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  2. 高橋 雅英

高橋 雅英

タカハシ マサヒデ  (Masahide Takahashi)

基本情報

所属
藤田医科大学 国際再生医療センター センター長、特命教授
(兼任)研究統括監理部 特命教授 (統括学術プログラムディレクター)
学位
医学博士(名古屋大学)
連絡先
masahide.takahashifujita-hu.ac.jp
J-GLOBAL ID
200901036145308243
researchmap会員ID
1000023347

研究キーワード

 9

研究分野

 1

経歴

 5

学歴

 2

受賞

 8
もっとみる

論文

 220
  • Hyogo Naoi, Yuzo Suzuki, Asuka Miyagi, Ryo Horiguchi, Yuya Aono, Yusuke Inoue, Hideki Yasui, Hironao Hozumi, Masato Karayama, Kazuki Furuhashi, Noriyuki Enomoto, Tomoyuki Fujisawa, Naoki Inui, Shinji Mii, Masatoshi Ichihara, Masahide Takahashi, Takafumi Suda
    Journal of immunology (Baltimore, Md. : 1950) 212(7) 1221-1231 2024年4月1日  
    Pulmonary fibrosis is a fatal condition characterized by fibroblast and myofibroblast proliferation and collagen deposition. TGF-β plays a pivotal role in the development of pulmonary fibrosis. Therefore, modulation of TGF-β signaling is a promising therapeutic strategy for treating pulmonary fibrosis. To date, however, interventions targeting TGF-β have not shown consistent efficacy. CD109 is a GPI-anchored glycoprotein that binds to TGF-β receptor I and negatively regulates TGF-β signaling. However, no studies have examined the role and therapeutic potential of CD109 in pulmonary fibrosis. The purpose of this study was to determine the role and therapeutic value of CD109 in bleomycin-induced pulmonary fibrosis. CD109-transgenic mice overexpressing CD109 exhibited significantly attenuated pulmonary fibrosis, preserved lung function, and reduced lung fibroblasts and myofibroblasts compared with wild-type (WT) mice. CD109-/- mice exhibited pulmonary fibrosis comparable to WT mice. CD109 expression was induced in variety types of cells, including lung fibroblasts and macrophages, upon bleomycin exposure. Recombinant CD109 protein inhibited TGF-β signaling and significantly decreased ACTA2 expression in human fetal lung fibroblast cells in vitro. Administration of recombinant CD109 protein markedly reduced pulmonary fibrosis in bleomycin-treated WT mice in vivo. Our results suggest that CD109 is not essential for the development of pulmonary fibrosis, but excess CD109 protein can inhibit pulmonary fibrosis development, possibly through suppression of TGF-β signaling. CD109 is a novel therapeutic candidate for treating pulmonary fibrosis.
  • Ryota Ando, Yukihiro Shiraki, Yuki Miyai, Hiroki Shimizu, Kazuhiro Furuhashi, Shun Minatoguchi, Katsuhiro Kato, Akira Kato, Tadashi Iida, Yasuyuki Mizutani, Kisuke Ito, Naoya Asai, Shinji Mii, Nobutoshi Esaki, Masahide Takahashi, Atsushi Enomoto
    The Journal of pathology 2023年10月5日  
    Pancreatic stellate cells (PSCs) are stromal cells in the pancreas that play an important role in pancreatic pathology. In chronic pancreatitis (CP) and pancreatic ductal adenocarcinoma (PDAC), PSCs are known to get activated to form myofibroblasts or cancer-associated fibroblasts (CAFs) that promote stromal fibroinflammatory reactions. However, previous studies on PSCs were mainly based on the findings obtained using ex vivo expanded PSCs, with few studies that addressed the significance of in situ tissue-resident PSCs using animal models. Their contributions to fibrotic reactions in CP and PDAC are also lesser-known. These limitations in our understanding of PSC biology have been attributed to the lack of specific molecular markers of PSCs. Herein, we established Meflin (Islr), a glycosylphosphatidylinositol-anchored membrane protein, as a PSC-specific marker in both mouse and human by using human pancreatic tissue samples and Meflin reporter mice. Meflin-positive (Meflin+ ) cells contain lipid droplets and express the conventional PSC marker Desmin in normal mouse pancreas, with some cells also positive for Gli1, the marker of pancreatic tissue-resident fibroblasts. Three-dimensional analysis of the cleared pancreas of Meflin reporter mice showed that Meflin+ PSCs have long and thin cytoplasmic protrusions, and are localised on the abluminal side of vessels in the normal pancreas. Lineage tracing experiments revealed that Meflin+ PSCs constitute one of the origins of fibroblasts and CAFs in CP and PDAC, respectively. In these