメントCAESARの構造と機能

    久保田 聡, 近藤 誠二, 江口 傑徳, 服部 高子, 中西 徹, 滝川 正春

    歯科基礎医学会雑誌   43 ( 5 )   554 - 554   2001.8

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  • 低酸素による結合組織成長因子(CTGF)及びマトリクスメタロプロテアーゼ(MMP)活性の協調的発現誘導

    近藤 誠二, 久保田 聡, 志茂 剛, 西田 崇, 吉道 玄, 江口 傑徳, 菅原 利夫, 滝川 正春

    歯科基礎医学会雑誌   43 ( 5 )   632 - 632   2001.8

  • ヒト軟骨様細胞株HCS-2/8におけるCTGF/Hcs24遺伝子のプロモーター活性決定因子

    江口 傑徳, 久保田 聡, 近藤 誠二, 志茂 剛, 中西 徹, 窪木 拓男, 矢谷 博文, 滝川 正春

    歯科基礎医学会雑誌   43 ( 5 )   557 - 557   2001.8

  • 軟骨様細胞株HCS-2/8における多機能成長因子CTGF/Hcs24の転写から分泌まで

    江口 傑徳, 久保田 聡, 志茂 剛, 近藤 誠二, 中西 徹, 矢谷 博文, 滝川 正春

    生化学   73 ( 8 )   778 - 778   2001.8

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  • Regulatory Mechanism of Human Connective Tissue Growth Factor (CTGF/Hcs24) Gene Expression in a Human Chondrocytic Cell Line, HCS-2/8 Reviewed

    T. Eguchi, S. Kubota, S. Kondo, T. Shimo, T. Hattori, T. Nakanishi, T. Kuboki, H. Yatani, M. Takigawa

    Journal of Biochemistry   130 ( 1 )   79 - 87   2001.7

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    CTGF/Hcs24 is a multi-functional growth factor that potentiates either the growth or differentiation of mesenchymal cells, according to the biological conditions. Among various functional aspects of CTGF/Hcs24, it is especially notable that CTGF/Hcs24 may promote endochondral ossification in growth cartilage through all stages, and it is highly expressed in a human chondrosarcoma-derived chondrocytic cell line (HCS-2/8). In this study, to clarify the regulatory mechanism of CTGF/Hcs24 gene expression in chondrocytes, we analyzed the transcriptional activity of the CTGF/Hcs24 promoter and the effect of the CTGF/Hcs24 3′-untranslated region (3′-UTR) on gene expression in HCS-2/8 by means of an established DNA transfection and luciferase reporter gene assay system. As a result, the luciferase activity of the CTGF/Hcs24 promoter was found to be remarkably high in HCS-2/8. The 3′-UTR of the CTGF/Hcs24 gene strongly repressed the luciferase activity in HCS-2/8, when it was linked to the downstream of the luciferase reporter gene, suggesting its functionality also in chondrocytic cells. Deletion analysis of the CTGF/Hcs24 promoter clarified a major segment responsible for the enhanced CTGF/Hcs24 promoter activity in HCS-2/8. The TGF-β response element in the DNA segment was active in HCS-2/8, and point mutations in the element moderately decreased the highly maintained promoter activity with total loss of TGF-β responsiveness. These results indicate that the strong expression of the CTGF/Hcs24 gene in HCS-2/8 was mainly caused by high transcriptional activity of the CTGF/Hcs24 promoter, and that the TGF-β response element is one of the critical elements that support the high transcription activity.

    DOI: 10.1093/oxfordjournals.jbchem.a002965

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  • 軟骨由来成長因子CTGF/Hcs24のヒト軟骨細胞株HCS-2/8におけるプロセシングと分泌の様態

    久保田 聡, 江口 傑徳, 志茂 剛, 西田 崇, 服部 高子, 近藤 誠二, 中西 徹, 滝川 正春

    日本骨代謝学会雑誌   19 ( 2 )   104 - 104   2001.7

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  • ヒト軟骨肉腫由来軟骨様細胞株HCS-2/8における結合組織成長因子CTGF/Hcs24遺伝子のプロモーター活性決定因子

    江口 傑徳, 久保田 聡, 近藤 誠二, 志茂 剛, 中西 徹, 窪木 拓男, 矢谷 博文, 滝川 正春

    日本骨代謝学会雑誌   19 ( 2 )   102 - 102   2001.7

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  • 結合組織成長因子CTGF/Hcs24の軟骨細胞様細胞株HCS-2/8での発現と動態制御

    久保田 聡, 江口 傑徳, 志茂 剛, 服部 高子, 近藤 誠二, 中西 徹, 滝川 正春

    Connective Tissue   33 ( 2 )   157 - 157   2001.6

  • Characterization of a Mouse ctgf3′-UTR Segment that Mediates Repressive Regulation of Gene Expression.

    近藤誠二, 久保田聡, 江口傑徳, 服部高子, 中西徹, 菅原利夫, 滝川正春

    日本口腔科学会雑誌   50 ( 6 )   2001

  • 市販のカリエスリスクテストを用いた唾液中齲蝕原性細菌に対するクロルヘキシジンの殺菌効果に関する臨床的研究

    杉山 英樹, 水口 一, 上原 淳二, 大西 栄史, 畑中 乾志, 吉田 栄介, 半井 雅昭, 峯 篤史, 縄稚 久美子, 江口 傑徳

    日本補綴歯科学会雑誌   44 ( 6 )   867 - 867   2000.12

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  • Adenoviral Gene Transduction of Connective Tissue Growth Factor to a Mouse Osteoblastic Cell Line in Vitro and to the Bone Marrow of the Rat Tibia in Vivo

    KANYAMA M, KUBOKI T, EGUCHI T, NAWACHI K, UEHARA J, MIZUSHIMA T, SONOYAMA W, FUJISAWA T, YATANI H

    44 ( 104 )   196 - 196   2000.11

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  • A novel RNA element that confers post-transcriptional repression of human connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) gene.

    S Kubota, S Kondo, T Eguchi, T Hattori, T Nakanishi, RJ Pomerantz, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   15   S340 - S340   2000.9

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  • 軟骨由来成長因子CTGF/Hcs24遺伝子の転写後調節エレメントCAESAR 変異体分析によって得られた新たな知見

    久保田聡, 近藤誠二, 江口傑徳, 服部高子, 中西徹, 滝川正春

    日本分子生物学会年会プログラム・講演要旨集   23rd   2000

  • ヒト軟骨細胞様培養細胞株HCS-2/8における結合組織成長因子ctgf/ecogenin遺伝子発現制御機構

    江口傑徳, 久保田聡, 近藤誠二, 服部高子, 中西徹, 窪木拓男, 矢谷博文, 滝川正春

    日本分子生物学会年会プログラム・講演要旨集   23rd   2000

  • A cis-acting repressive element in the 3 '-untranslated region of the CTGF gene.

    S Kubota, T Hattori, T Eguchi, S Kondo, T Nakanishi, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH   14   S436 - S436   1999.9

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Presentations

  • 軟骨由来成長因子CTGF/Hcs24のヒト軟骨細胞株HCS-2/8におけるプロセシングと分泌の様態

    久保田 聡, 江口 傑徳, 志茂 剛, 西田 崇, 服部 高子, 近藤 誠二, 中西 徹, 滝川 正春

    日本骨代謝学会雑誌  2001.7  (一社)日本骨代謝学会

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    Event date: 2001.7

    Language:Japanese  

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  • 結合組織成長因子CTGF/Hcs24の軟骨細胞様細胞株HCS-2/8での発現と動態制御

    久保田 聡, 江口 傑徳, 志茂 剛, 服部 高子, 近藤 誠二, 中西 徹, 滝川 正春

    Connective Tissue  2001.6  日本結合組織学会

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    Event date: 2001.6

    Language:Japanese  

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  • Regulatory mechanism of human connective tissue growth factor (ctgf/hcs24) gene expression in a human chondrocytic cell line, HCS-2/8.

