| Co-Investigator(Kenkyū-buntansha) |
UMEZAWA Akihiro National Institute of Child Health and Development, Reproductive Biology and Pathology, Head, 生殖医療研究部, 部長 (70213486)
SAKAMOTO Kei Tokyo Medical and Dental University, Graduate School, Research Associate, 大学院・医歯学総合研究科, 助手 (00302886)
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| Budget Amount *help |
¥11,600,000 (Direct Cost: ¥11,600,000)
Fiscal Year 2003: ¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 2002: ¥7,700,000 (Direct Cost: ¥7,700,000)
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| Research Abstract |
Notch is a trausmembrane receptor that plays a crucial role in differentiation of stem cells. Its role in the formation of mesenchymal tissues has also been implicated although detailed mechanism has not yet been analyzed. To elucidate the function of Notch signaling in osteogenesis, the constitutively active Notchi (Notch intracellular domain, NICD) was transfected into two different osteoblastic mesenchymal cell lines, Kusa-Al and Kusa-O, and established stable transformants (KusaANICD and KusaONICD) to examine the Notch signaling. NICD generally suppressed the expression of osteogenic marker genes, calcium deposition, in vitro mineralization, the promoter activities of the Cbfal and the Ose2 element in both Kusa-Al and KusaO. In vivo bone formation of Kusa-Al was significantly suppressed by NICD. These results imply that Notch signaling functions as a negative regulator on osteoblast differentiation. In order to confirm these results, the Notch signaling was suppressed by an evoluti
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onarily conserved transcription factor CBF1, also known as RBP-Jic. Transient transfection of CBF1 remarkably increased the promoter activities of Ose2 and Cbfal, but blocked the elevation of Hesi promoter activity induced by NICD transfection. In Kusa-AI/CBF1 cell line, which is a CBF1 expressing stable cell line, the osteogenic properties, including calcium deposition, in vitro mineralization and the RANKL expression were significantly promoted. The in vivo hard tissue formation was greatly facilitated in Kusa-A1/CBFI. Furthermore, the hard tissues formed by Kusa-A1/CBF1 showed more compact structure similar to trabecular bone tissues than those by Kusa-AI/host. The expression of CBF1 was ubiquitous, but it was especially high in bone tissues and bone forming structures. These, results indicated that over-expression of transcription mediator CBF1 strongly promoted the osteogenic differentiation of mesenchymal progenitor cells. Taken together, the Notch signaling should be essential in the osteoblastic differentiation, and modification of its signaling would be key to control the mineralization. Less
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