diseases, Meflin+ PSC-derived fibroblasts showed a distinctive morphology and distribution from Meflin+ PSCs in the normal pancreas. Furthermore, we showed that the genetic depletion of Meflin+ PSCs accelerated fibrosis and attenuated epithelial regeneration and stromal R-spondin 3 expression, thereby implying that Meflin+ PSCs and their lineage cells may support tissue recovery and Wnt/R-spondin signalling after pancreatic injury and PDAC development. Together, these data indicate that Meflin may be a marker specific to tissue-resident PSCs and useful for studying their biology in both health and disease. © 2023 The Pathological Society of Great Britain and Ireland.
  • Yuya Aono, Yuzo Suzuki, Ryo Horiguchi, Yusuke Inoue, Masato Karayama, Hironao Hozumi, Kazuki Furuhashi, Noriyuki Enomoto, Tomoyuki Fujisawa, Yutaro Nakamura, Naoki Inui, Shinji Mii, Masahide Takahashi, Takafumi Suda
    American journal of respiratory cell and molecular biology 68(2) 201-212 2022年10月10日  査読有り
    Asthma is a chronic airway inflammatory disease characterized by airway hyperreactivity (AHR) and eosinophilic airway inflammation. Dendritic cells (DCs) are essential for the development of asthma via presenting allergens, causing Th2 skewing and eosinophil inflammation. Recent studies have revealed that CD109, a glycosylphosphatidylinositol-anchored glycoprotein, is involved in the pathogenesis of inflammatory diseases such as rheumatoid arthritis and psoriasis. However, no study has addressed the role of CD109 in asthma. This study sought to address the role of CD109 on DCs in the development of AHR and allergic inflammation. CD109 deficient mice (CD109-/- mice) were sensitized with house dust mite (HDM) or ovalbumin and compared to wild-type (WT) mice for induction of AHR and allergic inflammation. CD109-deficient mice had reduced AHR and eosinophilic inflammation together with lower Th2 cytokine expression compared to WT mice. Interestingly, CD109 expression was induced in lung conventional DC2s (cDC2s), but not lung cDC1s, upon allergic challenge. Lung cDC2s from CD109-/- mice had a poor ability to induce cytokine production in ex vivo DC-T cell cocultures with high expression of RUNX3, resulting in suppression of Th2 differentiation. Adoptive transfer of bone-marrow-derived CD109-/- DCs loaded with HDM failed to develop AHR and eosinophilic inflammation. Finally, administration of monoclonal anti-CD109 antibody reduced airway eosinophils and significantly decreased AHR. Our results suggest the involvement of CD109 in asthma pathogenesis. CD109 is a novel therapeutic target for asthma.
  • 迫田 朋佳, 江崎 寛季, 安藤 良太, 宮井 雄基, 飯田 忠, 松山 誠, 白木 之浩, 三井 伸二, 西田 佳弘, 高橋 雅英, 榎本 篤
    日本癌学会総会記事 81回 E-2021 2022年9月  
  • Yuki Miyai, Daisuke Sugiyama, Tetsunari Hase, Naoya Asai, Tetsuro Taki, Kazuki Nishida, Takayuki Fukui, Toyofumi Fengshi Chen-Yoshikawa, Hiroki Kobayashi, Shinji Mii, Yukihiro Shiraki, Yoshinori Hasegawa, Hiroyoshi Nishikawa, Yuichi Ando, Masahide Takahashi, Atsushi Enomoto
    Life Science Alliance 5(6) e202101230-e202101230 2022年6月  
    Cancer-associated fibroblasts (CAFs) are an integral component of the tumor microenvironment (TME). Most CAFs shape the TME toward an immunosuppressive milieu and attenuate the efficacy of immune checkpoint blockade (ICB) therapy. However, the detailed mechanism of how heterogeneous CAFs regulate tumor response to ICB therapy has not been defined. Here, we show that a recently defined CAF subset characterized by the expression of Meflin, a glycosylphosphatidylinositol-anchored protein marker of mesenchymal stromal/stem cells, is associated with survival and favorable therapeutic response to ICB monotherapy in patients with non-small cell lung cancer (NSCLC). The prevalence of Meflin-positive CAFs was positively correlated with CD4-positive T-cell infiltration and vascularization within non-small cell lung cancer tumors. Meflin deficiency and CAF-specific Meflin overexpression resulted in defective and enhanced ICB therapy responses in syngeneic tumors in mice, respectively. These findings suggest the presence of a CAF subset that promotes ICB therapy efficacy, which adds to our understanding of CAF functions and heterogeneity.
もっとみる