    T.Eguchi, S.Kubota, S.Kondo, T.Shimo, T.Hattori, T.Nakanishi, T.Kuboki, H.Yatani, M.Takigawa

    Journal of Biochemistry  2001 

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  • A novel RNA element that confers post-transcriptional repression of human connective tissue growth factor/hypertrophic chondrocyte specific 24 (ctgf/hcs24) gene.

    S Kubota, S Kondo, T Eguchi, T Hattori, T Nakanishi, RJ Pomerantz, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH  2000.9  AMER SOC BONE & MINERAL RES

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    Event date: 2000.9

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  • A cis-acting repressive element in the 3 '-untranslated region of the CTGF gene.

    S Kubota, T Hattori, T Eguchi, S Kondo, T Nakanishi, M Takigawa

    JOURNAL OF BONE AND MINERAL RESEARCH  1999.9  AMER SOC BONE & MINERAL RES

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  • Extracellular Vesicles as a Drug Carrier for Targeting Cancer Metabolism Kinase

    Sheta M, Yoshida K, Okamoto K, Eguchi T

    Biomedical Society for Stress Response Japan, the 17th Annual Meeting.  2023.11 

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  • Discovery and targeting of metabolism kinase in cancer organoid EV

    Sheta M, Eguchi T, Yoshida K, Okamoto K

    The 10th JSEV Annual Meeting  2023.10 

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  • Learn from the oldoncology, know the new: RNA-seq analysis of human chondrosarcoma-derived cells. Frontiers in CCN family reseaerch Invited

    Takanori Eguchi, Eriko Aoyama, Masaharu Takigawa

    Japan CCN Family Study Group  2023.9 

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  • Extracellular Vesicle Kinases Secreted by Metastatic Cancer Organoids

    Eguchi T, Sheta M, Yoshida K, Ono K

    The 82nd Annual Meeting of the Japanese Cancer Association, English Oral Session  2023.9 

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  • Extracellular Vesicles (EVs) as a Drug Carrier for Targeting Cancer Metabolism Kinase

    Sheta M, Yoshida K, Taha EA, Okamoto K, Eguchi T

    Brainstorming-2023  2023.8 

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  • Structural analysis of MZF nuclear bodies involved in cancer stress resistance

    Takanori Eguchi

    2022.11.6 

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  • SCAND1 and MZF1 co-regulate epithelial-to-mesenchymal transition in cancer

    Takanori Eguchi, Benjamin Lang, Mona Sheta, Manh Tien Tran, Barbara Wegiel, Stuart Calderwood

    2022.10.1 

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  • Resident stroma-secreted chemokine CCL2 governs myeloid-derived suppressor cells in the tumor microenvironment

    Hotaka Kawai, Shuta Tomida, Takanori Eguchi, Toshiaki Ohara, Kisho Ono, Hitoshi Nagatsuka

    2022.10.1 

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  • Metalloproteinase on Extracellular Vesicles Regulates ECM in Tumor Microenvironment and Promotes Invasion and Metastasis

    Yuka Okusha, Eman A. Taha, Yanyin Lu, Mona Sheta, Hotaka Kawai, Chiharu Sogawa, Kuniaki Okamoto, Takanori Eguchi

    2022.9.30 

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  • Proinflammatory Mechanism Regulated by Circulating Extracellular Vesicles-derived microRNAs: Potential Diagnostic Biomarkers for Aggressive Periodontitis

    2022.9.2 

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  • Extracellular vesicles enriched with moonlighting metalloproteinase are highly transmissive, Pro-tumorigenic, and trans-activates cellular communication network factor(CCN2/CTGF): CRISPR against cancer

    2022.9 

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  • 腔癌間質由来のCCL2はCCR2陽性骨髄由来免疫抑制細胞の腫瘍間質への動員に関与する

    河合 穂高, メイ・ワトウ, 高畠 清文, 冨田 秀太, 小野 喜章, 江口 傑徳, 大原 利章, 中野 敬介, 長塚 仁

    第26回日本がん免疫学会総会  2022.7 

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  • Porphyromonas gingivalis-infected Macrophage Extracellular Vesicles And Pregnancy

    Airi Tanai, Yoko Fukuhara, Takanori Eguchi, Koji Ueda, Mika Ikegame, Hirohiko Okamura

    2022 AADOCR/CADR Annual Meeting & Exhibition  2022.3 

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  • 細胞外小胞を利用した新規骨再生療法の検討

    福岡 史朗, 江口傑徳, 陸 彦因, 上原 健敬, 藤原 智洋, 中田 英二, 尾﨑 敏文, 岡元 邦彰

    第36回創薬・薬理フォーラム岡山  2021.12.25 

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  • 口腔癌のエクソソームを介した腫瘍進展機序の解明と新規治療戦略の開発に向けて 〜分子シャペロン搭載エクソソームの可能性〜

    小野喜章, 江口傑徳, 十川千春, 奥舎有加, 岡元邦彰, 佐々木朗

    第57回日本口腔組織培養学会学術大会  2021.11 

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  • エクソソームに搭載される多機能タンパク質:HSP90とMMP3

    江口傑徳

    第15回日本臨床ストレス応答学会大会  2021.11 

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  • マクロファージ由来のHSP90搭載エクソソームは口腔癌細胞への効率的な分子送達能と細胞毒性を有す

    陸 彦因, 江口傑徳

    第15回日本臨床ストレス応答学会大会  2021.11 

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  • 歯周病原菌に感染したマクロファージ由来の細胞外小胞は胎盤の血管形成を阻害する

    棚井あいり, 福原瑶子, 江口傑徳, 植田幸嗣, 吉田賀弥, 岡村裕彦

    第8回日本細胞外小胞学会  2021.10.18 

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  • Porphyromonas gingivalis impairs placental and fetal development through macrophage-derived extracellular vesicles

    2021.10 

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  • 侵襲性歯周炎の血液診断バイオマーカーとしての細胞外小胞由来マイクロRNAの探索

    河本美奈, 山本直史, 河村麻理, 森綾乃, 山城圭介, 大森一弘, 小野喜章, 江口傑徳, 十川千春, 高柴正悟

    第41回岡山歯学会  2020.10 

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  • Application of Organoids and Extracellular Vesicles to Cancer Research

    2020.9 

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  • Effects of Fibroblast Growth Factor 1 (FGF-1) on CCN2 Gene Expression in Chondrocytic Cells

    Abdellatif El-Seoudi, Tarek Abd El Kader, Takashi Nishida, Takanori Eguchi, Eriko Aoyama, Takako Hattori, Masaharu Takigawa, Satoshi Kubota

    Experimental Biology 2019 (EB2019) Annual Meeting  2020.4 

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  • 三次元腫瘍オルガノイドの開発と細胞外小胞の新機能

    江口傑徳

    第125回日本解剖学会総会全国学術集会:シンポジウム「顎口腔領域の発生と疾患に見る細胞間情報伝達機構の新たなカタチ」  2020.3 

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  • 細胞間情報伝達・細胞外小胞研究国際シンポジウム開催報告と今後

    江口傑徳

    第50回 ARCOCSセミナー:次世代研究育成グループ研究成果中間報告  2020.2 

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  • パーキンソン病治療薬の抗癌作用(ドラッグリポジショニングの提案)

