Basic Research for Tissue Engineering of Periodontal Tissue Regeneration : Validity of Enamel Matrix Proteins and Periodontal Ligament Fibroblast Synthesized Proteins.

牙周组织再生的组织工程基础研究：牙釉质基质蛋白和牙周膜成纤维细胞合成蛋白的有效性。

基本信息

批准号：
14370714
负责人：
KAWASE Toshio
金额：
$ 8.51万
依托单位：
Kanagawa Dental College
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

Enamel matrix proteins (enamel matrix derivative, Emdogain^<【○!R】>, EMD) have been clinically used regenerate the periodontal tissues. Although it is clamed that EMD can be used to promote new attachment formation around periodontally invoved teeth, the underlying biological mechanism is not understood. We have demonstrated that the secreted proteins of HPLF have a potency for the cell attachment and spreading activity factor (CASF) promoting tissue regeneration.The present study investigated the effect of EMD on the behavior of rat bone marrow cells(RBMC) in vitro and compared it with that for the conditioned medium protein(CMP) obtained from human periodontal ligament-derived fibroblasts (HPLF), especially their attachment property, alkaline phosphatase(ALPase) activity and the mRNA levels for ALPase, bone sialoprotein(BSP), Osteopontin(OPN), and osteocalcin(OCN). The hydrophobic dishes were coated EMD (25μg/ml) or CMP (25μg/ml) for 24 hrs at 4℃. The dishes were treated with heat inactivated BSA.RBMCs were obtained from the femora of 7-week-old male rats and cultured in α-MEM containing 10% FCS and 0.25mM ascorbic acid phosphate on hydrophobic culture plates coated with EMD or CMP. RBMCs attached and spread within 3 hours. RBMCs cultured onto EMD coated dishes expressed high ALPase activity and mRNA levels when compared with CMP coated ones. Furthermore, the expression of OPN and OCN mRNAs of RBMCs was significantly enhanced by the presence of EMD. EMD did not play a chemotactic activity. CMP stimulate the migration activity of HPLF.Our results indicated that RBMCs responded differently to EMD or CMP, and that EMD stimulated osteogenic differentiation of RBMCs. Therefore, HPLF may interacted with EMD coated on dentin surface, proliferate and differentiate. And mesenchymal cells (RBMC) couled be migrated by CMP and differentiated to osteogenic cells.

牙釉质基质蛋白（牙釉质基质衍生物，Emdogain^<【○！R】>，EMD）已在临床上用于牙周组织的再生。生物学机制尚不清楚。我们已经证明 HPLF 的分泌蛋白具有促进组织再生的细胞附着和扩散活性因子 (CASF) 的效力。本研究调查了 EMD 对组织再生的影响。大鼠骨髓细胞（RBMC）的体外行为，并与人牙周膜来源的成纤维细胞（HPLF）获得的条件培养基蛋白（CMP）进行比较，特别是其附着特性、碱性磷酸酶（ALPase）活性和ALPase、骨唾液蛋白 (BSP)、骨桥蛋白 (OPN) 和骨钙素 (OCN) 的 mRNA 水平疏水培养皿涂有 EMD。 (25μg/ml)或CMP(25μg/ml)在4℃下处理24小时。用热灭活的BSA处理培养皿。从7周龄雄性大鼠的股骨中获得RBMC，并在含有10的α-MEM中培养。 % FCS 和 0.25mM 抗坏血酸磷酸盐在涂有 EMD 或 CMP 的疏水性培养板上，并在 3 小时内铺展。与 CMP 包被的培养皿相比，培养在 EMD 包被培养皿上的 RBMC 表达较高的 ALPase 活性和 mRNA 水平，此外，EMD 的存在显着增强了 RBMC 的 OPN 和 OCN mRNA 表达，但没有发挥 CMP 刺激作用。我们的研究结果表明，RBMCs 对 EMD 或 CMP 的反应不同，并且 EMD 刺激 RBMCs 的成骨分化，因此，HPLF 可能与 RBMCs 相互作用。 EMD涂覆在牙本质表面，增殖并分化，间充质细胞（RBMC）通过CMP耦合迁移并分化为成骨细胞。