MISC

 199
  • Noguchi M, Imaizumi K, Kawabe T, Wakayama H, Horio Y, Sekido Y, Hara T, Hashimoto N, Takahashi M, Shimokata K, Hasegawa Y
    Cancer Gene Therapy 8 421-429 2001年  査読有り
  • Shigemitsu, K, Sekido, Y, Usami, N, Mori, S, Sato, M, Horio, Y, Hasegawa, Y, Bader, S.A, Gazdar, A.F, Minna, J.D, Hida, T, Yoshioka, H, Imaizumi, M, Ueda, Y, Takahashi, M, Shimokata, K
    Oncogene 20 4249-57 2001年  査読有り
  • Kato M, Liu W, Akhand AA, Hossain K, Takeda K, Takahashi M, Nakashima I
    J Invest Dermatol 115(6) 1157-1158-1158 2000年12月  
  • H Hayashi, M Ichihara, T Iwashita, H Murakami, Y Shimono, K Kawai, K Kurokawa, Y Murakumo, T Imai, H Funahashi, A Nakao, M Takahashi
    ONCOGENE 19(39) 4469-4475 2000年9月  査読有り
    Glial cell line derived neurotrophic factor (GDNF) signals through a multicomponent receptor complex consisting of RET receptor tyrosine kinase and a member of GDNF family receptor alpha (GPR alpha). Recently, it was shown that tyrosine 1062 in RET represents a binding site for SHC adaptor proteins and is crucial for both RAS/mitogen activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathways. In the present study, we characterized how these two pathways diverge from tyrosine 1062, using human neuroblastoma and primitive neuroectodermal tumor cell lines expressing RET at high levels. In response to GDNF stimulation, SHC bound to GAB1 and GRB2 adaptor proteins as well as RET, and SHC and GAB1 were highly phosphorylated on tyrosine. The complex formation consisting of SHC, GAB1 and GRB2 was almost abolished by replacement of tyrosine 1062 in RET with phenylalanine. Tyrosine-phosphorylated GAB1 was also associated with p85 subunit of PI3-K, resulting in PI3-K and AKT activation, whereas SHC-GRB2-SOS complex was responsible for the RAS/ERK signaling pathway. These results suggested that the RAS and PI3K pathways activated by GDNF bifurcate mainly through SHC bound to tyrosine 1062 in RET. Furthermore, using luciferase reporter-gene assays, we found that the RAS/ERK and PI3-K signaling pathways are important for activation of CREB and NF-kappa B in GDNF-treated cells, respectively.
  • K Kawai, T Iwashita, H Murakami, N Hiraiwa, A Yoshiki, M Kusakabe, K Ono, K Iida, A Nakayama, M Takahashi
    CANCER RESEARCH 60(18) 5254-5260 2000年9月  査読有り
    Germ line mutations of the RET proto-oncogene are responsible for the development of multiple endocrine neoplasia type 2A (MEN 2A), an inherited cancer syndrome characterized by medullary thyroid carcinoma, pheochromocytoma, and parathyroid hyperplasia, To study the mechanism of tissue-specific tumor development by RET with a MEN2A (cysteine 634-->arginine) mutation, we generated transgenic mice by introducing the RET-MEN2A gene fused to Moloney murine leukemia virus long terminal repeat. Expression of the transgene and its product was detected at variable levels in a variety of tissues including thyroid, heart, liver, colon, parotid gland, and brain. All of 29 mice analyzed developed thyroid C-cell hyperplasia or medullary carcinoma, accompanying high levels of serum calcitonin, In addition, development of mammary or parotid gland adenocarcinoma was observed in one-half of the transgenic mice. RET dimerization and its complex formation with Shc and Grb2 adaptor proteins were detected in tumor tissues, Unexpectedly, no tumor formation was found in other tissues despite RET-MEN2A expression where RET dimerization was undetectable. Because these tissues but not tumors expressed glial cell line-derived neurotrophic factor family receptor alpha (GFR alpha) at high levels, this suggested that GFR alpha expression may interfere in the dimerization of the RET-MEN2A mutant proteins, leading to tissue-specific tumor development in vivo.
  • 川井 久美, 岩下 寿秀, 浅井 直也, 村上 秀樹, 平岩 典子, 日下部 守昭, 高橋 雅英
    日本病理学会会誌 89(1) 294-294 2000年3月  
  • Kato M, Isobe K, Dai Y, Liu W, Takahashi M, Nakashima I
    Invest Dermatol 114(3) 599-601 2000年3月  
  • Kato M, Iwashita T, Takeda K, Akhand AA, Liu W, Yoshihara M, Asai N, Suzuki H, Takahashi M, Nakashima I
    Mol Biol Cell 11(1) 93-101-101 2000年1月  
  • Sato, M, Sekido, Y, Horio, Y, Takahashi, M, Saito, H, Minna, J.D, Shimokata, K, Hasegawa, Y
    Jpn J Cancer Res 91(5) 504-9-509 2000年  査読有り
  • T Iwashita, M Kato, H Murakami, N Asai, Y Ishiguro, S Ito, Y Iwata, K Kawai, M Asai, K Kurokawa, H Kajita, M Takahashi
    ONCOGENE 18(26) 3919-3922 1999年7月  査読有り
    Several mutations were identified in the kinase domain of the RET proto-oncogene in patients with multiple endocrine neoplasia (MEN) 2B, familial medullary thyroid carcinoma (FMTC) or sporadic medullary thyroid carcinoma. We introduced seven mutations (glutamic acid 768-->aspartic acid (E768D), valine 804-->leucine (V804L), alanine 883-->phenylalanine (A1883F), serine 891-->alanine (S891A), methionine 918 -->threonine (M918T), alanine 919-->proline (A919P) and E768D/A919P) into the short and long isoforms of RET cDNA and transfected the mutant cDNAs into NIH3T3 cells, The transforming activity of the long isoform of Ret with each mutation was much higher that that of its short isoform, Based on the levels of the transforming activity, these mutant RET genes were classified into two groups; a group with high transforming activity (A883F, M918T and E768D/A919P) and a group with low transforming activity (E768D, V804L, S891A and A919P) (designated high group and low group). Interestingly, the level of transforming activity correlated with clinical phenotypes; high group Bet with the A883F or M918T mutation and low group Ret with the E768D, V804L or S891A mutation were associated with the development of MEN 2B and FMTC, respectively. in addition, we found that substitution of phenylalanine for tyrosine 905 present in the kinase domain abolished both transforming and autophosphorylation activities of low group Ret whereas it did not affect the activities of high group Ret.
  • 高橋 雅英, 浅井 直也, 岩下 寿秀, 村上 秀樹, 伊藤 信二
    日本内分泌学会雑誌 75(1) 42-42 1999年4月  
  • H Murakami, T Iwashita, N Asai, Y Iwata, S Narumiya, M Takahashi
    ONCOGENE 18(11) 1975-1982 1999年3月  査読有り