    十川千春, 江口傑徳, Tran Tien Manh, 石毛真行, 河合穂高, 奥舎有加, 中野敬介, 十川紀夫, 小崎健一, 岡元邦彰

    第45回 岡山脳研究セミナー  2020.1.28 

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  • The Roles of Macrophage-derived Exosomes on the Oral Cancer Cells

    The International Symposium of Medical-Dental-Pharmaceutical Education and Research in Okayama  2019.12 

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  • Roles of MMP3 in Cargo Signature and Transmissive Activity of Cancer Extracellular Vesicles (Poster Competition Award)

    The 4th International Symposium of Medical-Dental-Pharmaceutical Education in Okayama  2019.12 

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  • Exosome-mediated Intercellular Communication using Multiplexing Fluorescent and Bioluminescent Reporter Systems

    The 4th International, Symposium of Medical, Dental Education in Okayama

    2019.12 

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  • オルガノイドを応用したドラッグリポジショニング開発

    十川千春, 江口傑徳, Tran Tien Manh, 石毛真行, 河合穂高, 奥舎有加, 中野敬介, 十川紀夫, 小崎健一, 岡元邦彰

    岡山歯学会  2019.12 

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  • Tumoroidを応用したゲノム研究の可能性

    江口傑徳

    第2回岡山大学ゲノミクス研究会  2019.11.18 

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  • The Chaperone Trio Regulates Exosomes, Tumor Malignancy, and Macrophage Polarization

    BSSR, Japan  2019.11.3 

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  • Oncosome and Tumoroid

    Takanori Eguchi

    Intercellular Communication and Extracellular Vesicles The First International Symposium  2019.11 

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  • Cancer exosomes initiate EMT involving drug resistance and tumorigenesis.

    Takanori Eguchi, Kisho Ono, Chiharu Sogawa, Yuka Okusha, Eman Ahmed Taha, Manh Tien Tran, Yanyin Lu, Kuniaki Okamoto

    The 9th International Epithelial-Mesenchymal Transition International Association meeting (TEMTIA IX), the 35th International Kumamoto Medical Bioscience Symposium  2019.11 

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  • Roles of MMP3-containing extracellular vesicles in tumorigenesis, CD9-exosome release, and cell-cell communication in cancer

    Intercellular Communication and Extracellular Vesicles The First International Symposium  2019.11 

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  • Rab11A negatively regulates osteoclastogenesis through modulating the transport route of cell surface receptors

    Intercellular Communication and Extracellular Vesicles The First International Symposium  2019.11 

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  • Monitoring of exosome using multiplexing fluorescent and bioluminescent reporter systems

    Intercellular Communication and Extracellular Vesicles The First International Symposium  2019.11 

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  • Drug repositioning using the three-dimensional tumor organoid monitoring system for anti-cancer drug screening

    Intercellular Communication and Extracellular Vesicles The First International Symposium  2019.11 

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  • Diagnostic and therapeutic potential of HSP90-exosome in oral squamous cell carcinoma

    Kisho Ono, Taka Eguchi, Keisuke Nakano, Hotaka Kawai, Akira Sasaki

    2019.10 

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  • Development of a Tumor Organoid-based Drug Screening System

    2019.10 

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  • Crucial roles of extracellular vesicles in progression and resistance in oral cancer

    Takanori Eguchi

    2019.10 

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  • The molecular chaperone trio CDC37/HSP90 is essential for oral cancer exosomes-driven malignancy and macrophage M2 polarization.

    Takanori Eguchi, Kisho Ono, Hotaka Kawai, Manh Tran Tran, Chiharu Sogawa, Yuka Okusha, Kuniaki Okamoto

    The 6th JSEV  2019.10 

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  • Crucial roles of CDC37 and HSP90 in the resistance and microenvironment in tumor malignancy

    Takanori Eguchi

    2019.9.6 

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  • 高転移性癌細胞で高発現するMMP3が有するCtgf/Ccn2発現調節機能と細胞外小胞の関連

    奥舎 有加, 江口 傑徳, タハ・エマン, チャン・チェン・マン, 十川 千春, 青山絵里子, 滝川正春, 岡元 邦彰

    第11回日本CCNファミリー研究会  2019.8.31 

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  • Roles of MMP3-containing extracellular vesicles in tumorigenesis, CD9-exosome release, and cell-cell communication in cancer

    Eman A. Taha, Yuka Okusha, Chiharu Sogawa, Abdellatif El-Seoudi, Satoshi Kubota, Ayano Satoh, Kuniaki Okamoto, Taka Eguchi

    2019.8.31 

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  • The molecular chaperone trio CDC37/HSP90 is essential for oral cancer exosomes-driven malignancy and macrophage M2 polarization.

    Takanori Eguchi, Kisho Ono, Hotaka Kawai, Manh Tien Tran, Chiharu Sogawa, Yuka Okusha, Kuniaki Okamoto

    The 11th JARI  2019.8.28 

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  • Roles of MMP3 in cargo signature and transmissive activity of cancer extracellular vesicles and in tumorigenesis

    Eman A. Taha, Takanori Eguchi, Yuka Okusha, Chiharu Sogawa, Abdellatif El-Seoudi, Satoshi Kubota, Ayano Satoh, Kuniaki Okamoto

    The 11th JARI  2019.8.28 

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  • Anti-tumor effect by regulation of HSP90 / CDC37

    Takanori Eguchi

    Seminar at Sunshine Hospital Victoria University, Melbourne  2019.3.29 

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  • How do Exosomes promote Carcinogenesis?

    Takanori Eguchi

    Special Cancer Biology Seminar at QIMRB Medical Research Institute, Brisbane  2019.3.27 

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  • Exosomal and Transcriptional Roles of MMP in Cancer

    Takanori Eguchi

    Special Guest Lecture at the University of Sydney  2019.3.25 

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  • Depletion of cholesterol lipid efflux pump ABCG1 triggers accumulation of exosomes and regression of tumors

    Takanori Eguchi, Chiharu Sogawa, Yuri Namba, Yuka Okusha, Hotaka Kawai, Kisho Ono, Mami Itagaki, Jun Murakami, Kazumi Ohyama, Junichi Asaumi, Kuniaki Okamoto

    The 92nd Annual Meeting of the Japanese Pharmacological Society  2019.3 

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  • Canonical and Novel Roles for Matrix Metalloproteinases

    Eguchi, Takanori

    BioScience Retreat in Tosa 2017  2018.12.3 

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  • Tumoroid Never Knows? Tumoroids and Exosomes are New Hopes for Refractory Cancer Research and Therapeutics

    Eguchi, Takanori

    The 41st Annual Meeting of the Molecular Biology Society of Japan  2018.11.30 

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  • RASP: 癌のストレス抵抗性と小胞/HSP分泌特性

    Eguchi, Takanori

    2018 Bioscience Retreat in Awaji  2018.11.24 

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  • Secretion of Extracellular Vesicles with Heat Shock Proteins by Resistant Adenocarcinomas and Metastatic Oral Cancer International conference

    Eguchi, Takanori

    Cold Spring Harbor Asia  2018.11.16 

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  • Organoids with Cancer Stem Cell-like Properties Secrete EpCAM-Exosomes and H SP90 in a 3D NanoEnvironment

    Eguchi, Takanori

    Japanse Cancer Association, 77th Annual Meeting  2018.9.28 

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  • The intranuclear PEX domain of MMP involves proliferation, migration, and metastasis of aggressive adenocarcinoma cells

