2000 年度実績報告書

Cbfal遺伝子を用いた骨・軟骨再生の為の骨髄由来間葉系幹細胞の開発

研究課題

研究課題/領域番号	12557123
研究機関	東京医科歯科大学
研究代表者	野田政樹東京医科歯科大学, 難治疾患研究所, 教授 (50231725)
研究分担者	山下照仁東京医科歯科大学, 難治疾患研究所, 助手 (90302893) 二藤彰東京医科歯科大学, 難治疾患研究所, 助教授 (00240747)
キーワード	骨芽細胞 / 骨髄幹細胞 / Cbfa / 骨再生 / 転写因子 / プロモーター解析 / 軟骨細胞 / BMP
研究概要	本年においては骨芽細胞分化を決定する転写因子Cbfa1のプロモーター領域を解析した。このプロモーター領域のクローニングにより1.8kbのクローンを得て、そのフラッグメントをルシフェラーゼに繋ぎこれを骨芽細胞および軟骨細胞に導入してその活性を検討した。この結果、1.8kbは0.33kbよりも弱い活性を示し、従って1.8kbと0.33kbの間に抑制性のエレメントが存在することが推察された。次に0.33kbより0.16kbの間で検討すると、0.16kbの活性が弱くなり従って0.33kbと0.16kbの間に正の促進活性を示すシグナルがあることが推察された。このコンストラクトを骨芽細胞ならびに軟骨細胞に導入することによってその活性を検討した。軟骨細胞と骨芽細胞ではほぼ同様の活性のパターンを示した。次にこの活性に対するBMPの影響を検討した。1.8kbのフラグメントはBMPに対して正の統制を示し、また0.33kbのフラグメントをBMPに対して正の応答性を初代培養細胞の軟骨細胞で示した。一方0.16kbではその応答性が減少し従ってBMPによるこのフラグメントの水溶活性は0.33kbと0.16kbの間にあるものと推察された。骨髄の細胞としては、これまでに正常な骨芽細胞および遺伝子改変マウスからの骨芽細胞として、TOBのノックアウトマウスからの骨芽細胞ならびに、細胞に蛍光標識を行ったトランスジェニックマウスからの骨芽細胞からの樹立を行い、今後これらの細胞に対する遺伝子導入のための基礎となる材料が本年の研究により整えられた。今後このフラグメントを用い、また骨髄細胞株を樹立してこのフラグメントによる活性のもとに間葉系の幹細胞としての性質を持つ細胞の機能を得るべく検索を行う予定である。

