2001 Fiscal Year Annual Research Report

骨格形成における軟骨系シグナル分子群の機能解析

Research Project

Project/Area Number	13045011
Research Institution	Tokyo Medical and Dental University
Principal Investigator	野田政樹東京医科歯科大学, 難治疾患研究所, 教授 (50231725)
Co-Investigator(Kenkyū-buntansha)	辻邦和東京医科歯科大学, 難治疾患研究所, 助手 (20323694) 二藤彰東京医科歯科大学, 難治疾患研究所, 助教授 (00240747)
Keywords	軟骨形成 / 分化シグナル / 転写 / プロモーター / 骨格形成 / Cbfa1 / レチノイン酸
Research Abstract	本年度においては軟骨のシグナル系としてCbfa1の制御に関わるヘッジホック遺伝子の機能を検討した。この結果ヘッジホッグの存在により、初代培養軟骨細胞においてCbfa1の発現レベルの上昇が観察され、更にCbfa1のプロモーターの解析によりヘッジホッグによるシグナルが転写を制御する経路にあることを明らかにした。更に軟骨分化系シグナルであるSox9の発現とその制御について検討を行った。この結果、Sox9分子が初代軟骨培養細胞において高いレベルにおいて発現すると共に、レチノイン酸によってその発現が急激に抑制されることが明らかとなり、その抑制様式は用量依存性で0.5μMから観察され、1:0μMでピークで達した。更にレチノイン酸によるSox9の遺伝子発現は、時間依存性で12時間以内に最大の抑制が観察された。レチノイン酸の軟骨分化制御転写因子Sox9の遺伝子発現に対する作用が遺伝子発現のどのレベルにあるかを検討した結果、ヌクレアーランオンアッセイによって単離された核の中における転写がSox9により抑制される一方で、形成されたメッセンジャーRNAの安定性を検討した結果からは、レチノイン酸の存在によってもメッセンジャーRNAの半減期は殆ど変化がない。これらの結果から軟骨分化シグナルの転写因子であるSox9の発現はレチノイン酸によって転写を中心とした制御を介することが明らかとなった。またウエスタンブロッド法によってレチノイン酸によるSox9遺伝子の発現調節が69kDのSox9の蛋白そのものの発現を抑制することであることを示した。次にSox9の強制発現によってレチノイン酸によるII型コラーゲンプロモーターの活性への影響を調べたところ、Sox9は軟骨分化促進シグナルとしてII型コラーゲンプロモーターのエンハンサーの活性を促進したが、レチノイン酸の存在によってSox9による促進活性が阻止されることが明らかとなった。以上より、軟骨系シグナル分子であるSox9、ヘッジホッグの細胞の分化に対する機能が明らかとなった。

Research Products

(21 results)

All Other

All Publications (21 results)

[Publications] Ishijima M: "Enhancement of osteoctastic 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. 19. 399-404 (2001)
[Publications] Ihara H.: "Parathyroid hormone-induced bone resorption does not occur in the absence of osteopontin"Journal of Biological Chemistry. 276. 13065-13071 (2001)
[Publications] Asou Y: "Osteopontin facilitates angiogenesis, accumulation of osteoclasts and resorption in ectopic bone"ndocrinology. 142. 1325-1332 (2001)
[Publications] Nemoto H: "Osteopontin deficiency reduces experimental tumor cell metastasis to bone and soft tissues"Journal of Bone and Mineral Research. 6. 652-659 (2001)
[Publications] Yamashita T: "Klotho-deficient mice are resistant to bone loss induced by unloading due to sciatic neurectomy"Journal of Endocrinology. 168. 347-351 (2001)
[Publications] Kawaguchi N: "Spontaneous mutation in Mitf gene causes osteopetrosis in silver homozygote quail"Developmental Dynamics. 220. 133-140 (2001)
[Publications] Sekiya I: "Transcriptional suppression of SOX9 expression by retinoic acid in chondrocytes"Journal of Cellular Biochemistry. S36. 71-78 (2001)
[Publications] Tsuji K: "Transient suppression of core-binding factor alpha 1 expression by basic fibroblast growth factor in rat osteoblast-like osteosarcoma ROS17/2.8 cells"Journal of Bone Miner Metabolism. 19. 213-219 (2001)
[Publications] Sekiya I: "Dexamethasone enhances Sox9 expression in chondrocytes"Journal of Endocrinology. 169. 573-579 (2001)
[Publications] Denhardt, DT: "Osteopontin as a means to cope with environmental insults: regulation of inflammation, tissue remodeling, and cell survival"Journal of Clinical Investigation. 107. 1055-1061 (2001)
[Publications] Tsuji K: "Vitamin D_3 enhances the expression of I-mfa, an inhibitor of the MyoD family, in osteoblasts"Biochemica et Biophysica Acta. 1539. 122-130 (2001)
[Publications] Yoshida Y: "Negative regulation of BMP/Smad signaling by Tob in osteoblasts"Cell. 103. 1085-1097 (2000)
[Publications] Yamashita T: "Retardation in bone resorption after bone marrow ablation in klotho mutant mice"Endocrinology. 141. 438-445 (2000)
[Publications] Sekiya I: "OX9 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)
[Publications] Yamashita T: "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)
[Publications] Murakami S: "Indian hedgehog during fracture healing in adult rat femora"Calcified Tissue International. 66. 272-276 (2000)
[Publications] Takazawa Y: "CBFA1, is constitutively expressed in chondrocytic cell line, TC6, and its expression is upregulated by BMP2"Journal of Endocrinology. 162. 579-586 (2000)
[Publications] Tsuji K: "Identification and Expression of a novel 3-exon of mouse Runx1/Pebp2 α B/Cbfa2/AML1 gene"Biochemical and Biophysical Research Communications. 274. 171-176 (2000)
[Publications] Kawaguchi N: "Mitf is expressed in osteoclast progenitors in vitro"Experimental Cell Research. 260. 284-291 (2000)
[Publications] Furuya K: "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)
[Publications] Shiraki-Iida T: "Improvement of multiple pathophysiological phenotypes of klotho (kl/kl) mice by adenovirus-mediated expression of the klotho gene"Journal of Gene Med. 2. 233-242 (2000)

