1997 年度実績報告書

マウスAGM領域からの造血幹細胞自己複製因子のクローニング

研究課題

研究課題/領域番号	09357007
研究種目	基盤研究(A)
研究機関	東京大学
研究代表者	中畑龍俊東京大学, 医科学研究所, 教授 (20110744)
研究分担者	渡辺すみ子東京大学, 医科学研究所, 助手 (60240735) 前川平東京大学, 医科学研究所, 講師 (80229286) 辻浩一郎東京大学, 医科学研究所, 助教授 (50179991)
キーワード	AGM領域 / 造血幹細胞 / 自己複製因子 / マウス / ヒト / ストローマ細胞株 / AGM-S3 / 遺伝子クローニング
研究概要	造血幹細胞の自己複製因子を同定するため造血幹細胞の発生の場である胎生9〜11.5日目のマウスAGM領域を器官培養することにより多数のストローマ細胞を得た。その中でヒト臍帯血CD34細胞との共培養で未分化造血前駆細胞の増殖を支持するストローマ細胞が胎生10.5,11.5日目のAGM領域の培養より得られた。このストローマ細胞から限界希釈法を用いてストローマ細胞株の樹立を試み、3種類(AGMS1,AGMS2,AGMS3)のストローマ細胞株を得た。マウス骨髄より分離したlin^-Sca-1^+c-Kit^<high>細胞をこれらに重層後10日間培養し、day12CFU-Sを測定したところAGMS3上で著明な増幅が得られた。AGM-S3はVCAM-1,CD13,Sca-1陽性であり血管内皮細胞の性格を有していた。AGM-S3とヒト臍帯血CD34^+CD38^-細胞を共培養すると、6週間以上の長期にわたり、ヒトCFU-Mixの増幅が得られた。また、AGMS3上で培養前後の臍帯血単核球1x10^6をNOD/SCIDマウスに移植し8週後のマウス骨髄を解析したところ、AGMS3と共培養後移植したマウス骨髄中にヒト細胞が検出された。以上より、AGM-S3は種を越えて働く造血幹細胞自己複製因子を発現している可能性が示唆された。AGM-S3が発現している既知のサイトカインmRNAを検討したところ、IL-6,OSM,SCFの発現のみが認められが、これらではヒト造血幹細胞の増殖は起こらないことから、AGM-S3は造血幹細胞の増殖に働く未知の分子を発現している可能性が高いことが明らかとなった。種々の検討により。予想される分子は膜結合型であることから、現在、AGMS3からcDNAライブラリーを作製し、動物細胞発現ベクターに組み込み、ヒトCD34^+CD38^-細胞の増殖を指標に発現クローニングを試みている。

研究成果
(28件)

すべてその他

すべて文献書誌 (28件)