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] Ishijima,M.,Rittling,S.R.,Yamashita,T.,Tsuji,K.,Kurosawa,H.,Nifuji,A.,Denhardt,D.T.,and Noda,M.: "Enhancement of osteoclastic bone resorption and suppression of osteoblastic bone formation in response to reduced mechanical stress do not occur in the absence of osteopontin."Journal of Experimental Medicine. 193. 399-404 (2001)
[文献書誌] Ihara H.Denhardt D.T.,Furuya,K.,Yamasita,T.,Muguruma,Y.,Tsuji,K.,Hruska.K.A,Higashio,K.,Enomoto,S.,Nifuji,A.,Rittling S.R.,and Noda,M.: "Parathyroid hormone-induced bone resorption does not occur in the absence of osteopontin."Journal of Biological Chemistry. 276(In Press). (2001)
[文献書誌] Asou Y.,Rittling S.R.,Yoshitake H.,Tsuji K.,Shinomiya K.,Nifuji A.,Denhardt D.T.,and Noda M.: "Osteopontin facilitates angiogenesis, accumulation of osteoclasts and resorption in ectopic bone."Endocrinology. 142. 1325-1332 (2001)
[文献書誌] Nemota H.,Rittling S.R,Yoshitako H.,Furuya K,Amagasa T.,Tsuji K,Nifuji A,Denhardt D.T,and Noda M.: "Osteopontin deficiency reduces experimental tumor cell metastasis to bone and soft tissues."Journal of Bone and Mineral Research. (In Press). (2001)
[文献書誌] Sekiya,I.,Koopman,P.,Tsuji, K.,Mertin,S.,Harley,V.,Yamada,Y.,Shinomiya,K.,Nifuji,A.,and Noda,M.: "Dexamethasone Enhances SOX9 Expression in Chondrocytes."Journal of Endocrinology. (In Press). (2001)
[文献書誌] Tsuji,K.,Kraut,N.,Groudine,M.,and Noda,M.: "Vitamin D3 Enhances the Expression of I-mfa, an Inhibitor of MyoD Family, in Osteoblasts."Biochimica et Biophysica Acta. (In Press). (2001)
[文献書誌] Tsuji,K.and Noda,M.: "Transient Suppression of Cbfa1 Expression by Basic Fibroblast Growth Factor (bFGF) in Rat Osteoblast-like Osteosarcoma ROS17/2.8 Cells."Journal of Bone and Mineral Metabolism. (In Press). (2001)
[文献書誌] Sekiya,I.,Koopman,P.,Tsuji,K.,Mertin,S.,Harley,V.,Yamada,Y.,Shinomiya,K.,Nifuji,A.,and Noda,M.: "Transcriptional suppression of SOX9 expression by retinoic acid in chondrocytes."Journal of Cellular Biochemistry. (In Press). (2001)
[文献書誌] Kawaguchi N.,Ono T.,Mochii M.and Noda M.: "Spontaneous mutation in Mitf gene causes osteopetrosis in silver homozygote quail."Developmental Dynamics. 220. 133-140 (2001)
[文献書誌] Yamashita T.,Sekiya I.,Kawaguchi N.,Kashimada,K.,Nifuji,A.,Nabeshma Y.and Noda M. : "Klotho-deficient mice are resistant to bone loss induced by unloading due to sciatic neurectomy."Journal of Endocrinology. 168. 347-351 (2001)
[文献書誌] Yoshida Y.,Tanaka S.Umamon H.Minowa O.,Usul M.,Ikomatsu N.Hosoda E.,Imamura T.Kuro J.Yamashita T.,Miyazono K.,Noda M.,Noda T.,and Yamamoto T.: "Negative regulation of BMP/Smad signaling by Tob in osteoblasts."Cell. 103. 1085-1097 (2000)
[文献書誌] Shiraki-Iida T,Iida A,Nabeshima Y,Anazawa H,Nishikawa S,Noda M,Kuro-o M,Nabeshima Y.: "Improvement of multiple pathophysiological phenotypes of klotho (kl/kl) mice by adenovirus-mediated expression of the klotho gene."J Gene Med. 2. 233-42 (2000)
[文献書誌] Furuya K.,Nakamoto T.,Shen Z. J.,Tsuji K.,Nifuji A.,Hirai H.and Noda M.: "Overexpression of Cas-interactiong zinc finger protein (CIZ) suppresses proliferation and enhances expression of type I collagen gene in osteoblast-like MC3T3E1 cells."Experimental Cell Research. 261. 329-335 (2000)
[文献書誌] Kawaguchi N.and Noda M.: "Mitf is expressed in osteoclast progenitors in vitro."Experimental Cell Research. 260. 284-291 (2000)
[文献書誌] Tsuji K.and Noda M.: "Identification and Expression of a novel 3′-exon of mouse Runx1/Pebp2αB/Cbfa2/AML1 gene."Biochemical and Biophysical Research Communications. 274. 171-176 (2000)
[文献書誌] Sekiya I.,Tsuji K.,Koopman P.,Watanabe H.,Yamada Y.,Shinomiya K.,Nifuji A.,and Noda M: "SOX9 enhances aggrecan gene promoter/enhancer activity and is upregulated by Retinoic Acid in a Cartilage-Derived Cell Line, TC6."Journal of Biological Chemistry. 275. 10738-10744 (2000)
[文献書誌] Takazawa Y.,Tsuji K.,Nifuji A.,Kurosawa H.,Ito Y.and Noda M.: "CBFA1, IS Constitutively expressed in chondrocytic cell line, TC6, and its expression is upregulated byBMP2."Journal of Endocrinology. 165. 579-586 (2000)
[文献書誌] Murakami S.and Noda M.: "Indian hedgehog during fracture healing in adult rat femora"Calcified Tissue International. 66. 272-276 (2000)
[文献書誌] Yamashita T.,Yoshitake H.,Tsuji K.,Kawaguchi N.,Nabeshima Y.,and Noda M.: "Retardation in bone resorption after bone marrow ablation in klotho mutant mice."Endocrinology. 141. 438-445 (2000)
[文献書誌] Yamashita T.,Nabeshima Y. and Noda M.: "High-resolution uCT (micro-computed tomography) analyses of the abnormal trabecular bone structures in klotho gene mutant mice."Journal of Endocrinology. 164. 239-245 (2000)
[文献書誌] Hijikata M,Matsumoto HN,Kobayashi A,Nifuji A,Noda M,Natori S.: "Induction of apoptosis of monocyte-macrophage lineage cells by 5-S-GAD."FEBS Lett.. 457. 405-408 (1999)
[文献書誌] Nifuji A.,and Noda M.: "Coordinated expression of noggin and BMPs during early skeletogenesis and induction of noggin expression by BMP7."Journal of Bone and Mineral Research. 14. 2057-66 (1999)
[文献書誌] Yoshitake Y.,Ritting S.R.,Denhardt D.T.and Noda M.: "Osteopontin-deficient mice are resistant to ovariectomy-induced bone resorption."Proceedings of the National Academy of Sciences. 96. 8156-8160 (1999)
[文献書誌] Takazawa Y.,Nifuji A.,Mataga N.,Yamauchi Y.,Kurosawa H.,and Noda M.: "Articular cartilage immortalized by a temperature sensitive mutant of SV40 large T antigen survive and form cartilage tissue in articular cartilage environment."Journal of Cellular Biochemistry. 74. 338-345 (1999)
[文献書誌] Nifuji A.,Odile Kellermann,and Noda M.: "Noggin expression in a mesodermal pluripotent cell line C1 and its regulation by BMP."Journal of Cellular Biochemistry. 73. 437-444 (1999)
[文献書誌] Furuya K.,Nifuji A.,Rosen V.,and Noda M.: "Effects of GDF7/BMP12 on proliferation and alkaline phosphatase expression in rat osteoblastic osteosarcoma ROS 17/2.8 cells."Journal of Cellular Biochemistry. 72. 177-180 (1999)
[文献書誌] Tamura M.and Noda M.: "Identification of DERMO-1 as a member of helix-loop-helix type transcription factors expressed in osteoblastic cells."Journal of Cellular Biochemistry. 72. 167-176 (1999)

