Cell Biological Responses of Human Periodontium Derived Cells by Mechanical Stress

机械应力对人牙周组织来源细胞的细胞生物学反应

基本信息

批准号：
08835022
负责人：
KAWASE Toshio
金额：
$ 1.6万
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 (C)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

项目摘要

Periodontal ligament lies between tooth and alveolar bone and functions as a cushion mitigating mechanical stress such as occlusion and mastication, and the mechanical stress may affect the periodontal metabolism. The object of this study was to demonstrate the effect the mechanical stretching on the level of ^<35>S-Met labeled proteins and the synthesis and the gene expression of osteonectin (ON) in human periodontal ligament-derived fibroblasts (HPLF). HPLF were cultured on Flex I,type I collagen coated flexible-bottomed culture plates on a Flexercell strain unit. HPLF were flexed at 3 cycles (10 sec strain and 10 sec relaxation) at 3 (7,18 and 24% increase in surface area) for 1,3 and 5 days).After macromolecules in media labeled with ^<35>S-Met applied to IEC-DEAE column eluted with 0-0.5M NaCl in PBS (-), SDS-PAGE (5-15%gel) were performed and autoradiograms were obtained. The application of stretching increased of several radiolabeled bands between 200 to 100 and 70 and 30 kDa compared with the non-stress control were shifted to the neutral fraction of of ion-exchange chromatography. The shift of elution position suggested that there may be two possibilities which are the decrease of posttranslational modification such as phosphorylation and acidic glycosylation or enzymatic degradation of modifications.We have focused to ON which has established as one of bone non-collagenous proteins regulating bone mineralization and remodeling. The synthesis of ON subjected to mechanical stress, as examined by Western blotting analysis were increased in HPLF by 18% for the first day, but results were not significantly different as compared to the synthesis of ON to mechanical stress of 7 and 24%. These findings suggest that ON may be involved in periodontal metabolism in response to the mechanical stress.

牙周韧带位于牙齿和牙槽骨之间，起到缓冲作用，减轻咬合、咀嚼等机械应力，机械应力会影响牙周代谢。本研究的目的是证明机械拉伸对人牙周膜来源的成纤维细胞(HPLF)中^ 35 S-Met标记蛋白的水平以及骨连接蛋白(ON)的合成和基因表达的影响。 HPLF 在 Flexercell 菌株单元上的 Flex I、I 型胶原涂层软底培养板上培养。 HPLF在3个循环(10秒应变和10秒松弛)下弯曲(表面积增加7,18和24％)，持续1,3和5天)。在用^ 35 S-标记的介质中的大分子之后将 Met 应用于 IEC-DEAE 柱，用 0-0.5M NaCl 的 PBS (-) 溶液洗脱，进行 SDS-PAGE (5-15%gel) 并获得放射自显影图。与非应激对照相比，拉伸的应用增加了200至100和70和30 kDa之间的几个放射性标记带，这些带被转移至离子交换色谱的中性部分。洗脱位置的变化表明可能存在两种可能性，即磷酸化和酸性糖基化等翻译后修饰的减少或修饰的酶促降解。我们重点关注ON，它已被确定为调节骨矿化的骨非胶原蛋白之一和改造。通过蛋白质印迹分析检测，在 HPLF 中，受到机械应力的 ON 合成第一天增加了 18%，但与机械应力下 ON 的合成分别为 7% 和 24% 相比，结果没有显着差异。这些发现表明，ON 可能参与牙周代谢以响应机械应力。