    Glial cell line-derived neurotrophic factor (GDNF) signals through a unique receptor system that includes Ret receptor tyrosine kinase and a glycosyl-phosphatidylinositol-linked cell surface protein. In the present study, we have identified several proteins in neuroblastoma cells that are phosphorylated on tyrosine in response to GDNF, The phosphorylated proteins include focal adhesion kinase (FAK), paxillin and Crk-associated substrate, p130(Cas), all of which are known to be associated with focal adhesions. Of these, paxillin and p130(Cas) interacted with Crk proteins in GDNF-treated neuroblastoma cells. GDNF also induced reorganization of the actin cytoskelton, Tyrosine phosphorylation of FAK, paxillin and p130(Cas) was inhibited by cytochalasin D or two specific inhibitors of phosphatidylinositol-3' kinase (PI-3' kinase), wortmannin and LY294002, indicating that their tyrosine phosphorylation depends on the formation of actin stress fiber and activation of PI-3' kinase. In addition, phosphorylation of FAK but not of paxillin and p130(Cas) was markedly impaired by the Clostridium botulinum C3 exoenzyme that specifically ADP-ribosylates and inactivates Rho, These results suggested the presence of Rho-dependent and -independent signaling pathways downstream of PI-3' kinase that mediate tyrosine phosphorylation of FAK, paxillin and p130(Cas) through Pet kinase.
  • Kato M, Liu W, Akhand AA, Dai Y, Ohbayashi M, Tuzuki T, Suzuki H, Isobe K-I, Takahashi M, Izumi Nakashima
    Oncogene 18(3) 837-842-842 1999年1月21日  
  • Asai M, Kato M, Asai N, Iwashita T, Murakami H, Kawai K, Nakashima I, Takahashi M
    Jpn J Cancer Res 90(1) 86-92 1999年1月  
  • M Takahashi, T Iwashita, M Santoro, S Lyonnet, GM Lenoir, M Billaud
    HUMAN MUTATION 13(4) 331-336 1999年  査読有り
    Multiple endocrine neoplasia type 2 (MEN2) and Hirschsprung's disease (HSCR) are two dominantly inherited neurocristopathies ascribed to mutations in the RET gene [Chakravarti, 1996; Pasini et al., 1996; Eng and Mulligan, 1997]. MEN2 is a cancer syndrome comprising three related clinical subtypes: (1) MEN type 2A (MEN2A; MIM# 171400) characterized by the association of medullary thyroid carcinoma (MTC), pheochromocytoma (Pheo), and hyperparathyroidism; (2) MEN type 2B (MEN2B; MIM# 162300), which includes MTC, Pheo, mucosal neuromas, ganglioneuromatosis of the digestive tract, and skeletal abnormalities; and (3) familial MTC (FMTC; MIM# 155240), defined by the sole occurrence of MTC. HSCR (MIM# 142623) is a congenital malformation caused by the absence of enteric plexuses in the hindgut, leading to bowel obstruction in neonates. The RET gene (MIM# 164761) codes for a transmembrane tyrosine kinase, a component of a multimeric complex that also comprises one of four members of a novel family of glycosylphosphatidylinositol (GPI)-anchored receptor, GFR alpha(1-4) (e.g., GFRA1, MIM# 601496; references are detailed in Baloh et al. [1998]. Four structurally related soluble factors-glial cell line-derived neurotrophic factor (GDNF), neurturin, persephin, and artemin-are the ligands of these multimolecular receptors in which the nature of the GFR alpha determines the ligand specificity of the complex [see Baloh et al., 1998, for references]. It is well documented that RET/GFR alpha-1/GDNF delivers a signal critical for the survival of the early neural crest derived precursors that colonize the intestine below the rostral foregut and give rise to the enteric nervous plexuses [Gershon, 1997; Cacalano et al., 1998; Enomoto et al., 1998]. Hum Mutat 13:331-336, 1999. (C) 1999 Wiley Liss, Inc.
  • Kiyoshi Yanagisawa, Hirotaka Osada, Akira Masuda, Masashi Kondo, Toshiko Saito, Yasushi Yatabe, Kenzo Takagi, Toshitada Takahashi, Takashi Takahashi
    Oncogene 17(13) 1743-1747 1998年10月  
    Smad family members are essential intracellular signaling components of the transforming growth factor-beta (TGF-β) superfamily involved in a range of biological activities. Two highly homologous molecules, Smad2 and Smad3, have so far been identified as receptor-activated Smads for TGF-β signaling and have become the focus of intensive studies. However, no definite differences in regulation or function have been established between these TGF-β signaling molecules. In the present study, we show that the expression of Smad3, but not its dose relative, Smad2, is down-regulated by TGF-β mediated signals themselves in human lung epithelial cells. This down-regulation of Smad3 by TGF-β treatment did not appear to result from shortening of the half-life of Smad3 mRNA. Constitutive expression of Smad3 in the presence of TGF-β induced apoptotic cell death, with an adverse effect on the cell growth of human lung epithelial cells. Apoptotic cell death could also be induced by forced expression of Smad2 in the presence of TGF-β, but less efficiently than by that of Smad3. These findings clearly define the distinctions between Smad2 and Smad3 for the first time in that a qualitative difference was observed with regard to the regulation of their expression in response to TGF-β, while Smad2 and Smad3 appeared to have quantitatively different capabilities regarding the induction of apoptotic cell death in human lung epithelial cells.
  • M Kato, M Takahashi, AA Akhand, W Liu, Y Dai, S Shimizu, T Iwamoto, H Suzuki, Nakashima, I
    ONCOGENE 17(14) 1885-1888 1998年10月  
    We report here on a novel metallothionein-I (MT)/ret transgenic mouse line in which skin melanosis, benign melanocytic tumor and malignant melanoma metastasizing to distant organs develop stepwise. The process of tumor development and its malignant transformation in this line may resemble that of the human giant congenital melanocytic nevus that is present at birth and that frequently gives rise to malignant melanoma during aging. We observed an increase in the expression level and activity of the ret transgene during the disease progression. That increase in transgene expression accompanied an activation of mitogen-activated protein kinases (MAPKs) and c-Jun as well as matrix metalloproteinases. These results suggest that progressive dysregulation of the expression level of the ret transgene might play a crucial role in the malignant transformation of melanocytic tumors developed in the MT/ret transgenic mouse line.
