1998 年度実績報告書

サイトカインレセプタートランスジェニックマウスを用いた造血幹細胞の分化機構

研究課題

研究課題/領域番号	10307020
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) 辻浩一郎東京大学, 医科学研究所, 助教授 (50179991)
キーワード	造血幹細胞 / 造血前駆細胞 / 分化 / サイトカインレセプター / サイトカイン / in vivo / in vitro / トランスジェニック(Tg)マウス
研究概要	造血幹細胞の増殖・分化におけるサイトカインの役割を明らかにするため、本年度は好中球、マクロファージの特異的増殖・分化因子と考えられているGM-CSF,G-CSFの受容体を発現するトランスジェニック(Tg)マウスを作成し解析した。H2-Ldプロモーター下流にヒトGM-CSFRα,βcDNA,ヒトおよびマウスG-CSFR cDNAをそれぞれつないだコンストラクトを作成し、型のごとく3系統のTgマウスを作成した。誕生したマウスの尾よりDNAを調整し、hGM-CSFRα,βおよびhG-CSFR、mG-CSFRのgenomicな発現をPCR法、Southem blot法で確認したところ、7匹にhGM-CSFRα,β遺伝子が、11匹にhG-CSFR遺伝子が、5匹にmG-CSFR遺伝子が確認された。それぞれのマウスを交配し得られたFlマウス骨髄における蛋白の発現をFACSを用いて確認したところ、2系統のhGM-CSFRα,β-Tgマウス、2系統のhGCSFR-Tgマウスが得られた。mGCSFR-Tgマウスは最近出来たばかりであり、現在、Flマウスを作成中である。 hGM-CSFR-Tgマウス、hG-CSFR-Tgマウスおよびそれぞれのリッターメイトより骨髄細胞を採取し、メチルセルロース培養法を用いてhGM-CSF、hG-CSFの作用を検討した。 hGM-GSFR-Tgマウス骨髄細胞をGM-CSF存在下で培養すると好中球系前駆細胞のみならず未分化造血前駆細胞、赤芽球系前駆細胞、巨核球系前駆細胞、肥満細胞前駆細胞等の増殖と最終的な分化、成熟が得られることが明らかとなった。また、TgマウスへのGM-CSFの投与により赤血球、好中球、単球、NK細胞の著明な増加、著明な胸腺の縮小、CD4+CD8+細胞の消失などが観察されている。またhG-CSF-Tgマウスでも同様にG-CSF刺激により様々な前駆細胞の増殖・分化・成熟が見られたが、-CSF単独での赤血球までの最終分化は見られなかった。エリスロポエチン(EPO)存在下にG-CSFを添加すると赤芽球バーストの著明な増加が見られた。以上のG-CSFの作用はFACSでソーティングした細胞の単細胞培養でも確認され、G-CSFの直接作用であることが明らかとなった。さらに、ヒトG-CSF存在下で形成された芽球コロニーを取り出して、再度ヒトG-CSF存在下で培養すると、再びさまざまな血球系のコロニーが形成された。このことはGM-CSFやG-CSFは多能性造血幹細胞から好中球、マクロファージへの分化を決定する因子として作用するのではなく、受容体を発現している細胞に単なる増殖因子として働き、全ての血球の分化がもたらされることを示唆している。

研究成果

(27件)

すべてその他

すべて文献書誌 (27件)