    Eguchi, Takanori

    Japanse Cancer Association, 77th Annual Meeting  2018.9.28 

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  • Roles for cancer EVs in antibody therapeutic resistance and epithelial-to-mesenchymal transition

    Eguchi T, Ono K, Fujiwara T, Sogawa C, Calderwood SK

    The 10th JARI Annual Meeting / The 5th JSEV Annual Meeting  2018.8.30 

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  • MMP-CCN Regulatory Axis is a Common Key to Cancer and Cartilage Metabolisms. (Symposium)

    Eguchi, Takanori

    CCN family society Japan, 10th Annual Meeting  2018.8.25 

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  • RASP: Resistance-associated secretory phenotype in refractory cancer (Symposium)

    Eguchi, Takanori

    Biomedical society for stress response  2018 

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    Presentation type:Symposium, workshop panel (nominated)  

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  • Basic, latest findings, and future prospects of Exosomes

    Eguchi, Takanori

    ARCOCS seminar  2018 

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  • がんをなくす:New Approaches Revealing Novel Secretory Phenotype, Pathology, and Therapeutic Targets in Resistant / Refractory Cancer (ベストプレゼンテーション賞受賞)

    Eguchi, Takanori

    Brainstorming2018  2018 

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  • Roles for HSP90 and MMP3 in tumor resistance and metastasis

    Eguchi T

    2018 

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  • A novel HSP-inducing pathway via MMP3-heterochromatin protein interaction in cancer and inflammatory diseases

    Eguchi, Takanori

    Biomedical society for stress response, 12th annual meeting  2017.11.5 

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  • Intracellular role of Matrix Metalloproteinases

    Eguchi, Takanori

    Japanese Association for Oral Biology, annual meeting  2017.9.17 

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  • MMP3 interaction with heterochromatin proteins regulate HSP gene expression

    Eguchi, Takanori

    2017.8.26 

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  • Challenging drug discovery using 3D culture technology

    Eguchi, Takanori

    ARCOCS seminar  2017.2.15 

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  • Challenging mechanism underlying resistant cancer using trans-omics

    Eguchi, Takanori

    第4回次世代がんインフォマティクス研究会  2016.12.16 

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  • MZF1 and SCAND1 Control EMT, Oncogenic Signaling and Molecular Chaperone Expression

    Eguchi, Takanori

    Molecular Biology Society of Japan, the 41st annual meeting  2016.11.30 

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  • 癌細胞ストレス耐性における細胞内MMP3の役割

    江口 傑徳

    第37回岡山歯学会学術集会  2016.10.16 

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  • SCAN-zinc finger transcription factors control EMT, oncogenic kinase signaling pathways and molecular chaperone expression in prostate cancer cells

    Eguchi, Takanori

    Japanese Cancer Association, the 75th Annual Meeeting of the JCA  2016.10.6 

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  • OstemiR: A Novel Panel of MicroRNA Biomarkers in Osteoblastic and Osteocytic Differentiation from Mesencymal Stem Cells.

    Eguchi, Takanori

    American Society for Biochemistry and Molecular Biology / Experimental Biology 2015  2015.3 

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  • Intra-nuclear MMP-3 controls transcription of HSP70 gene through interaction with heterochromatin proteins

    Eguchi, Takanori

    American Society for Biochemistry and Molecular Biology / Experimental Biology 2015  2015.3 

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  • 転写コンポーネントによる分子シャペロンおよび腫瘍の制御(講演) Invited

    江口 傑徳

    Molecular Biology of Calcified Tissueセミナー / BioForum@Dental School 合同講演会  2014.3.18 

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  • 間葉系幹細胞の骨細胞分化における新規の マイクロRNAバイオマーカー(講演)

    江口 傑徳

    Molecular Biology of Calcified Tissueセミナー  2013.6.3 

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  • Control of Cdc37/Hsp90 and kinase signaling in prostate cancer by SCAN domain proteins. Invited

    Eguchi, Takanori

    The 11th International Congress of Hyperthermic Oncology, The 29th Japanese Congress of Thermal Medicine.  2012.8 

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  • 岡山歯学会賞受賞講演

    江口 傑徳

    2009.11.8 

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  • 日本軟骨代謝学会賞受賞講演

    江口 傑徳

    2009.3.7 

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  • Novel transcriptional Regulation of CCN2/CTGF by Nuclear Translocation of MMP3 (ICCNS Springer Scholarship Award Presentation)

    Eguchi, Takanori

    The 5th Workshop for International CCN Society  2008.10.22 

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Awards

  • Braingstorming2018 Best Presentation Award

    2018.9  

    Takanori Eguchi

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  • Japanese Society of Cartilage Metabolism Award

    2009  

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  • International CCN Society Springer Scholarship

    2008  

    Takanori Eguchi

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  • 岡山歯学会優秀論文賞

    2020.11  

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  • 生物学研究奨励賞

    2016.10   両備檉園記念財団  

    江口 傑徳

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  • 岡山歯学会奨励論文賞

    2009  

    江口傑徳

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Research Projects

  • Development of In Vitro Diagnostic Agents for Intractable Cancer Using Extracellular Vesicles

    2024.04 - 2025.03

    Japan Agency for Medical Research and Development  橋渡し研究プログラム  Seeds A

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    Authorship:Principal investigator 

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  • Roles of cancer extracellular vesicles in hijacking macrophages and metastatic niche formation

    Grant number:23H03310  2023.04 - 2026.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research  Grant-in-Aid for Scientific Research (B)

    江口 傑徳

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    Grant amount:\18720000 ( Direct expense: \14400000 、 Indirect expense:\4320000 )

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  • Understanding MMP3-rich exosomes on neoplastic cell communication in tumor microenvironment

    Grant number:22F22409  2022.09 - 2025.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for JSPS Fellows  Grant-in-Aid for JSPS Fellows

    江口 傑徳, SHETA MONA

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    Grant amount:\2300000 ( Direct expense: \2300000 )

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  • MZF1-Driven Cancer Stem-Like Resistance Against Cell Stress

    Grant number:17K11642  2017.04 - 2021.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    Eguchi Takanori

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    Authorship:Principal investigator  Grant type:Competitive

    Grant amount:\4680000 ( Direct expense: \3600000 、 Indirect expense:\1080000 )

    Refractory cancers are highly resistant to cellular stress. In this study, we demonstrated: 1) 3D cell aggregative structures of 66 cancer cell types, 2) cancer stem-like properties of metastatic prostate cancer cell-derived organoid, 3) stress-responsive production of extracellular heat shock protein (eHSP) and exosome, 4) reciprocal regulatory mechanism of CDC37 gene by oncogenic transcription factor MZF1 and tumor-suppressing transcription factor SCAND1, 5) “stresssome” including eHSP90 and damaged membrane vesicles, 6) eHSPs as prognostic markers in exosomes in metastatic oral carcinoma, 7) roles of metastatic cancer-derived exosomes in cancer progression and microenvironment such as M2-type macrophages, and 8) a triple knockdown method as a novel therapeutic strategy targeting refractory metastatic cancers.