  • 高橋 雅英, 浅井 直也, 岩下 寿秀, 村上 秀樹, 伊藤 信二
    生化学 70(8) 690-690 1998年8月  
  • 高橋 雅英, 野崎 千佳, 浅井 直也, 岩下 寿秀, 村上 秀樹
    末梢神経 = Peripheral nerve 8(1) 25-29 1998年6月10日  
  • 伊藤 信二, 岩下 寿秀, 浅井 直也, 村上 秀樹, 高橋 雅英
    G.I.Research 6(3) 225-230 1998年6月  
  • RD Klein, D Sherman, WH Ho, D Stone, GL Bennett, B Moffat, R Vandlen, L Simmons, QM Gu, JA Hongo, B Devaux, K Poulsen, M Armanini, C Nozaki, N Asai, A Goddard, H Phillips, CE Henderson, M Takahashi, A Rosenthal
    NATURE 392(6672) 210-210 1998年3月  査読有り
  • M. Takahashi, N. Asai, T. Iwashita, H. Murakami, S. Ito
    Recent results in cancer research. Fortschritte der Krebsforschung. Progrès dans les recherches sur le cancer 154 229-236 1998年  
    The ret proto-oncogene encodes a receptor tyrosine kinase whose ligands belong to the glial cell line-derived neurotrophic factor (GDNF) protein family. Its germline mutations are responsible for the development of multiple endocrine neoplasia (MEN) types 2A and 2B and Hirschsprung's disease (HSCR). MEN2A and MEN2B mutations result in the constitutive activation of Ret by different molecular mechanisms. MEN2A mutations involve cysteine residues present in the Ret extracellular domain and induce disulfide-linked Ret dimerization on the cell surface. MEN2B mutations were identified in methionine 918 in the tyrosine kinase domain and activate Ret without dimerization, probably due to a conformational change of its catalytic core region. In contrast to MEN2 mutations, HSCR mutations represent loss of function mutations. We found that most of HSCR mutations detected in the extracellular domain impair the Ret cell surface expression. More interestingly, ret mutations in cysteines 618 and 620 were reported in several families who developed both MEN2A and HSCR. It was suggested that these mutations might have two biological effects on Ret function, leading to the development of different clinical phenotypes in the same patients.
  • C Nozaki, N Asai, H Murakami, T Iwashita, Y Iwata, K Horibe, RD Klein, A Rosenthal, M Takahashi
    ONCOGENE 16(3) 293-299 1998年1月  査読有り
    Glial cell line-derived neurotrophic factor (GDNF) and neurturin (NTN) define a new family of neurotrophic factors that play crucial roles in survival and differentiation of various neurons. Recent studies demonstrated that GDNF and NTN use a multicomponent receptor system in which glycosyl-phosphatidylinositol (GPI)-linked cell surface proteins and Ret receptor tyrosine kinase function as the ligand-binding and signalling components, respectively. In the present study, we investigated the role of Ca2+ ions for biochemical and biological activities of Ret because Ret has a unique structure of the extracellular domain with the cadherin-like motif. The results demonstrated that Ca2+ ions might be required for the complex formation of Ret and GDNF or NTN that induces Ret oligomerization and autophosphorylation. Full morphological differentiation of neuroblastoma cells by these neurotrophic factors was also Ca2+-dependent. These findings thus suggested that, in addition to GPI-linked cell surface proteins, Ca2+ ions are components of the signal transducing complex formed by Ret and GDNF protein family.
  • Akira Masuda, Masashi Kondo, Toshiko Saito, Yasushi Yatabe, Tomoko Kobayashi, Mitsumasa Okamoto, Motokazu Suyama, Toshitada Takahashi, Takashi Takahashi
    Cancer Research 57(21) 4898-4904 1997年11月  
    Novel human epithelial cell lines retaining characteristic features of normal peripheral airway cells were established by transfecting the SV40 large T antigen gene into primary in vitro outgrowths from normal peripheral lung specimens. These lines, designated as HPL1A to HPL1E, showed the polygonal shapes typical of epithelial cells and expressed cytokeratin in abundance. Ultrastructural examination revealed the presence of microvilli, multivesicular bodies, and multilamellar body-like structures that are characteristic of type II pneumocytes, but expression of CC10 transcripts, a highly specific marker for Clara cells, was also observed. Response to transforming growth factor β, epidermal growth factor (EGF), and hepatocyte growth factor, all of which are thought to be important growth-regulatory molecules for cellular proliferation and developmental processes of peripheral lung, was apparent. In the HPL1A case, markedly altered cell morphology and cytoskeletal organization, potent inhibition of cell growth, and increased expression of an extracellular matrix protein were noted with transforming growth factor β. Interestingly, both EGF and hepatocyte growth factor stimulated anchorage-dependent growth, whereas only EGF could sustain anchorage-independent proliferation. The HPL1 lines are, to our knowledge, the first series of stable epithelial lines of human peripheral lung to be described. They should be valuable for investigating various aspects of growth regulation and oncogenic processes, including the mechanisms of acquisition of anchorage independence and the interrelationships of genetic changes identified previously in lung cancers. In addition, the HPL1 lines may also prove useful for development of in vitro models for other human lung disorders as well as to elucidate the mechanisms of peripheral lung differentiation.
  • Sharma Nidhi, Ohyabu Atsushi, Murakumo Yoshiki, TAKAHASHI Masahide, SAITO Mitsuhiro, AMO Hiroyuki, MURAYAMA Shin-ichiro, OHNO Kyoko, ODA Sen-ichi, MATSUYAMA Mutsushi
    Pathology international 47(7) 436-441 1997年7月  
  • S Ito, T Iwashita, N Asai, H Murakami, Y Iwata, G Sobue, M Takahashi
    CANCER RESEARCH 57(14) 2870-2872 1997年7月  査読有り