[文献書誌] Muraoka K.: "Defective response to thrombopoietin and impared expression of c-mpl mRAD in bone marrow cells in congenital amegakaryocytic thrombocytopenia." Brit.J.Haematol.96. 287-292 (1997)
[文献書誌] Eguchi M.: "Morphological and cytochemical changes in human mast cells during culture." Med.Electron Microsc.30. 25-30 (1997)
[文献書誌] Toru H.: "Interleukin-4 induces homotypic aggregation of human mast cells by promoting LFA-1/ICAM-1 adhesion molecules." Blood. 89. 3296-3302 (1997)
[文献書誌] Yanagida M.: "Effect of a chymotrypsin-like inhibitor,TPCK,on histamine release from cultured human mast cells." J.Pharm.Pharmth.49. 537-541 (1997)
[文献書誌] Ishiguro A.: "Effects and serum levels of thrombopoietin in a case of chronic thrombocytopenia with achondroplasia." Int.J.Hematol.66. 99-102 (1997)
[文献書誌] Muraoka K.: "Thrombopoietin-independent effect of interferon-γ on the proliferation of human megakaryocyte progenitors." Brit.J.Haematol.98. 265-273 (1997)
[文献書誌] Nishijima I.: "Hemopoietic and lymphopoietic responses in human GM-CSF receptor transgenic mice injected with human GM-CSF." Blood. 90. 1031-1038 (1997)
[文献書誌] Yoshida M.: "Thrombopoietin alone stimulates the early proliferation and survival of human erythroid,myeloid,and multipotential progenitors in serum-free culture." Brit.J.Haematol.98. 254-264 (1997)
[文献書誌] Umemoto Y.: "Leptin stimulates the proliferation of murine myelocytic and primitive hematopoietic progenitor cells." Blood. 90. 3438-3443 (1997)
[文献書誌] Akasaka T.: "The role of mel-18,a Mammalian Polycomb group gene,during the IL-7-dependent proliferation of lymphocyte precursors." Immunity. 7. 135-146 (1997)
[文献書誌] Ebihara Y.: "Synergistic action of Flt3 and gp 130 signalings in human hematopoiesis." Blood. 90. 4363-4368 (1997)
[文献書誌] Tsuji K.: "Role of soluble interleukin-6 receptor with interleukin-6 in human hematopoiesis." Hematology. 2. 143-149 (1997)
[文献書誌] Suzuki H.: "An increase in peripheral blood progenitor cells in primates by co-administration of recombinant human interleukin 6 and recombinant human granulocyte colony-stimulating factor." Transplantation. 64. 1468-1473 (1997)
[文献書誌] Bessho M.: "Treatment of the anemia of aplastic anemia patients with recombinant human erythropoietin in combination with granulocyte colony-stimulating factor : a multicenter randomized controlled study." Eur.J.Haematol.58. 265-272 (1997)
[文献書誌] Suzuki H.: "Early and late events in FcεRI signal transduction in human cultured mast cells." The Journal of Immunology. 159. 5881-5888 (1997)
[文献書誌] Takahashi T.: "A novel molecular approach to ex vivo hematopoietic expansion with recombinant EGFR-expressing adenovirus vector." Blood. (in press.).
[文献書誌] Toru H.: "IL-4 promotes the development of tryptase and chymase double positive human mast cells accompanied with cell maturation." Blood. (in press.).
[文献書誌] Manabe A.: "Characterization of leukemic cells in CD2/CD19 double positive acute lymphoblastic leukemia." Int.J.Hematol.(in press.).
[文献書誌] Sui X.: "Soluble IL-6 receptor with IL-6 stimulates megakaryopoiesis from human CD34_+ cells through gp 130 signaling." Blood. (in press.).
[文献書誌] Nakahata T.: "Ex vivo expansion of human primitive hemopoietic progenitors." Stem Cells.(in press.).
[文献書誌] Mukouyama Y.: "Induction of hematopoiesis by oncostatin M in the AGM region." Immunity. (in press.).
[文献書誌] Nishijima I.: "Human granulocyte-macrophage colony stimulating factor (hGM-CSF) dependent in vitro and in vivo proliferation and differentiation of all hematopoietic progenitor cells in hGM-SCF receptor transgenic mice." Allergy and Clin.Immunol.(in press.).
[文献書誌] Nishihara M.: "A combination of stem cell factor and gramulocyte xolony-stimulating factor enhances proliferation and differentiation of human progenitor cells supported by murine stromal cell line MS-5." Euro.J.Immunol.(in press.).
[文献書誌] Tsuji K.: "Interferon-γ and human megakaryopoiesis." Leukemia and Lymphoma. (in press.).
[文献書誌] Tajika K.: "IL-6 and SCF exart different effects on megakaryocyte maturation." Brit.J.Haematol.(in press.).
[文献書誌] Maruyama K.: "Characterization of peripheral blood progenitor cells mobilized by Nartograstim (N-terminal replaced granulocyte colony-stimulating factor) in normal volunteers." Bone Marrow Transplant.(in press.).
[文献書誌] Xu M.: "Stimulation of human primitive hematopoiesis by murine AGM-derived stromal cells." Blood. (in press.).
[文献書誌] Ishiguro A.: "Elevation of serum thrombopoietin precedes thrombocytopoiesis in Kawasaki disease." Thromb.Haemostasis. (in press.).

1997 年度 実績報告書

マウスAGM領域からの造血幹細胞自己複製因子のクローニング

研究代表者

中畑 龍俊 東京大学, 医科学研究所, 教授 (20110744)

研究成果

[文献書誌] Muraoka K.: "Defective response to thrombopoietin and impared expression of c-mpl mRAD in bone marrow cells in congenital amegakaryocytic thrombocytopenia." Brit.J.Haematol.96. 287-292 (1997)

[文献書誌] Eguchi M.: "Morphological and cytochemical changes in human mast cells during culture." Med.Electron Microsc.30. 25-30 (1997)

[文献書誌] Toru H.: "Interleukin-4 induces homotypic aggregation of human mast cells by promoting LFA-1/ICAM-1 adhesion molecules." Blood. 89. 3296-3302 (1997)

[文献書誌] Yanagida M.: "Effect of a chymotrypsin-like inhibitor,TPCK,on histamine release from cultured human mast cells." J.Pharm.Pharmth.49. 537-541 (1997)