2000 年度 実績報告書

Cbfal遺伝子を用いた骨・軟骨再生の為の骨髄由来間葉系幹細胞の開発

研究代表者

野田 政樹 東京医科歯科大学, 難治疾患研究所, 教授 (50231725)

研究成果

[文献書誌] Ihara H.Denhardt D.T.,Furuya,K.,Yamasita,T.,Muguruma,Y.,Tsuji,K.,Hruska.K.A,Higashio,K.,Enomoto,S.,Nifuji,A.,Rittling S.R.,and Noda,M.: "Parathyroid hormone-induced bone resorption does not occur in the absence of osteopontin."Journal of Biological Chemistry. 276(In Press). (2001)

[文献書誌] Asou Y.,Rittling S.R.,Yoshitake H.,Tsuji K.,Shinomiya K.,Nifuji A.,Denhardt D.T.,and Noda M.: "Osteopontin facilitates angiogenesis, accumulation of osteoclasts and resorption in ectopic bone."Endocrinology. 142. 1325-1332 (2001)

[文献書誌] Nemota H.,Rittling S.R,Yoshitako H.,Furuya K,Amagasa T.,Tsuji K,Nifuji A,Denhardt D.T,and Noda M.: "Osteopontin deficiency reduces experimental tumor cell metastasis to bone and soft tissues."Journal of Bone and Mineral Research. (In Press). (2001)

[文献書誌] Sekiya,I.,Koopman,P.,Tsuji, K.,Mertin,S.,Harley,V.,Yamada,Y.,Shinomiya,K.,Nifuji,A.,and Noda,M.: "Dexamethasone Enhances SOX9 Expression in Chondrocytes."Journal of Endocrinology. (In Press). (2001)

[文献書誌] Tsuji,K.,Kraut,N.,Groudine,M.,and Noda,M.: "Vitamin D3 Enhances the Expression of I-mfa, an Inhibitor of MyoD Family, in Osteoblasts."Biochimica et Biophysica Acta. (In Press). (2001)

[文献書誌] Tsuji,K.and Noda,M.: "Transient Suppression of Cbfa1 Expression by Basic Fibroblast Growth Factor (bFGF) in Rat Osteoblast-like Osteosarcoma ROS17/2.8 Cells."Journal of Bone and Mineral Metabolism. (In Press). (2001)

[文献書誌] Sekiya,I.,Koopman,P.,Tsuji,K.,Mertin,S.,Harley,V.,Yamada,Y.,Shinomiya,K.,Nifuji,A.,and Noda,M.: "Transcriptional suppression of SOX9 expression by retinoic acid in chondrocytes."Journal of Cellular Biochemistry. (In Press). (2001)

[文献書誌] Kawaguchi N.,Ono T.,Mochii M.and Noda M.: "Spontaneous mutation in Mitf gene causes osteopetrosis in silver homozygote quail."Developmental Dynamics. 220. 133-140 (2001)

[文献書誌] Yamashita T.,Sekiya I.,Kawaguchi N.,Kashimada,K.,Nifuji,A.,Nabeshma Y.and Noda M. : "Klotho-deficient mice are resistant to bone loss induced by unloading due to sciatic neurectomy."Journal of Endocrinology. 168. 347-351 (2001)