    We investigated the transforming activity of the ret proto-oncogene with a mutation in cysteine 609, 611, 618, 620, 630, or 634 detected in patients with multiple endocrine neoplasia type 2A (MEN 2A), familial medullary thyroid carcinoma (FMTC), or Hirschsprung's disease. Of these cysteine mutations, codon 634 mutations are known to be correlated with the development of MEN 2A, whereas codon 609, 618, or 620 mutations were detected in two-thirds of FMTCs and in several cases of Hirschsprung's disease. Analysis of a total of 18 mutant genes revealed that codon 634 mutations have the highest transforming activity, The activity of ret with a codon 609, 611, 618, or 620 mutation and with a codon 630 mutation was approximately 3- to 5-fold and 2-fold lower than that of ret with a codon 634 mutation, respectively, In addition, different amino acid substitutions for the same cysteine displayed comparable transforming activity. The expression of the cell surface form of Ret with codon 609, 611, 618, or 620 mutation was very low compared with that of Ret with codon 634 mutation, indicating that the former four mutations might impair transport of Ret to the plasma membrane, as observed for several Hirschsprung mutations affecting the Ret extracellular domain. These results thus suggest that mutations in cysteine 609, 611, 618, or 620 may have the potential to develop Hirschsprung's disease in addition to MEN 2A and FMTC.
  • 高橋 雅英, 浅井 直也, 岩下 寿秀
    モレキュラ-メディシン 34(6) 750-755 1997年6月  
  • OHIWA M, MURAKAMI H, IWASHITA T, ASAI N, IWATA Y, IMAI T, FUNAHASHI H, TAKAGI H, TAKAHASHI M
    Biochemical and Biophysical Research Communications 237(3) 747-751 1997年  
  • T Iwashita, H Murakami, N Asai, M Takahashi
    HUMAN MOLECULAR GENETICS 5(10) 1577-1580 1996年10月  査読有り
    Hirschsprung disease (HSCR) is a congenital disorder associated with the absence of intrinsic ganglion cells in the distal gastrointestinal tract, Recently, many missense, nonsense and frameshift mutations of the ref proto-oncogene were found in familial and sporadic cases of HSCR, Consistent with the view that the HSCR phenotype is the result of inactivation of Ret, the missense mutations detected in the tyrosine kinase domain were demonstrated to result in a marked decrease of the kinase activity of Ret, However, the effects of missense mutations found in the extracellular domain remain unknown, We now report that five mutations in the extracellular domain examined inhibit transport of the Ret protein to the plasma membrane, As a consequence, they significantly decreased the transforming activity of Ret with multiple endocrine neoplasia (MEN) 2A mutation for which cell surface expression is required, Our results also demonstrated that long segment HSCR mutations more severely impair transport of Ret to the plasma membrane than a short segment HSCR mutation, suggesting that the level of its cell surface expression may correlate to the HSCR phenotype.
  • N Asai, H Murakami, T Iwashita, M Takahashi
    JOURNAL OF BIOLOGICAL CHEMISTRY 271(30) 17644-17649 1996年7月  査読有り
    Germ line mutations of the ret proto-oncogene are associated with the development of three dominantly inherited neoplastic disorders, multiple endocrine neoplasia (MEN) 2A, MEN 2B, and familial medullary thyroid carcinoma, It has been demonstrated that the mutations result in constitutive activation of the Ret protein, leading to transformation of MH 3T3 cells, In the present study we investigated the role of tyrosine residues present in the carboxyl-terminal sequence for the transforming activity of Ret with the MEN 2A or MEN 2B mutation (MENSA-Ret or MEN2B-Ret). Substitution of phenylalanine for tyrosine 1062 (designated Y1062F) markedly impaired the transforming activity of both MEN2A-Ret and MEN2B-Ret, whereas substitution or deletion for four other tyrosines (codons 981, 1015, 1090, and 1096) did not affect their activity, The She adaptor proteins bound to the MEN2A-Ret and MEN2B-Ret proteins and were phosphorylated on tyrosine in the transfectants. The binding of She to the Y1062F mutant proteins was reduced by approximately 80%, indicating that tyrosine 1062 is a major binding site for She. In addition, phosphopeptide analysis of MENSA-Ret demonstrated that tyrosine 1062 represents an autophosphorylation site of the mutant Ret proteins.
  • JJS Treanor, L Goodman, F deSauvage, DM Stone, KT Poulsen, CD Beck, C Gray, MP Armanini, RA Pollock, F Hefti, HS Phillips, A Goddard, MW Moore, A BujBello, AM Davies, N Asai, M Takahashi, R Vandlen, CE Henderson, A Rosenthal
    NATURE 382(6586) 80-83 1996年7月  査読有り
    GLIAL-CELL-LINE-DERIVED neurotrophic factor (GDNF)(1) is a potent survival factor for central and peripheral neurons(2-6), and is essential for the development of kidneys and the enteric nervous system(7-9). Despite the potential clinical and physiological importance of GDNF, its mechanism of action is unknown. Here we show that physiological responses to GDNF require the presence of a novel glycosyl-phosphatidylinositol (GPI)-linked protein (designated GDNFR-alpha) that is expressed on GDNF-responsive cells and binds GDNF with a high affinity. We further demonstrate that GDNF promotes the formation of a physical complex between GDNFR-alpha and the orphan tyrosine kinase receptor Ret(10-12), thereby inducing its tyrosine phosphorylation. These findings support the hypothesis that GDNF uses a multi-subunit receptor system in which GDNFR-alpha and Ret function as the ligand-binding and signalling components, respectively.
  • T Iwashita, N Asai, H Murakami, M Matsuyama, M Takahashi
    ONCOGENE 12(3) 481-487 1996年2月  査読有り
    The c-ret proto-oncogene with multiple endocrine neoplasia (MEN) 2A or 2B mutation can transform NIH3T3 cells with high efficiencies as a consequence of its constitutive activation. The MEN2A mutation induces ligand-independent homodimerization of the Ret protein on the cell surface while the MEN2B mutation appears to alter the catalytic activity without dimerization. In the present study, we investigated the role of tyrosine residues present in the kinase domain for the transforming activity of the mutant Ret proteins. Substitution of phenylalanine for tyrosine 905 (Y905F) that corresponds to tyrosine 416 of the Src protein abolished the transforming activity of Ret with the MEN2A mutation (MEN2A-Ret) but not with the MEN2B mutation (MEN2B-Ret). On the other hand, the transforming activity of MEN2B-Ret but not MEN2A-Ret significantly decreased by changing tyrosine 864 or 952 to phenylalanine. In addition, double mutations of these tyrosines (Y864/952F) completely abolished the activity of MEN2B-Ret. The Y905F and Y864/952F mutations resulted in severe impairment of the kinase activity of MEN2A-Ret and MEN2B-Ret, respectively. These results thus indicated that tyrosine residues essential for the transforming activity are different between MEN2A-Ret and MEN2B-Ret.