[文献書誌] Manabe A., Mori T., Ebihara Y., Nakahata T., et al: "Characterization of leukemic cells in CD2/CD19 double positive acute lymphoblastic leukemia." Int.J.Hematol.67. 45-52 (1998)
[文献書誌] Mukouyama Y., Hara T., Xu M., Nakahata T., et al: "Induction of hematopoiesis by oncostatin M in the AGM region." Immunity. 8. 105-114 (1998)
[文献書誌] Toru H., Eguchi M., Yata J., Nakahata T., et al: "IL-4 promotes the development of tryptase and chymase double positive human mast cells accompanied with cell maturation." Blood. 91. 187-195 (1998)
[文献書誌] Nakahata T.: "Characteristics of hemopoietic stem/progenitor cells in bone marrow,peripheral blood and cord blood." Int.J.Pediatr.Hematol.Oncology. 5. 60-61 (1998)
[文献書誌] Takahashi T., Yamada K., Nakahata T., Hirai H., et al: "A novel molecular approach to ex vivo hematopoietic expansion with recombinant EGFR-expressing adenovirus vector." Blood. 91. 4509-4515 (1998)
[文献書誌] Nishihara M., Wada Y., Nakahata T., Maekawa T., et al: "A combination of stem cell factor and granulocyte colony-stimulating factor enhances the growth of human progenitor B cells supported by murine stromal cell line MS-5." Eur.J.Immunol.28. 855-864 (1998)
[文献書誌] Tsuji K., Muraoka K., Nakahata T., et al: "Interferon-g and human megakaryopoiesis." Leukemia and Lymphoma. 31. 107-113 (1998)
[文献書誌] Tajika K., Ikebuchi K., Nakahata T., Asano S., et al: "IL-6 and SCF exart different effects on megakaryocyte maturation." Brit.J.Haematol.100. 105-111 (1998)
[文献書誌] Ishiguro A., Ishikita T., Naritake S., Nakahata T., et al: "Elevation of serum thrombopoietin precedes thrombocytopoiesis in Kawasaki disease." Thromb.Haemostasis. 79. 1096-1100 (1998)
[文献書誌] Yoshida K., Taga T., Nakahata T., Kishimoto T., et al: "Myocardial,hematological, and placental disorders caused by targeted disruption of gp130, acommon signal transducer for IL-6 family of cytokines." Contemporaty Immunology. 259-289 (1998)
[文献書誌] Suzuki H., Takei M., Nakahata T., Fukamachi H., et al: "Inhibitory effect of adenosine on degranulation of human cultured mast cells upon cross-linking of FceRI." Biochem.Biophy.Res.Comm.242. 697-702 (1998)
[文献書誌] Sato T., Watanabe S., Tsuji K., Nakahata T., et al: "Induction of the erythropoietin receptor gene and acquisition of responsiveness to erythropoietin by stem cell factor in HML/SE, a human leukemic cell line." J.Biol.Chem.273. 16921-16926 (1998)
[文献書誌] Maruyama K., Tsuji K., Kodera Y., Nakahata T., et al: "Characterization of peripheral blood progenitor cells mobilized by Nartograstim (N-terminal replaced granulocyte colony-stimulating factor) in normal volunteers." Bone Marrow Transplant.22. 313-320 (1998)
[文献書誌] Xu M., Tsuji K., Miyajima A., Nakahata T., et al: "Stimulation of mouse and human primitive hematopoiesis by murine embryonic aorta-gonad-mesonephros-derived stromal cells." Blood. 192. 2032-2040 (1998)
[文献書誌] Shichijo M., Nakahata T., Saito H.,Nagai H., et al: "The effect of anti-asthma drage on mediator release from cultured human mast cells." Clin.Exp.Allergy. 28. 1229-1236 (1998)
[文献書誌] Sako M., Nakahaat T., Kishitomo T., Sugiyama H., et al: "Abnormal expression of Wilms'tumor gene WT1 in juvenile chronic myeloid leukemia and infantile monosomy 7 syndrome." Leukemia Res.22. 965-967 (1998)
[文献書誌] Toru H., Pawankar R., Yata J., Nakahata T., et al: "Human mast cells produce interleukin-13 by high affinity IgE receptor cross-linking : enhanced IL-13 production by IL-4 primed human mast cells." J.Allergy and Clin.Immunol.(in press.). (1998)
[文献書誌] Yang FC, Watanabe S., Ebihara Y., Nakahata T., et al: "Human G-CSF stimulates the development of primitive multipotential progenitors of human G-CSF receptor-transgenic mice,but does not affect their commitment in vitro and in vivo." Blood. 92. 1632-1640 (1998)
[文献書誌] Nakahata T., Sui X., Kishimoto T., et al: "Ex vivo expansion of human primitive hemopoietic progenitors." Stem Cells.(in press.). (1998)
[文献書誌] Sui X., Tsuji K., Kishimoto T., Nakahata T., et al: "Soluble IL-6 receptor with IL-6 stimulates megakaryopoiesis from human CD34+cells through gp130 signaling." Blood. (in press.). (1998)
[文献書誌] Yang FC., Tsuji K., Ikeda Y., Nakahata T., et al: "Effects of human granulocyte colony-stimulating factor (hG-CSF) on megakaryopoiesis and platelet function in hG-CSF receptor-transgenic mice." Blood. (in press.). (1998)
[文献書誌] Taniguchi T., Endo H., Nakahata T., Motokura T., et al: "Expression of p21Cip1/Waf1/Sdi1 and p27 Kip1 cyclin-dependent kinase inhibitors during human hematopoiesis." Blood. (in press.). (1998)
[文献書誌] Duraisamy K., Saito H., Tsuji K., Nakahata T., et al: "Characterization of mast-cell-committed progenitors present in human cord blood." Blood. (in press.). (1998)
[文献書誌] Nakahata T., Toru H., et al: "Development of human mast cells." Adv.Immunol.(in press.). (1998)
[文献書誌] Hasegawa S., Nakahata T., Okumura K., RaC., et al: "Functional expression of the high affinity receptor for IgE (Fc epsilon RI) in human platelets and its intracellular expression in human megakaryocytes." Blood. (in press.). (1998)
[文献書誌] Hibino H., Tani K., Nakahata T., Asano S., et al: "Common Mamoset as a target preclinical primate for cytokine and gene therapy studies." Blood. (in press.). (1998)
[文献書誌] Kobayashi M., Ueda K., Nishihira H., Nakahata T., et al: "Serum granulocyte colony-stimulating factor levels in patients with chronic neutropenia of childhood : Modulation of G-CSF levels by myeloid precursor cell mass." Brit.J.Haematol.(in press.). (1998)

