Interactions between cell adhesion and siganaling molecules in mechanical stress response of chondrocytes

软骨细胞机械应激反应中细胞粘附和信号分子之间的相互作用

基本信息

批准号：
12557180
负责人：
MITANI Hideo
金额：
$ 8.45万
依托单位：
TOHOKU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
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/en/grant/KAKENHI-PROJECT-12557180/
关键词：
chondrocytes mechanica lstress cell adhesion integrin extracellular matrix type II collagen focal adhesion kinase fibronectin Focal Adehesion kinase 細胞-基質間接着細胞内情報伝達

项目摘要

Biomechanical forces are major epigenetic factors that determine the form and differentiation of skeletal tissues, and may be transduced by cell adhesion to the extracellular matrix (ECM). To test the hypothesis that stepwise tension forces are transduced into molecular signals during early chondrogenesis, we developed a culture system studying the proliferation and differentiation of chondrocytes. Rat embryonic day-12 limb buds were microdissected and dissociated into cells which were then micro-mass cultured on a silicone membrane and maintained for up to 6 days. Stepwise tension was applied to these cultures from day 3. The time course of the expression pattern and level of cartilage-specific markers and non-chondrogenic markers were analyzed and compared with those in non-stretched control cultures. Under tension conditions, Alcian blue staining showed an apparent decrease in the rate and extent of chondrogenesis, and confocal laser scanning microscopy demonstrated apparent stretching of the cells. Quantitatively, type II collagen and aggrecan expression were significantly inhibited by 20 to 30% and 10 to 20%, respectively, after 12 hrs of tension force loading, and this difference was maintained through 3 days of culture. In contrast, the expression of type I collagen and fibronectin remained relatively constant throughout the experimental period. This down-regulation in the expression of chondrogenic markers was completely rescued when cell-ECM attachment was inhibited by GRGDSPK (Gly-Arg-Gly-Asp-Ser-Pro-Lys) peptide. We conclude that stepwise tension inhibits chondrogenesis through integrins in embryonic limb bud mesenchyme, and propose that signal transduction from biomechanical stimuli may be mediated by cell-ECM adhesion.

生物力学是确定骨骼组织形式和分化的主要表观遗传因素，并且可以通过细胞粘附到细胞外基质（ECM）来转导。为了检验以下假设：在早期软骨发生期间，逐步张力将其转导为分子信号，我们开发了一种研究软骨细胞增殖和分化的培养系统。将大鼠胚胎第12天的肢体芽显微分解并分离成细胞，然后将微质量培养在硅胶膜上，并保持长达6天。从第3天开始将逐步张力应用于这些培养物。分析了软骨特异性标记的表达模式和水平的时间过程，并分析了非发育性标记物，并将其与非伸展的对照培养物中的时间过程进行了比较。在张力条件下，Alcian蓝色染色显示软骨发生率和程度明显降低，共聚焦激光扫描显微镜表现出明显的细胞拉伸。定量地，II型胶原蛋白和脂肪蛋白表达分别在12小时的张力力加载后分别显着抑制20％至30％和10％至20％，并在3天的培养过程中维持这种差异。相反，在整个实验期间，I型胶原蛋白和纤连蛋白的表达保持相对恒定。当GrgDSPK（Gly-Arg-Gly-Gly-Asp-ser-Pro-Lys）抑制细胞ECM附着时，完全挽救了软骨形成标志物表达中的这种下调。我们得出的结论是，逐步张力通过胚胎肢体芽中充质抑制软骨发生，并提出可能通过细胞ECM粘附来介导的来自生物力学刺激的信号转导。