  • WADA M, ASAI N, TSUZUKI T, MARUYAMA S, OHIWA M, IMAI T, FUNAHASHI H, TAKAGI H, TAKAHASHI M
    Biochemical and Biophysical Research Communications 218(2) 606-609 1996年  
  • MURAKUMO Yoshiki, TAKAHASHI Masahide, HAYASHI Naomi, TAGUCHI Masako, ARAKAWA Atsushi, SHARMA Nidhi, SAKATA Keita, SAITO Mitsuhiro, AMO Hiroyuki, KATOH Hideki, MATSUYAMA Mutsushi
    Pathology international 45(4) 261-265 1995年4月  
  • N ASAI, T IWASHITA, M MATSUYAMA, M TAKAHASHI
    MOLECULAR AND CELLULAR BIOLOGY 15(3) 1613-1619 1995年3月  査読有り
    Transforming activity of the c-ret proto-oncogene,vith multiple endocrine neoplasia (MEN) 2A mutations was investigated by transfection of NIH 3T3 cells. Mutant c-ret genes driven by the simian virus 40 or cytomegalovirus promoter induced transformation with high efficiencies, The 170-kDa Ret protein present on the cell surface of transformed cells was highly phosphorylated on tyrosine and formed disulfide-linked homodimers. This result indicated that MEN 2A mutations induced ligand-independent dimerization of the c-Ret protein on the cell surface, leading to activation of its intrinsic tyrosine kinase. In addition to the MEN 2A mutations, we further introduced a mutation (lysine for asparaginic acid at codon 300 [D300K]) in a putative Ca2+-binding site of the cadherin-like domain. When c-ret cDNA with both MEN 2A and D300K mutations was transfected into NIH 3T3 cells, transforming activity drastically decreased. Western blot (immunoblot) analysis revealed that very little of the 170-kDa Ret protein with the D300K mutation was expressed in transfectants while expression of the 150-kDa Ret protein retained in the endoplasmic reticulum was not affected. This result also demonstrated that transport of the Ret protein to the plasma membrane is required for its transforming activity.
  • T TSUZUKI, M TAKAHASHI, N ASAI, T IWASHITA, M MATSUYAMA, JP ASAI
    ONCOGENE 10(1) 191-198 1995年1月  査読有り
    Immunohistochemical analysis with the anti-Ret antibody was performed to investigate the expression of the c-ret proto-oncogene product (c-Ret protein) in embryonic, infant and adult rat tissues. During embryogenesis, the c-Ret expression became detectable by day 11.5 in the developing peripheral and central nervous systems as well as in the excretory system. In the peripheral nervous system of the trunk, it was expressed at high levels in the enteric neuroblasts and the autonomic and dorsal root ganglia, c-Ret positive cells appeared in the mesenchyme around the foregut and the dorsal aorta at day 11.5 and formed the myenteric plexus of the whole embryonic gut and the sympathetic trunk at later stages respectively. Examination of the cranial region revealed that the c-Ret protein was expressed in neural crest cells migrating from rhombomere 4 at day 11.5 and then became positive in the facial, glossopharyngeal and vagus cranial ganglia at day 12.5-13.5. After day 16.5 of gestation, the c-Ret expression was also observed in the trigeminal ganglion. In the central nervous system, the c-Ret protein was expressed in the neuroepithelial cells of the ventral neural tube (day 11.5-14.5), the motor neurons of the spinal cord (day 18.5) as well as in the embryonic neuroretina (day 18.5). In addition to the nervous system, the c-Ret expression was detected in the nephric duct (day 11.5), the ureteric bud (day 13.5) and the collecting ducts of the kidney (day 16.5). After birth, neurons in the nervous systems mentioned above continued to express the c-Ret protein at variable levels while no c-Ret expression was observed in the kidney of adult rats. Furthermore, the c-Ret expression was found in the acinar cells of the salivary gland, the epithelial cells of the thymus and the follicular dendritic cells of the spleen and lymph node in infant and adult rats. A small number of adrenal chromaffin cells and thyroid C cells also expressed the c-Ret protein. These results demonstrated that the c-Ret protein is expressed in several cell lineages in spatially and temporally controlled manner throughout life of the rat.
  • M KATO, T HATTORI, M TAKAHASHI, N YANAGITA, NAKASHIMA, I
    BRITISH JOURNAL OF CLINICAL PRACTICE 48(6) 299-301 1994年11月  
    In this study, prophylactic treatment with ketotifen and examination of subjective symptoms, blood eosinophil count and serum eosinophil cationic protein (ECP) values were carried out from the pre-to postpollinosis season to evaluate the effects of ECP and characterise the effects of prophylactic treatment on the appearance of these mediators in relation to clinical symptoms. We found that: serum ECP values during the season and post-season were significantly higher than those during the pre-season; there was a significant correlation between serum ECP value and blood eosinophil count during the season; and the levels for all three of the above parameters in the prophylactic treatment group were significantly lower than those in the no-treatment group during the season.
  • Maruyama S, Iwashita T, Imai T, Funahashi H, Ceccherini I, Luo Y, Romeo G, Matsuo S, Matsuyama M, Takahashi M
    Jpn J Cancer Res 85(9) 879-882-882 1994年9月  査読有り