1998 年度 実績報告書

サイトカインレセプタートランスジェニックマウスを用いた造血幹細胞の分化機構

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] Manabe A., Mori T., Ebihara Y., Nakahata T., et al: "Characterization of leukemic cells in CD2/CD19 double positive acute lymphoblastic leukemia." Int.J.Hematol.67. 45-52 (1998)

[文献書誌] Mukouyama Y., Hara T., Xu M., Nakahata T., et al: "Induction of hematopoiesis by oncostatin M in the AGM region." Immunity. 8. 105-114 (1998)

[文献書誌] Toru H., Eguchi M., Yata J., Nakahata T., et al: "IL-4 promotes the development of tryptase and chymase double positive human mast cells accompanied with cell maturation." Blood. 91. 187-195 (1998)

[文献書誌] Nakahata T.: "Characteristics of hemopoietic stem/progenitor cells in bone marrow,peripheral blood and cord blood." Int.J.Pediatr.Hematol.Oncology. 5. 60-61 (1998)

[文献書誌] Takahashi T., Yamada K., Nakahata T., Hirai H., et al: "A novel molecular approach to ex vivo hematopoietic expansion with recombinant EGFR-expressing adenovirus vector." Blood. 91. 4509-4515 (1998)

[文献書誌] Nishihara M., Wada Y., Nakahata T., Maekawa T., et al: "A combination of stem cell factor and granulocyte colony-stimulating factor enhances the growth of human progenitor B cells supported by murine stromal cell line MS-5." Eur.J.Immunol.28. 855-864 (1998)

[文献書誌] Tsuji K., Muraoka K., Nakahata T., et al: "Interferon-g and human megakaryopoiesis." Leukemia and Lymphoma. 31. 107-113 (1998)

[文献書誌] Tajika K., Ikebuchi K., Nakahata T., Asano S., et al: "IL-6 and SCF exart different effects on megakaryocyte maturation." Brit.J.Haematol.100. 105-111 (1998)

[文献書誌] Ishiguro A., Ishikita T., Naritake S., Nakahata T., et al: "Elevation of serum thrombopoietin precedes thrombocytopoiesis in Kawasaki disease." Thromb.Haemostasis. 79. 1096-1100 (1998)

[文献書誌] Yoshida K., Taga T., Nakahata T., Kishimoto T., et al: "Myocardial,hematological, and placental disorders caused by targeted disruption of gp130, acommon signal transducer for IL-6 family of cytokines." Contemporaty Immunology. 259-289 (1998)

[文献書誌] Suzuki H., Takei M., Nakahata T., Fukamachi H., et al: "Inhibitory effect of adenosine on degranulation of human cultured mast cells upon cross-linking of FceRI." Biochem.Biophy.Res.Comm.242. 697-702 (1998)