[文献書誌] Yoshida Y.,Tanaka S.Umamon H.Minowa O.,Usul M.,Ikomatsu N.Hosoda E.,Imamura T.Kuro J.Yamashita T.,Miyazono K.,Noda M.,Noda T.,and Yamamoto T.: "Negative regulation of BMP/Smad signaling by Tob in osteoblasts."Cell. 103. 1085-1097 (2000)

[文献書誌] Shiraki-Iida T,Iida A,Nabeshima Y,Anazawa H,Nishikawa S,Noda M,Kuro-o M,Nabeshima Y.: "Improvement of multiple pathophysiological phenotypes of klotho (kl/kl) mice by adenovirus-mediated expression of the klotho gene."J Gene Med. 2. 233-42 (2000)

[文献書誌] Kawaguchi N.and Noda M.: "Mitf is expressed in osteoclast progenitors in vitro."Experimental Cell Research. 260. 284-291 (2000)

[文献書誌] Tsuji K.and Noda M.: "Identification and Expression of a novel 3′-exon of mouse Runx1/Pebp2αB/Cbfa2/AML1 gene."Biochemical and Biophysical Research Communications. 274. 171-176 (2000)

[文献書誌] Sekiya I.,Tsuji K.,Koopman P.,Watanabe H.,Yamada Y.,Shinomiya K.,Nifuji A.,and Noda M: "SOX9 enhances aggrecan gene promoter/enhancer activity and is upregulated by Retinoic Acid in a Cartilage-Derived Cell Line, TC6."Journal of Biological Chemistry. 275. 10738-10744 (2000)

[文献書誌] Takazawa Y.,Tsuji K.,Nifuji A.,Kurosawa H.,Ito Y.and Noda M.: "CBFA1, IS Constitutively expressed in chondrocytic cell line, TC6, and its expression is upregulated byBMP2."Journal of Endocrinology. 165. 579-586 (2000)

[文献書誌] Murakami S.and Noda M.: "Indian hedgehog during fracture healing in adult rat femora"Calcified Tissue International. 66. 272-276 (2000)

[文献書誌] Yamashita T.,Yoshitake H.,Tsuji K.,Kawaguchi N.,Nabeshima Y.,and Noda M.: "Retardation in bone resorption after bone marrow ablation in klotho mutant mice."Endocrinology. 141. 438-445 (2000)

[文献書誌] Yamashita T.,Nabeshima Y. and Noda M.: "High-resolution uCT (micro-computed tomography) analyses of the abnormal trabecular bone structures in klotho gene mutant mice."Journal of Endocrinology. 164. 239-245 (2000)

[文献書誌] Hijikata M,Matsumoto HN,Kobayashi A,Nifuji A,Noda M,Natori S.: "Induction of apoptosis of monocyte-macrophage lineage cells by 5-S-GAD."FEBS Lett.. 457. 405-408 (1999)

[文献書誌] Nifuji A.,and Noda M.: "Coordinated expression of noggin and BMPs during early skeletogenesis and induction of noggin expression by BMP7."Journal of Bone and Mineral Research. 14. 2057-66 (1999)

[文献書誌] Yoshitake Y.,Ritting S.R.,Denhardt D.T.and Noda M.: "Osteopontin-deficient mice are resistant to ovariectomy-induced bone resorption."Proceedings of the National Academy of Sciences. 96. 8156-8160 (1999)

[文献書誌] Takazawa Y.,Nifuji A.,Mataga N.,Yamauchi Y.,Kurosawa H.,and Noda M.: "Articular cartilage immortalized by a temperature sensitive mutant of SV40 large T antigen survive and form cartilage tissue in articular cartilage environment."Journal of Cellular Biochemistry. 74. 338-345 (1999)

[文献書誌] Nifuji A.,Odile Kellermann,and Noda M.: "Noggin expression in a mesodermal pluripotent cell line C1 and its regulation by BMP."Journal of Cellular Biochemistry. 73. 437-444 (1999)

[文献書誌] Furuya K.,Nifuji A.,Rosen V.,and Noda M.: "Effects of GDF7/BMP12 on proliferation and alkaline phosphatase expression in rat osteoblastic osteosarcoma ROS 17/2.8 cells."Journal of Cellular Biochemistry. 72. 177-180 (1999)

[文献書誌] Tamura M.and Noda M.: "Identification of DERMO-1 as a member of helix-loop-helix type transcription factors expressed in osteoblastic cells."Journal of Cellular Biochemistry. 72. 167-176 (1999)

2000 年度実績報告書

野田政樹東京医科歯科大学, 難治疾患研究所, 教授 (50231725)