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  • 腫瘍微小環境の細胞間コミュニケーションにおけるMMP3含有エクソソームの機能解明

    Grant number:22KF0268  2023.03 - 2025.03

    日本学術振興会  科学研究費助成事業 特別研究員奨励費  特別研究員奨励費

    江口 傑徳

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    Grant amount:\1300000 ( Direct expense: \1300000 )

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  • エクソソームに搭載される新規キナーゼの癌促進機能の解明

    2022.08 - 2023.03

    公益財団法人ウエスコ学術振興財団  公益財団法人ウエスコ学術振興財団研究助成金 

    江口傑徳

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  • エクソソームに搭載される新規キナーゼの癌促進機能の解明

    2022.08 - 2023.03

    公益財団法人ウエスコ学術振興財団  公益財団法人ウエスコ学術振興財団研究助成金 

    江口傑徳

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    Authorship:Principal investigator  Grant type:Competitive

    Grant amount:\400000 ( Direct expense: \400000 )

  • Molecular mechanisms of the inhibition of placenta and fetus development induced by periodontal disease

    Grant number:22H03511  2022.04 - 2026.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)  Grant-in-Aid for Scientific Research (B)

    岡村 裕彦, 十川 千春, 江口 傑徳, 大森 一弘, 福原 瑶子, 池亀 美華

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    Authorship:Coinvestigator(s) 

    Grant amount:\17160000 ( Direct expense: \13200000 、 Indirect expense:\3960000 )

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  • 四肢長管骨骨折の偽関節・骨欠損に対する新規エクソソーム・マスカレ再生療法の開発

    Grant number:21K20992  2021.08 - 2023.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Research Activity Start-up  Grant-in-Aid for Research Activity Start-up

    福岡 史朗

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    Grant amount:\3120000 ( Direct expense: \2400000 、 Indirect expense:\720000 )

    マウスマクロファージ様細胞(RAW264.7)を培養し、試薬(IL-4)を添加することでM2マクロファージに分極させ、免疫細胞染色にて確認した。分化させたM2マクロファージからサイズ排除クロマトグラフィー法を用いてエクソソームを抽出し、ウェスタンブロットにてエクソソームマーカー(CD9、63)を、透過電子顕微鏡、静的光散乱ナノサイズ粒子分析器にてエクソソームのサイズを確認することでエクソソームと同定した。
    実際の臨床で行ったMasquelet法により採取したInduced membraneをHE染色を行ったところ、マクロファージ、特にM2マクロファージ優位に浸潤を認め、同様にエクソソームを抽出することが出来た。
    またvivoではラットから骨髄を採取し、MCSFを添加することでM2マクロファージに分極させ、同様にエクソソームを抽出することができた。

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  • 細胞外小胞の口腔トロピズムを基軸とする侵襲性歯周炎の病態解明と診断への応用展開

    Grant number:21H03119  2021.04 - 2025.03

    日本学術振興会  科学研究費助成事業 基盤研究(B)  基盤研究(B)

    山本 直史, 井手口 英隆, 宮地 孝明, 江口 傑徳, 江國 大輔, 高柴 正悟

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    Grant amount:\16900000 ( Direct expense: \13000000 、 Indirect expense:\3900000 )

    侵襲性歯周炎(Aggressive periodontitis:AgP)は全身的には健康な若年者に発症し,急速に進行する特殊な歯周炎であるが、その発症病態は不明なままである。本研究では,臓器特異的な作用(臓器トロピズム)が近年注目されている血中の細胞外小胞(EV)とAgPの病態関与の可能性を調べた。
    今年度は、AgP患者6名と健常者3名の初診時血中EVから,AgPで高発現するmiRNAをRNAシーケンスにて調べ,マーカー候補となるそれらのmiRNA mimicをヒト歯肉線維芽細胞と歯周炎モデルマウスに遺伝子導入した。誘導された炎症性サイトカインの発現量をリアルタイムPCR法とELISA法にて測定し,歯槽骨吸収量をマイクロCTにて調べた。
    健常者と比較して,AgP患者で発現量が2倍以上増加したmiRNAを500種類以上同定した。それらのうち5種のmiRNAとmiR-181b-5pを歯肉線維芽細胞に導入すると,IL-6とIL-1βの産生が増加した。とりわけ,miR-181b-5p を歯肉組織に導入すると歯槽骨吸収が進行した。
    すなわち、AgP患者の血中EVには診断マーカー候補となるmiRNAが多く発現しており,miR-181b-5pはIL-6とIL-1β発現を伴う炎症を助長することによってAgPを重症化する可能性が示された。

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  • Exploration of the novel therapeutic target for lung cancer based on the analysis of the characteristics of tumor cells derived from 3D culture

    Grant number:21K08902  2021.04 - 2024.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    山本 寛斉, 豊岡 伸一, 冨田 秀太, 諏澤 憲, 江口 傑徳, 加藤 竜司

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    Grant amount:\4160000 ( Direct expense: \3200000 、 Indirect expense:\960000 )

    近年、免疫チェックポイント阻害剤の登場により、がん細胞周囲の微小環境(腫瘍微小環境)が治療標的として注目されている。三次元(3D)培養での腫瘍細胞は二次元(2D)培養と比べ、細胞間インタラクションが生体内に近く、腫瘍微小環境を反映していることが知られている。また、がん難治性に関与する幹細胞性の維持や治療抵抗性の評価にも従来の2D 培養によるアッセイ系よりも適している。申請者らは、その中でも腫瘍細胞の凝集形態には多様性があり、特にがんの悪性度が高い集団が存在することを発見した。本研究は、より生体内の環境を反映する3D 培養により肺がん細胞の特性を評価し、悪性度と関連する遺伝子を同定し、これを標的とする治療戦略の確立を目指すものである。
    令和3年度は、肺がん細胞株 (n = 30) を用いて3D 培養による細胞凝集塊の形態学的特徴の確認を進め、Grape-like, Spheroid, Monolayer sheet, Other typeの4つに分類した。また、腫瘍学的特徴を明らかにするために、これらの形態学的特徴と増殖能・浸潤能・遊走能の表現型との関連について解析を進めたところ、Monolayer sheet typeの細胞株は浸潤能・遊走能がSpheroid typeの細胞株よりも高いという結果であった。また、各肺がん細胞株のin vivoでの腫瘍形成能をマウスモデルを用いて検討を進めている。

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  • Pioneering chondroneutrigenomics research and its development into chondroneutrigenetics

    Grant number:20K20611  2020.07 - 2024.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Research (Pioneering)  Grant-in-Aid for Challenging Research (Pioneering)

    滝川 正春, 青山 絵理子, 星島 光博, 久保田 聡, 西田 崇, 江口 傑徳

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    Grant amount:\25870000 ( Direct expense: \19900000 、 Indirect expense:\5970000 )

    1.昨年度メチオニンの代謝産物であるS-アデノシルメチオニン(SAM)をヒト軟骨細胞様細胞株HCS-2/8の培養系に添加すると、まずCCN2の遺伝子発現が亢進し、次いで2型コラーゲンの遺伝子発現が上昇し、その後、アグリカンの蓄積量(アルシアンブルー染色)も増加すること、また、ポリアミンの前駆体の一つSAM脱炭酸物を合成するSAM 炭酸酵素AMD1の阻害剤、SardomizideをSAMと共に添加するとSAMによるアグリカンの蓄積が抑制されることを見いだした。今年度はこれらの知見を、染色の場合は生化学的手法で測定するなど他の手法を用いて再確認するとともに、1培養細胞株では不十分との考えのもと、ラット軟骨肉腫由来の軟骨細胞様細胞株RCS細胞を用いて確認した。これらの結果はSAMがCCN2の発現を誘導する機能分子であることを示している。また、同培養系にSAMを添加して、スペルミジン、スペルミン等のポリアミンレベルをHPLCで測定すると、両ポリアミン濃度の増加が見られた。従って、SAMは少なくとも一部はポリアミン合成を介して軟骨細胞の分化機能を亢進させることを示唆している。
    2.CCN2が関節軟骨形成因子GDF5と結合することはすでに報告済みであるが、CCN2はGDF5とBMPRIbとの結合には影響しないこと、NogginはCCN2のGDF5への結合を阻害することを見いだした。また、CCN2は、軟骨細胞においてGDF5によるSmad1/5/8のリン酸化を増強し、アグリカン遺伝子発現促進作用をさらに増強した。
    3.齧歯類の変形性関節症の予防・修復作用を有するCCN2と「陰と陽」の関係があるとされているCCN3の発現が、ヒト変形性肩関節症および変形性股関節症の症状と正に相関することを明らかにした。2と3の知見は本課題後半のコンドロニュートリジェネティクス研究に繋がる重要な基礎的知見となる。