[文献書誌] Ishiguro A.: "Effects and serum levels of thrombopoietin in a case of chronic thrombocytopenia with achondroplasia." Int.J.Hematol.66. 99-102 (1997)

[文献書誌] Muraoka K.: "Thrombopoietin-independent effect of interferon-γ on the proliferation of human megakaryocyte progenitors." Brit.J.Haematol.98. 265-273 (1997)

[文献書誌] Nishijima I.: "Hemopoietic and lymphopoietic responses in human GM-CSF receptor transgenic mice injected with human GM-CSF." Blood. 90. 1031-1038 (1997)

[文献書誌] Yoshida M.: "Thrombopoietin alone stimulates the early proliferation and survival of human erythroid,myeloid,and multipotential progenitors in serum-free culture." Brit.J.Haematol.98. 254-264 (1997)

[文献書誌] Umemoto Y.: "Leptin stimulates the proliferation of murine myelocytic and primitive hematopoietic progenitor cells." Blood. 90. 3438-3443 (1997)

[文献書誌] Akasaka T.: "The role of mel-18,a Mammalian Polycomb group gene,during the IL-7-dependent proliferation of lymphocyte precursors." Immunity. 7. 135-146 (1997)

[文献書誌] Ebihara Y.: "Synergistic action of Flt3 and gp 130 signalings in human hematopoiesis." Blood. 90. 4363-4368 (1997)

[文献書誌] Tsuji K.: "Role of soluble interleukin-6 receptor with interleukin-6 in human hematopoiesis." Hematology. 2. 143-149 (1997)

[文献書誌] Suzuki H.: "An increase in peripheral blood progenitor cells in primates by co-administration of recombinant human interleukin 6 and recombinant human granulocyte colony-stimulating factor." Transplantation. 64. 1468-1473 (1997)

[文献書誌] Bessho M.: "Treatment of the anemia of aplastic anemia patients with recombinant human erythropoietin in combination with granulocyte colony-stimulating factor : a multicenter randomized controlled study." Eur.J.Haematol.58. 265-272 (1997)

[文献書誌] Suzuki H.: "Early and late events in FcεRI signal transduction in human cultured mast cells." The Journal of Immunology. 159. 5881-5888 (1997)

[文献書誌] Takahashi T.: "A novel molecular approach to ex vivo hematopoietic expansion with recombinant EGFR-expressing adenovirus vector." Blood. (in press.).

[文献書誌] Toru H.: "IL-4 promotes the development of tryptase and chymase double positive human mast cells accompanied with cell maturation." Blood. (in press.).

[文献書誌] Manabe A.: "Characterization of leukemic cells in CD2/CD19 double positive acute lymphoblastic leukemia." Int.J.Hematol.(in press.).

[文献書誌] Sui X.: "Soluble IL-6 receptor with IL-6 stimulates megakaryopoiesis from human CD34_+ cells through gp 130 signaling." Blood. (in press.).

[文献書誌] Nakahata T.: "Ex vivo expansion of human primitive hemopoietic progenitors." Stem Cells.(in press.).

[文献書誌] Mukouyama Y.: "Induction of hematopoiesis by oncostatin M in the AGM region." Immunity. (in press.).

[文献書誌] Nishijima I.: "Human granulocyte-macrophage colony stimulating factor (hGM-CSF) dependent in vitro and in vivo proliferation and differentiation of all hematopoietic progenitor cells in hGM-SCF receptor transgenic mice." Allergy and Clin.Immunol.(in press.).

[文献書誌] Nishihara M.: "A combination of stem cell factor and gramulocyte xolony-stimulating factor enhances proliferation and differentiation of human progenitor cells supported by murine stromal cell line MS-5." Euro.J.Immunol.(in press.).

[文献書誌] Tsuji K.: "Interferon-γ and human megakaryopoiesis." Leukemia and Lymphoma. (in press.).

[文献書誌] Tajika K.: "IL-6 and SCF exart different effects on megakaryocyte maturation." Brit.J.Haematol.(in press.).

[文献書誌] Maruyama K.: "Characterization of peripheral blood progenitor cells mobilized by Nartograstim (N-terminal replaced granulocyte colony-stimulating factor) in normal volunteers." Bone Marrow Transplant.(in press.).

[文献書誌] Xu M.: "Stimulation of human primitive hematopoiesis by murine AGM-derived stromal cells." Blood. (in press.).

[文献書誌] Ishiguro A.: "Elevation of serum thrombopoietin precedes thrombocytopoiesis in Kawasaki disease." Thromb.Haemostasis. (in press.).

1997 年度実績報告書

中畑龍俊東京大学, 医科学研究所, 教授 (20110744)