[文献書誌] Sato T., Watanabe S., Tsuji K., Nakahata T., et al: "Induction of the erythropoietin receptor gene and acquisition of responsiveness to erythropoietin by stem cell factor in HML/SE, a human leukemic cell line." J.Biol.Chem.273. 16921-16926 (1998)

[文献書誌] Maruyama K., Tsuji K., Kodera Y., Nakahata T., et al: "Characterization of peripheral blood progenitor cells mobilized by Nartograstim (N-terminal replaced granulocyte colony-stimulating factor) in normal volunteers." Bone Marrow Transplant.22. 313-320 (1998)

[文献書誌] Xu M., Tsuji K., Miyajima A., Nakahata T., et al: "Stimulation of mouse and human primitive hematopoiesis by murine embryonic aorta-gonad-mesonephros-derived stromal cells." Blood. 192. 2032-2040 (1998)

[文献書誌] Shichijo M., Nakahata T., Saito H.,Nagai H., et al: "The effect of anti-asthma drage on mediator release from cultured human mast cells." Clin.Exp.Allergy. 28. 1229-1236 (1998)

[文献書誌] Sako M., Nakahaat T., Kishitomo T., Sugiyama H., et al: "Abnormal expression of Wilms'tumor gene WT1 in juvenile chronic myeloid leukemia and infantile monosomy 7 syndrome." Leukemia Res.22. 965-967 (1998)

[文献書誌] Toru H., Pawankar R., Yata J., Nakahata T., et al: "Human mast cells produce interleukin-13 by high affinity IgE receptor cross-linking : enhanced IL-13 production by IL-4 primed human mast cells." J.Allergy and Clin.Immunol.(in press.). (1998)

[文献書誌] Yang FC, Watanabe S., Ebihara Y., Nakahata T., et al: "Human G-CSF stimulates the development of primitive multipotential progenitors of human G-CSF receptor-transgenic mice,but does not affect their commitment in vitro and in vivo." Blood. 92. 1632-1640 (1998)

[文献書誌] Nakahata T., Sui X., Kishimoto T., et al: "Ex vivo expansion of human primitive hemopoietic progenitors." Stem Cells.(in press.). (1998)

[文献書誌] Sui X., Tsuji K., Kishimoto T., Nakahata T., et al: "Soluble IL-6 receptor with IL-6 stimulates megakaryopoiesis from human CD34+cells through gp130 signaling." Blood. (in press.). (1998)

[文献書誌] Yang FC., Tsuji K., Ikeda Y., Nakahata T., et al: "Effects of human granulocyte colony-stimulating factor (hG-CSF) on megakaryopoiesis and platelet function in hG-CSF receptor-transgenic mice." Blood. (in press.). (1998)

[文献書誌] Taniguchi T., Endo H., Nakahata T., Motokura T., et al: "Expression of p21Cip1/Waf1/Sdi1 and p27 Kip1 cyclin-dependent kinase inhibitors during human hematopoiesis." Blood. (in press.). (1998)

[文献書誌] Duraisamy K., Saito H., Tsuji K., Nakahata T., et al: "Characterization of mast-cell-committed progenitors present in human cord blood." Blood. (in press.). (1998)

[文献書誌] Nakahata T., Toru H., et al: "Development of human mast cells." Adv.Immunol.(in press.). (1998)

[文献書誌] Hasegawa S., Nakahata T., Okumura K., RaC., et al: "Functional expression of the high affinity receptor for IgE (Fc epsilon RI) in human platelets and its intracellular expression in human megakaryocytes." Blood. (in press.). (1998)

[文献書誌] Hibino H., Tani K., Nakahata T., Asano S., et al: "Common Mamoset as a target preclinical primate for cytokine and gene therapy studies." Blood. (in press.). (1998)

[文献書誌] Kobayashi M., Ueda K., Nishihira H., Nakahata T., et al: "Serum granulocyte colony-stimulating factor levels in patients with chronic neutropenia of childhood : Modulation of G-CSF levels by myeloid precursor cell mass." Brit.J.Haematol.(in press.). (1998)

1998 年度実績報告書

中畑龍俊東京大学, 医科学研究所, 教授 (20110744)