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  • 口腔扁平上皮癌における新規遠隔転移抑制治療の開発

    Grant number:20H03888  2020.04 - 2025.03

    日本学術振興会  科学研究費助成事業 基盤研究(B)  基盤研究(B)

    長塚 仁, 山近 英樹, 中野 敬介, 河合 穂高, 江口 傑徳, 辻極 秀次, 高畠 清文

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    Grant amount:\17030000 ( Direct expense: \13100000 、 Indirect expense:\3930000 )

    本年度は、リンパ節における転移niche形成の検討を行うために、GFP骨髄移植を行なったマウスにヒト好転移ヒト扁平上皮癌細胞株を頬粘膜に移植して、リンパ節転移モデルの作成を行なった。腫瘍組織は、通報に従い標本を作成し、免疫組織化学染色、蛍光免疫二重染色、蛍光免疫多重染色を行い検討を行なった。
    ●蛍光免疫多重染色(Opal 7 color manual IHC)による目的細胞の組織内での局在の検討
    転移に関与するBMDCのプロファイルがある程度特定されれば、次に蛍光免疫多重染色(Opal 7 color manual IHC)を用いることにより、目的BMDCの組織での局在を明らかとする。Opal 7 color manual IHCは、同一切片内で最大7色までの多重染色を行うことが可能な染色手法で、FACSで得られた情報をそのまま切片内に再現することが可能である。これにより、組織内での目的BMDCの局在や動態の詳細を継時的に観察した。

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  • 三次元腫瘍オルガノイド評価系により見出された新規癌転移抑制化合物の創薬展開

    Grant number:20K09904  2020.04 - 2023.03

    日本学術振興会  科学研究費助成事業 基盤研究(C)  基盤研究(C)

    十川 千春, 岡元 邦彰, 江口 傑徳, 河合 穂高, 青山 絵理子

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    Grant amount:\4420000 ( Direct expense: \3400000 、 Indirect expense:\1020000 )

    癌の悪性化、転移および再発を制御するには、分子機序の解明と有効な抗腫瘍薬の開発が不可欠である。近年、癌の悪性化には癌細胞を取りまく腫瘍微小環境が影響するといわれ、癌細胞のみならず腫瘍微小環境を制御する因子も標的分子の候補に入れた検討が必要である。本研究課題では、三次元腫瘍オルガノイド形成とMMP9発現をモニタリングすることによる、独自の多元薬物評価系によって見出したヒット化合物をもとに、新規癌転移抑制薬開発に向けた創薬展開を行うことを目的とする。
    令和3年度は、これまで得られたヒット化合物のさらなるブラッシュアップのため、腫瘍微小環境に対する薬剤の効果を評価可能な高次アッセイ系の構築を引き続き行った。エクソソームをはじめとする細胞外小胞(Extracellular Vesicles: EV)は腫瘍微小環境制御に関わるとされるが、免疫系細胞が分泌するEVの腫瘍細胞に対する影響について、昨年度確立した蛍光または発光モニタリングシステムを応用することにより検討した。マクロファージ様に分化させたTHP-1細胞から分泌されたEVをサイズ排除クロマトグラフィーを用いて粒子径により分画し、CD9陽性の大型EV(80-300 nm)とCD63/HSP90陽性小型EV(20-200 nm)を得た。蛍光標識したこれらのEVは口腔癌細胞HSC-3に効率的に取り込まれ、口腔癌細胞の生存率を大幅に低下させることが明らかとなった。
    以上のことから、マクロファージ様細胞におけるEVの分泌を促進する薬剤は抗がん作用を発揮する可能性が高く、腫瘍微小環境に対する薬剤の効果を検討する上で重要であると考えられた。

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  • 細胞外小胞による生体ネットワークの解明と医療への応用 International coauthorship

    2020 - 2021

    岡山大学次世代研究育成グループ 

    江口傑徳

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  • The role of extracellular vesicles of oral bacteria in systemic disease

    Grant number:19H04051  2019.04 - 2023.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)  Grant-in-Aid for Scientific Research (B)

    岡村 裕彦, 江口 傑徳, 宝田 剛志, 吉田 賀弥, 池亀 美華, 江國 大輔, 伊原木 聰一郎

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    Grant amount:\17160000 ( Direct expense: \13200000 、 Indirect expense:\3960000 )

    歯周病原菌によって惹起される歯周病は,慢性炎症をともなう生活習慣病である。歯周病の病態の悪化が糖尿病などの全身性疾患の重症化に関与することが明らかになってきたが,現在でも高齢者を中心に8割以上の人々が何らかの歯周疾患を患っている。口腔は全身状態を示す鏡であり,健全な歯と口腔を維持することは,全身の健康にとって重要と認識されながらも,現状との間には依然として乖離がある。この原因として,歯周病と全身性疾患の重症化を関連づける明確な分子生物学的根拠が乏しいことが挙げられる。我々は,これらの疾患を関連づける新たな因子として歯周病原菌由来の『細胞外分泌小胞』に注目した。
    当該年度は,1.歯周病原菌由来の『細胞外分泌小胞』の組織・細胞障害性を調べる。2.細胞外分泌小胞』に含まれる病原因子を同定し,その細胞障害性について調べることを目的とした。
    歯周病原菌由来の『細胞外分泌小胞』を標識し,生体内での動向を可視化することに成功した。『細胞外分泌小胞』は,肝臓を含む遠隔臓器に集積した。また,このマウスではインスリンに対する応答性が減弱し,高いレベルの血糖値を示した。培養細胞を用いた実験により,『細胞外分泌小胞』は肝細胞において糖代謝に関わるシグナル伝達経路を阻害することが分かった。回収した『細胞外分泌小胞』からタンパク質を抽出し,質量分析により解析したところ,菌固有のタンパク質分解酵素などが含まれていた。以上の結果より,歯周病原菌は『細胞外分泌小胞』を介して細胞障害性因子を肝臓に到達させ,肝細胞の糖取り込みを抑制することで,糖尿病の重症化に関与すると考えられる。

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  • Intracellular function and new extracellular signaling pathways of CCN proteins and their common molecular base

    Grant number:19H03817  2019.04 - 2023.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)  Grant-in-Aid for Scientific Research (B)

    滝川 正春, 青山 絵理子, 星島 光博, 久保田 聡, 西田 崇, 江口 傑徳, 大野 充昭, 鈴木 守

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    Authorship:Coinvestigator(s) 

    Grant amount:\17290000 ( Direct expense: \13300000 、 Indirect expense:\3990000 )

