Exploring new candidate genes for pulpal mineralization using micro array system

利用微阵列系统探索牙髓矿化的新候选基因

• 批准号: 12671849
• 负责人: KAWASHIMA Nobuyuki
• 金额: $ 2.5万
• 依托单位: Tokyo Medical and Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671849/
• 关键词: Mineralization; micro array; osteoblasts; Kusa A; Kusa O; Alkaline phosphatase; osteocalcin; pulpal cells; オステロカルシン; 歯髄生物; cDNAアレイ; β-glycerophosphate; ascorbic acid; Kusa A; 骨髄間質細胞

The purpose of this study is to find out new candidates that are responsible to mineralization, and to determine their expression in the dental pulp in order to establish a new method of hard tissue formation in the pulp. First, gene expression profiles were compared between Kusa A and Kusa.O cells, which are matured and immatured osteoblastic cell lines respectively. Next, gene expression profiles were compared between Kusa cells cultured in calcifying condition and in normal condition, that are presence of ascorbic acid (AA) and b-glycerophosphate (bGP) or absence of them respectively. Kusa A and O cells are derived from mouse stromal cells, and both cell lines exhibit high alkaline phosphatase activity. Kusa A cells showed in vivo calcification using diffusion chamber transfer into the mice peritoneal, but Kusa O cells did not. Furthermore, Kusa A demonstrated osteocalcin gene expression and in vitro mineralization, therefore Kusa A thought, to be highly differentiated osteoblasts. On the other hand, Kusa 0 cells thought to be immature osteoblasts. Gene expression profile were compared between Kusa A and O cells, and several genes were up regulated and dowirregulated. AA and bGP rapidly depressed the many gene expression. Originally used CDNA array were plotted only 588 genes, and further studies were performed with the Mouse GEM CDNA array (Kurabo) in which 8700 clones were plotted. Several interesting genes were picked, and functional analysis is ongoing.

本研究的目的是寻找新的与矿化有关的候选基因，并确定它们在牙髓中的表达，以期建立一种新的牙髓硬组织形成方法。首先，比较了成骨细胞系Kusa A和成骨细胞系Kusa.O的基因表达谱。然后，比较在钙化条件下培养的Kusa细胞和正常条件下培养的Kusa细胞在有无抗坏血酸(AA)和b-甘油磷酸(BGP)情况下的基因表达谱。KusaA和O细胞来源于小鼠基质细胞，这两种细胞系都表现出很高的碱性磷酸酶活性。扩散腔移植到小鼠腹膜后，Kusa A细胞显示体内钙化，而Kusa O细胞未见钙化。此外，Kusa A还表现出骨钙素基因的表达和体外矿化，因此Kusa A认为是高度分化的成骨细胞。另一方面，Kusa 0细胞被认为是未成熟的成骨细胞。比较Kusa A和O细胞的基因表达谱，发现有几个基因表达上调和下调。AA和BGP迅速抑制了许多基因的表达。最初使用的CDNA阵列仅绘制了588个基因，并使用小鼠GEM CDNA阵列(Kurabo)进行了进一步的研究，其中绘制了8700个克隆。挑选了几个有趣的基因，功能分析正在进行中。

项目成果

N.Kawashima, et al.: "-Eraluafion of Geue oxpressia. drofile of movel osteoblastic cell livemkuxcells An in vitvo madel of ferminal diferentiation of mineralization"Bone. (in submission).

N.Kawashima 等人：“-Geue oxpressia 的消除。移动成骨细胞 livemkuxcells 的活体成骨细胞矿化分化的体内研究”骨。

N.Kawashima et al.: "An inciter model of terminal differentiation of mineralization -Evaluation of gene expression profile of novel osteoblastic cell line, Kusa cells"Bone. (in submission).

N.Kawashima 等人：“矿化终末分化的激发器模型 - 新型成骨细胞系 Kusa 细胞的基因表达谱的评估”Bone。

Nobuyuki Kawashima et al.: "An in vitro model of terminal differentiation of mineralization - Evaluation of gene expression profile of novel ostoblastic cell line, kusa cells"Bone. (in submission). (2001)

Nobuyuki Kawashima 等人：“矿化终末分化的体外模型 - 新型成骨细胞系 kusa 细胞基因表达谱的评估”骨。