    We investigated mutations of the ret proto-oncogene in Japanese patients with multiple endocrine neoplasia (MEN) type 2A and type 2B. DNAs from pheochromocytomas and/or medullary thyroid carcinomas (MTCs) of five MEN 2A and three MEN 2B patients were amplified by a polymerase chain reaction (PCR) and analyzed. Tumors of four MEN 2A patients had missense mutations in Cys 634 in the extracellular domain of the ret proto-oncogene. The same mutations were detected in normal tissues of the patients, indicating that the mutations had arisen in the germ line. Using a reverse transcriptase(RT)-PCR, both normal and mutant transcripts of the ret proto-oncogene were detected in a tumor of one patient with MEN 2A mutation. In addition, three MEN 2B patients examined had the same point mutation (ATG-->ACG) at codon 918 in the tyrosine kinase domain of the ret proto-oncogene. Since all mutations identified in this study generated new restriction enzyme sites or eliminated a restriction site, the mutant alleles of affected family members could be readily detected without sequencing.
  • Takahashi M, Asai N, Iwashita T, Isomura T, Miyazaki K, Matsuyama M
    Oncogene. 8(11) 2925-9-29259 1993年11月  査読有り
  • Y MURAKUMO, M TAKAHASHI, A ARAKAWA, M SAITO, H AMO, H KATOH, M MATSUYAMA
    JAPANESE JOURNAL OF CANCER RESEARCH 84(8) 838-840 1993年8月  
    The thymoma-prone rat of the BUF/Mna strain is a useful model for human thymoma. In this strain thymoma development is regulated by a single autosomal susceptible gene, Tsr-1. At pre-thymoma age, BUF/Mna rats have extremely large thymuses, when compared to those of other strains of rats. Genetic studies in crosses between BUF/Mna rats with large thymuses and WKY/NCrj rats with small thymuses suggested the presence of a major autosomal gene, Ten-1, which contributes to thymus enlargement in a backcross population. Linkage studies between Ten-1 and microsatellite markers in backcross rats of (WKY/NCrj x BUF/Mna)F1 x BUF/Mna have led to the localization of Ten-1 in chromosome 1. This result may provide an approach to clone Tsr-1, which could be allelic to Ten-1.
  • Y SEKIDO, T TAKAHASHI, R UEDA, M TAKAHASHI, H SUZUKI, K NISHIDA, T TSUKAMOTO, T HIDA, K SHIMOKATA, KM ZSEBO, T TAKAHASHI
    CANCER RESEARCH 53(7) 1709-1714 1993年4月  
    Accumulating evidence suggests that c-kit and its ligand, stem cell factor (SCF), play an important role in the regulation of at least three lineages of stem cell growth and possibly in leukemogenesis, while only limited data are available that suggest possible involvement of c-kit/SCF in the development of human solid tumors such as lung cancer. We have recently reported that c-kit is aberrantly expressed almost exclusively in small-cell lung cancer (SCLC) among various types of solid tumors. The present study revealed that c-kit protein ectopically expressed in SCLC is indistinguishable from that in leukemia cell lines with megakaryocy tic characteristics with respect to amount, molecular size, and autophosphorylation status in response to recombinant human SCF. Furthermore, significant chemotactic response as well as moderate in vitro cell growth was induced in SCLC cell lines by the addition of recombinant human SCF, suggesting that c-kit/SCF may play an important biological role in the development of SCLC. Our extensive search for activating mutations naturally occurring in the c-kit gene revealed an amino acid substitution in the transmembrane domain of an SCLC cell line, although the functional consequences of this variant allele are yet to be determined.
  • T IWAMOTO, M TAKAHASHI, M ITO, K HAMATANI, M OHBAYASHI, W WAJJWALKU, K ISOBE, NAKASHIMA, I
    EMBO JOURNAL 10(11) 3167-3175 1991年11月  査読有り
    We generated four independent transgenic mouse lines that showed severe melanosis of the whole body by introducing the ret oncogene fused to the mouse metallothionein (MT)-I promoter-enhancer (MT/ret). Whereas melanogenesis was accelerated without distinct proliferative disorders in one line, melanocytic tumours frequently developed in the other three lines. Northern hybridization and in situ hybridization analyses showed that tumour cells and non-tumorous melanin-producing cells expressed the transgene at high levels. The aberrant melanogenesis and tumour development were influenced by genetic and environmental factors. Furthermore, crossbreeding experiments between the transgenic mice and W(nu) mice suggested that the ret gene product can partially compensate for the defect of melanocyte development in W(nu) mice. This is a novel mammalian model in which melanosis and melanocytic tumours develop stepwise, triggered by a single transgene.
  • M TAKAHASHI, Y BUMA, M TANIGUCHI
    ONCOGENE 6(2) 297-301 1991年2月  査読有り
    Monoclonal and/or polyclonal antibodies were generated against the products synthesized from two portions of the ret proto-oncogene (c-ret) cDNA expressed in Escherichia coli. These antibodies were reactive in immunoblotting with 150 kd and 170 kd proteins in cell lysates from three human neuroblastoma cell lines expressing the ret proto-oncogene. When the neuroblastoma cells were treated with tunicamycin, a protein with an apparent molecular weight of 120 kd, which is consistent with that of the c-ret protein predicted from the cDNA sequence, appeared on immunoblots. These results indicated that the 150 kd and 170 kd proteins in neuroblastoma cells are produced from a single polypeptide of 120 kd by post-translational glycosylation. Furthermore, the antibodies detected a unique 190 kd protein as well as 150 kd protein in a cell lysate from THP-1 human monocytic leukemia cell line, suggesting that glycosylated forms of the c-ret protein are different between neuroblastoma and leukemia cells.
  • Sekido Y, Obata Y, Ueda R, Hida T, Suyama M, Shimokata K, Ariyoshi Y, Takahashi T
    Cancer Res. 51 2416-2419 1991年  
  • M TAKAHASHI, Y BUMA, H HIAI
    ONCOGENE 4(6) 805-806 1989年6月  
  • M TAKAHASHI, Y BUMA, T IWAMOTO, Y INAGUMA, H IKEDA, H HIAI
    ONCOGENE 3(5) 571-578 1988年11月  査読有り
  • M TAKAHASHI, Y INAGUMA, H HIAI, F HIROSE
    MOLECULAR AND CELLULAR BIOLOGY 8(4) 1853-1856 1988年4月  査読有り
  • M TAKAHASHI, GM COOPER
    MOLECULAR AND CELLULAR BIOLOGY 7(4) 1378-1385 1987年4月  査読有り
  • M TAKAHASHI, J RITZ, GM COOPER
    CELL 42(2) 581-588 1985年  査読有り

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