    サブテーマ1: CCNタンパク質の意外な新機能について。1-1)CCN2結合因子として分泌小胞の細胞内輸送に関与するタンパク質Rab14をyeast two-hybrid法で同定し、両者がCCN2のIGFBPドメインを介して結合することを解明した。また、Cos7細胞に両タンパク質を強制発現して、両者が細胞内で共局在することを確認した。さらに、軟骨細胞においてRab14あるいはCCN2の発現を阻害すると小胞体ストレスマーカーの発現が上昇すること、Rab14とCCN2の相互作用が軟骨細胞のアグリカン分泌に重要であることも解明した。即ち、分泌タンパク質CCN2が細胞内で機能するという意外な事実を見いだした。1-2)CCN2 が神経栄養因子様の活性を持っていることを新たに見いだした。また、脂肪細胞分化を抑制すること、骨細胞から産生され骨のリモデリングに重要な役割を果たすことなどCCN2の新機能を見いだした。
    サブテーマ2: CCNタンパク質の細胞外小胞を介した情報ネットワークの形成について。好転移性がん細胞LuM1が産生する細胞外ベジクル (EV)が低転移性大腸がん細胞Colon26のEVよりMMP3とCCN2を多量に含有すること、このEVが血流を介して、皮下に移植したColon26癌の増殖を強く増強することを見いだした。また、 同オンコゾーム由来のMMP3が標的細胞中でCCN2を誘導することを見いだした。
    サブテーマ3: 構造ー機能解析とその展開について。ヒトCCN2全長タンパク質の立体構造を決めるため、ヒトCCN2組換えタンパク質を用いて、BINDS(創薬等先端技術支援基盤プラットホーム) の支援のもと、約800条件で結晶化を試みたが結晶は確認できず、また、FGF2との複合体の結晶化も進めるべく、同複合体の分離を試みたが、沈殿が生じてしまった。現在、これらの条件検討を継続中である。

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  • 細胞間情報伝達・細胞外小胞研究会第一回国際シンポジウム(ICEV-1)開催

    2019 - 2020

    岡山大学理事裁量経費シンポジウム開催経費支援 

    江口傑徳

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  • Regulation of the biological character of cancer and induction of cancer by the cancer stroma.

    Grant number:18K09789  2018.04 - 2021.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    Nakano Keisuke

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    Authorship:Coinvestigator(s)  Grant type:Competitive

    Grant amount:\4550000 ( Direct expense: \3500000 、 Indirect expense:\1050000 )

    The purpose of this study is to verify that the stroma of oral cancer can act on cancer cells to alter their biological properties and that the tumor stroma can act on the normal cells surrounding the cancer to cause tumorigenesis. The effects of the combination of oral squamous cell carcinoma cell lines and stromal cells in culture and tumor stroma on the biological character of tumor cells (proliferative capacity, invasive capacity, migratory capacity, and morphology) were examined histopathologically. The results showed that tumor tissues significantly change their properties depending on differences in tumor stroma. The results of this study strongly support that cancer stroma directly modulates the biological character of cancer.

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  • 難治性がん克服のための腫瘍オルガノイドを利用した創薬研究

    2018 - 2019

    公益財団法人鈴木謙三記念医科学応用研究財団  生活習慣病における医学,薬学の萌芽的研究 

    江口 傑徳

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    Authorship:Principal investigator  Grant type:Competitive

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  • Australia Tour Revealing Exosome Code

    2018 - 2019

    Supporting Advance Activities of Key YoUng Researchers 

    Takanori Eguchi

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  • Development of Novel Assay Systems to Evaluate Drug Sensitivity and Resistance that Reflects Tumor Heterogeneity

    Grant number:17K11669  2017.04 - 2021.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    Ohyama Kazumi

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    Grant amount:\4550000 ( Direct expense: \3500000 、 Indirect expense:\1050000 )

    We aimed to establish a novel system to evaluate drug sensitivity and resistance that reflects intratumoral heterogeneity. We demonstrated: 1) a novel cancer exosome-driven epithelial-to-mesenchymal transition (EMT), 2) pro-tumorigenic roles of the lipid excretion pump ABCG1 and hypoxic inducible factor 1 alpha (HIF-1α) in the intratumoral hypoxic environment in three-dimensional cancer organoids, 3) a novel exosome-driven drug resistance mechanism by which antibody-based drugs such as anti-EGFR medication (cetuximab) is secreted with exosomes from resistant cancer cells, and 4) gene expression profile of cancer stem cell-like organoids and drug resistance to cisplatin.

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  • Drug repositioning using the gene promoter activity-based anticancer drug screening system

    Grant number:17K11643  2017.04 - 2021.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    Sogawa Chiharu

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    Grant amount:\4550000 ( Direct expense: \3500000 、 Indirect expense:\1050000 )

    In the present study, we aimed to develop a novel reporter system evaluating tumorigenesis, invasiveness, metastasis, and druggability. High expression and genetic amplification of matrix metalloproteinase 9 (MMP9) were found in rapid metastatic colon cancer cases. Furthermore, the properties of three-dimensional (3D) tumor-like organoids (tumoroids) more closely resemble in vivo tumors. We screened the pharmacologically active compounds using an original tumoroid-based multiplex phenotypic screening system with an MMP9 promoter-driven fluorescence reporter to evaluate tumoroid formation and progression. The anti-Parkinson drug benztropine was the most effective compound uncovered by the screen. Benztropine significantly inhibited in vitro tumoroid formation, cancer cell survival, and MMP9 promoter activity. Benztropine also inhibited the tumor growth, circulating tumor cell number, and rate of metastasis in a tumor allograft model in mice.

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  • ストレス応答細胞間コミュニケーション研究コア形成

    2017 - 2018

    岡山大学若手研究者育成支援事業 

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  • Development of prognosis method for oral cancer prognosis with body fluid diagnosis using exosome as an index

    Grant number:16K11722  2016.04 - 2020.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    Murakami Jun

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    Grant amount:\4550000 ( Direct expense: \3500000 、 Indirect expense:\1050000 )

    Cetuximab (brand name: Erbitux) is an anti-epithelial growth factor receptor (EGFR) targeted anti-cancer drug. In Japan, it was approved for squamous cell carcinoma of the head and neck in 2012. Cetuximab binds to EGFR, inhibits receptor function, and exhibits an anticancer effect. The mechanism of resistance of new anticancer drugs via extracellular vesicles has been clarified. Therefore, we also reached the idea that extracellular vesicles might be involved in the resistance to cetuximab exhibited by oral cancer cells, and examined the mechanism of resistance to cetuximab mediated by extracellular vesicles secreted by cavity cancer cells.

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  • Development of Supplement having anti-cancer action based on Flavonoid

    Grant number:16K11863  2016.04 - 2019.03

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)  Grant-in-Aid for Scientific Research (C)

    OKAMOTO Kuniaki

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    Grant amount:\4680000 ( Direct expense: \3600000 、 Indirect expense:\1080000 )

    Lots of researchers reported that some flavonoids have an anti-cancer effect in vitro. But any effects of osteoclast and osteoblast correlated bone invasion and metastasis of cancer cells remain to investigate. In this research, we classified flavonoid into six gropes from its structure and analyzed the effect of each flavonoid in osteoclastogesesis. In result, Liquiritigenin, a component obtained from a Chinese herb, showed a significant inhibition effect during osteoclastogenesis. On the other hand, it did not inhibit osteoblast formation. Liquiritigenin is an aglycone of liquiritin, and it is one of the flavonoids present in Glycyrrhizae radix known as one of Kampo drug. Liquiritigenin has been shown to have various pharmacological effects, such as antioxidative, antitumor, and anti-inflammatory effects. Our data suggest that liquiritigenin may act as an inhibited supplement of osteoclasts in bone diseases such as osteoporosis.

