Molecular cloning of the factors that biologically accelerate reparative dentin formation
Grant-in-Aid for Scientific Research (B)
|Allocation Type||Single-year Grants |
|Research Institution||Okayama University |
KUBOKI Takuo Graduate School of Medicine and Dentistry, Associate Professor, 大学院・医歯学総合研究科, 助教授 (00225195)
TAKIGAWA Masaharu Graduate School of Medicine and Dentistry, Professor, 大学院・医歯学総合研究科, 教授 (20112063)
TAKASHIBA Shougo Graduate School of Medicine and Dentistry, Professor, 大学院・医歯学総合研究科, 教授 (50226768)
SONOYAMA Wataru Dental Hospital, Research Associate, 歯学部附属病院, 助手 (40325121)
KANYAMA Manabu Hospital of Dentistry, Assistant Professor, 歯学附属病院, 講師 (90294420)
NAKANISHI Tohru Graduate School of Medicine and Dentistry, Associate Professor, 大学院・医歯学総合研究科, 助教授 (30243463)
|Project Period (FY)
2000 – 2001
Completed (Fiscal Year 2001)
|Budget Amount *help
¥13,800,000 (Direct Cost: ¥13,800,000)
Fiscal Year 2001: ¥5,100,000 (Direct Cost: ¥5,100,000)
Fiscal Year 2000: ¥8,700,000 (Direct Cost: ¥8,700,000)
|Keywords||reparative dentin / gene transfection / adenovirus vector / connective tissue growth factor / transforming growth factor-beta / odontoblast / 炎症 / クローニング / アデノウイルスベクター|
1. Gene delivery to cultured cells
We prepared recombinant adenovirus vector carrying beta-galactosidase gene. Mouse osteoblast-like cells, MC-3T3 -E1, were cultured with this vector. As a result, almost all cells were transfected with beta-galactosidase gene.
2. Localization of known factors (TGF-beta 1 and CTGF) in vivo animal model
Localization of TGF-beta 1, that is supposed to be involved in reparative dentinogenesis, and CTGF were confirmed with immunohistochemical staining in animal (wister rat) experimental model. As a result, in 2 weeks from tooth reduction reparative dentin-like tissues were observed, and strong staining of TGF-beta 1 and CTGF were observed around these tissues.
3. Gene expression of TGF-beta 1 and CTGF in cultured cells stimulated with proinflammatory factors
MDPC-23, mouse-derived odontoblast-like cells were used in this study. The cells were stimulated with IL-1 beta and bacterial LPS. Changes in CTGF and TGF-b1 genes expression were examined by RT-PCR. As a result, MDPC-23 cells were expressing the CTGF and TGF-beta 1 genes coastitutively, and both factors increased CTGF gene expression and decreased TGF-b1 gene expression in the odontoblast-like cells (MDPC-23) within one-day period after stimulation.
4. Effect of TGF-beta 1 and CTGF to cultured cells
The effects of rCTGF and rTGF-beta 1 on cell proliferation were determined by the MTT assay. rTGF-beta 1 tended to decrease the proliferation dose-dependently, whereas effect of rCTGF was not evident. Next, to investigate the effects of rCTGF on calcification of MDPC-23 cells, cells were cultured with medium containing rCTGF. Sequential addition of AA and b-GP up-regulated the ALPase activity, while addition of rCTGF had no obvious effects.
Report (3 results)
Research Products (16 results)