2001 Fiscal Year Annual Research Report

骨格形成における軟骨系シグナル分子群の機能解析

Principal Investigator

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

Research Products

[Publications] Ishijima M: "Enhancement of osteoctastic 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. 19. 399-404 (2001)

[Publications] Ihara H.: "Parathyroid hormone-induced bone resorption does not occur in the absence of osteopontin"Journal of Biological Chemistry. 276. 13065-13071 (2001)

[Publications] Asou Y: "Osteopontin facilitates angiogenesis, accumulation of osteoclasts and resorption in ectopic bone"ndocrinology. 142. 1325-1332 (2001)

[Publications] Nemoto H: "Osteopontin deficiency reduces experimental tumor cell metastasis to bone and soft tissues"Journal of Bone and Mineral Research. 6. 652-659 (2001)

[Publications] Yamashita T: "Klotho-deficient mice are resistant to bone loss induced by unloading due to sciatic neurectomy"Journal of Endocrinology. 168. 347-351 (2001)

[Publications] Kawaguchi N: "Spontaneous mutation in Mitf gene causes osteopetrosis in silver homozygote quail"Developmental Dynamics. 220. 133-140 (2001)

[Publications] Sekiya I: "Transcriptional suppression of SOX9 expression by retinoic acid in chondrocytes"Journal of Cellular Biochemistry. S36. 71-78 (2001)

[Publications] Tsuji K: "Transient suppression of core-binding factor alpha 1 expression by basic fibroblast growth factor in rat osteoblast-like osteosarcoma ROS17/2.8 cells"Journal of Bone Miner Metabolism. 19. 213-219 (2001)

[Publications] Sekiya I: "Dexamethasone enhances Sox9 expression in chondrocytes"Journal of Endocrinology. 169. 573-579 (2001)

[Publications] Denhardt, DT: "Osteopontin as a means to cope with environmental insults: regulation of inflammation, tissue remodeling, and cell survival"Journal of Clinical Investigation. 107. 1055-1061 (2001)

[Publications] Tsuji K: "Vitamin D_3 enhances the expression of I-mfa, an inhibitor of the MyoD family, in osteoblasts"Biochemica et Biophysica Acta. 1539. 122-130 (2001)

[Publications] Yoshida Y: "Negative regulation of BMP/Smad signaling by Tob in osteoblasts"Cell. 103. 1085-1097 (2000)

[Publications] Yamashita T: "Retardation in bone resorption after bone marrow ablation in klotho mutant mice"Endocrinology. 141. 438-445 (2000)

[Publications] Sekiya I: "OX9 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)

[Publications] Yamashita T: "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)

[Publications] Murakami S: "Indian hedgehog during fracture healing in adult rat femora"Calcified Tissue International. 66. 272-276 (2000)

[Publications] Takazawa Y: "CBFA1, is constitutively expressed in chondrocytic cell line, TC6, and its expression is upregulated by BMP2"Journal of Endocrinology. 162. 579-586 (2000)

[Publications] Tsuji K: "Identification and Expression of a novel 3-exon of mouse Runx1/Pebp2 α B/Cbfa2/AML1 gene"Biochemical and Biophysical Research Communications. 274. 171-176 (2000)

[Publications] Kawaguchi N: "Mitf is expressed in osteoclast progenitors in vitro"Experimental Cell Research. 260. 284-291 (2000)

[Publications] Furuya K: "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)

[Publications] Shiraki-Iida T: "Improvement of multiple pathophysiological phenotypes of klotho (kl/kl) mice by adenovirus-mediated expression of the klotho gene"Journal of Gene Med. 2. 233-242 (2000)

野田政樹東京医科歯科大学, 難治疾患研究所, 教授 (50231725)