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  • MMPを介した腫瘍細胞リプログラミングの制御の試み

    2016 - 2017

    公益財団法人両備檉園記念財団 

    江口 傑徳

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  • Regulation of HSPs in Cancer by MMP3 and Heterochromatin Protein 1

    2015.01 - 2015.12

    Harvard Medical School Joint Center for Radiation Therapy Foundation Grant 

    Eguchi, Takanori

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  • Research and Development of Periodontal Bone-Regenerative Method and Agent

    2010

    MHLW  Incentive for young scientists 

    EGUCHI Takanori

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  • 歯周病治療薬と歯槽骨再生方法の開発

    2010

    厚労省  創薬基盤推進研究事業 

    江口 傑徳

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  • Strategy for treatment of periodontitis with regulating exocytosis using functional bionano particle

    Grant number:21659436  2009 - 2011

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research  Grant-in-Aid for Challenging Exploratory Research

    MATSUSHITA Kenji, EGUCHI Takanori

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    Grant amount:\3360000 ( Direct expense: \3000000 、 Indirect expense:\360000 )

    We explored factors for intracellular vesicle trafficking in gingival epithelial cells and examined a possibility of the regulation. A small G-protein Rab5 and the associated protein vinculin were identified as important factors for vesicle trafficking in the cells. Rab5 and ICAM-1 were associated with uptake of Porphyromonas gingivals by the cells. Furthermore, we clarified that E-selectin is important for regulating endothelial exocytosis by stimulation with P. gingivalis.

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  • Matrix metalloproteinase-3 in a stable form, a novel medicament for pulpitis

    Grant number:21390512  2009 - 2011

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)  Grant-in-Aid for Scientific Research (B)

    NAKAMURA Hiroshi, NAKASHIMA Misako, NAKATA Kazuhiko, EGUCHI Takanori

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    Grant amount:\18070000 ( Direct expense: \13900000 、 Indirect expense:\4170000 )

    In this study, the canine model of irreversible pulpitis was established to examine the effect of MMP-3 in the inflamed pulp tissues. Regeneration of pulp tissue with vasculature and nerves was observed in this pulpitis model 14 days after MMP-3 treatment, followed by dentin matrix formation on the exposed pulp tissues by day 28.In the absence of MMP-3, whole pulp necrosis was observed on day 14 to 28.Infiltration of macrophage and antigen-presenting cells was significantly inhibited in MMP-3 treated pulp tissues compared with control on day 3 to 7.IL-6 expression was significantly decreased as early as day 3 in MMP-3 treated pulp. These results suggest that MMP-3 has anti-inflammatory effect and is promising drug candidates for irreversible pulpitis.

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  • 長寿社会への布石: 水平性骨吸収に対する歯槽骨再生方法の開発

    2009

    武田科学振興財団  医学系研究奨励(生活習慣病) 

    江口傑徳

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  • 安定型マトリックスメタロプロテナーゼ3を用いた新しい歯髄炎治療薬の開発

    2009

    文科省  科研費 基盤研究B 

    中村洋

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  • 歯髄幹細胞を用いた象牙質・歯髄再生医療によるウ蝕・歯髄疾患等のための治療技術の開発

    2009

    国立長寿医療センター  長寿委託費 

    中島美砂子

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    Grant type:Competitive

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  • Fuseki for Longevity Society: Development of Bone Regenerative Medicine Targetting Holizontal Loss of the Periodontal Bone

    2009

    Takeda Foundation  Incentive for young scientists 

    EGUCHI Takanori

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  • Evaluation of Physiological and Pathological Properties of MMP via Molecular Interactions

    2008 - 2010

    MEXT  Grant in aid for young scientists (B) 

    EGUCHI Takanori

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  • 新規分子間相互作用を介したMMPの生理的・病理的機能発現機序の解明

    2008 - 2009

    文科省  科研費 若手研究 (B) 

    江口傑徳

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  • Physiological and Pathological Function of MMP3 by a Novel Molecular Interaction

    Grant number:20791378  2008 - 2009

    Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research Grant-in-Aid for Young Scientists (B)  Grant-in-Aid for Young Scientists (B)

    EGUCHI Takanori

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    Grant amount:\4160000 ( Direct expense: \3200000 、 Indirect expense:\960000 )

    マトリックス・メタロ・プロテアーゼ3(MMP3)は細胞外マトリックスや成長因子、そのレセプターを切断することで、組織リモデリング、血管新生、創傷治癒、骨・軟骨リモデリングに関与し、病理的にはリウマチのマーカーでもある。そこにきて我々は、MMP3が核移行し、転写因子としてCTGF/CCN2遺伝子の発現を誘導することを見出した。今回、細胞内で発現させたMMP3およびその機能ドメインであるPEXドメインおよびCatalyticドメインを細胞内で発現させその機能を解析した。全長MMP3およびそれらの変異体は細胞死を誘導することなく一部は核移行した。さらにはマイクロアレイによって、細胞内MMP3の特異的な新しいターゲット遺伝子としてHSP70B'を見出した。細胞内MMP3によってHSP70B'が誘導され、細胞がUV、感染、メカニカル・ロードなどのストレスに対して耐性を獲得すると考えられる。

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  • Identification and characterization of cartilage-specific transcription factor

    2003 - 2006

    JSPS  Incentive for JSPS postdoc 

    EGUCHI Takanori

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  • 軟骨細胞特異的な新規転写因子の単離とその機能解析

    2003 - 2006

    日本学術振興会  特別研究員奨励費 

    江口傑徳

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  • 軟骨細胞特異的な新規転写因子の単離とその機能解析

    Grant number:03J02535  2003 - 2005

    日本学術振興会  科学研究費助成事業 特別研究員奨励費  特別研究員奨励費

    江口 傑徳

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    Grant amount:\3300000 ( Direct expense: \3300000 )

    内軟骨性骨化、乳癌の骨転移、組織再生などへの強い関与が報告されているタンパク質CCN2/CTGFは、それら生命現象の実験的モデルとして多用されている細胞株HCS-2/8やMDA231でもまた高発現しており、そのメカニズムの一つがCCN2プロモーター上のエンハンサーTRENDICとタンパク質(複合体)の相互作用であることは既に我々が論文報告している。その後TRENDIC結合因子を同定するために行ったサウスウェスタンスクリーニングにより得られたcDNAの1つは細胞外基質分解酵素として知られるMMP-3をコードしていた。そこでMMP-3による転写調節について研究を行っている。
    1.軟骨組織および培養軟骨細胞様細胞株HCS-2./8の免疫染色では、MMP-3は細胞核に局在化していた。
    2.HCS-2/8の細胞成分を核・細胞質に分画し、ウェスタンブロッティングにより検討したところ、両方の画分にMMP-3が検出された。
    3.クロマチン免疫沈降により、通常培養条件下のHCS-2/8細胞においてMMP-3が染色体上のCCN2のプロモーター領域(TRENDICを含む)に結合することが分かった。
    4.TRENDICと核タンパク質複合体の結合は抗MMP-3抗体によって阻害されるというスーパーシフトアッセイの結果からも、MMP-3がTRENDICに結合することが分かる。
    5.各種細胞株で強制発現したMMP-3は細胞質および核に局在した。この強制発現MMP-3によるCCN2プロモーターの活性化は特定の細胞株でのみ認められた。
    6.HCS-2/8におけるCCN2のプロモーター活性はMMP-3インヒビターの添加により抑制された。
    7.質量分析装置を応用することでMMP-3結合性転写調節因子・核輸送因子を同定した。
    現在上記の結果を統合し、論文発表に向け準備中である。

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  • 総合歯科医学1 (2021academic year) 第1学期

  • Pharmacology 1 (2021academic year) 1st semester  - 水1,水2,水3

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