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

• 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

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.

生物力学是决定骨骼组织形态和分化的主要表观遗传因素，可能通过细胞与细胞外基质(ECM)的黏附传递。为了验证在早期软骨形成过程中逐步张力转化为分子信号的假设，我们开发了一种培养系统，研究软骨细胞的增殖和分化。大鼠胚胎第12天肢芽被显微解剖分离成细胞，然后在硅胶膜上进行微团培养，最长维持6天。从第3天开始对这些培养物施加逐步拉伸，分析软骨特异标记和非软骨形成标记的表达模式和水平的时间进程，并与未拉伸的对照组培养的软骨特异性标记和非软骨形成标记的表达模式和水平进行比较。在张力条件下，Alcian蓝染色显示软骨形成的速度和程度明显降低，激光共聚焦扫描显微镜显示细胞明显拉伸。定量分析显示，张力加载12h后，II型胶原和聚集素的表达分别被显著抑制20%~30%和10%~20%，并持续到培养3天。相比之下，I型胶原和纤维连接蛋白的表达在整个实验期间保持相对稳定。当GRGDSPK(Gly-Arg-Gly-Asp-Ser-Pro-Lys)肽抑制细胞-ECM附着时，这种软骨形成标志物表达的下调被完全挽救。结论：步进式张力通过整合素抑制胚胎肢芽间充质的软骨形成，提示生物力学刺激的信号转导可能是通过细胞-细胞外基质黏附来实现的。

项目成果

K.Onodera, I.Takahashi, Y.sasano et. al.: "Mechanical tension inhibits chondrogenesis of embryonic rat limb bud cells"Journal of Dental Research. 80. 634-634 (2001)

K.Onodera，I.Takahashi，Y.sasano 等。

K.Onodera, I.Takahashi, Y.Sasano et al.: "Mechanical stretch inhibits chondrogenesis through integrins in embryonic mesenchymal cells"Journal of Dental Research. 81. A382-A382 (2002)

K.Onodera、I.Takahashi、Y.Sasano 等人：“机械拉伸通过胚胎间充质细胞中的整合素抑制软骨形成”《牙科研究杂志》。

K. Onodera, I. Takahashi, Y. Sasano et. al.: "Mechanical tension inhibits chondrogenesis of embryonic rat limb bud cells"Journal of Dental Research. 80. 634-634 (2001)

K. Onodera、I. Takahashi、Y. Sasano 等。

K. Onodera, I. Takahashi, Y. Sasano et. al.: "Mechanical stretch inhibits chondrogenesis through integrins in embryonic mesenchymal cells"Journal of Dental Research. 81. A382-A382 (2002)

K. Onodera、I. Takahashi、Y. Sasano 等。

K.Onodera, I.Takahashi, Y.Sasano et al.: "Mechanical tension inhibits chondrogenesis of embryonic rat limb bud cells"Journal of Dental Research. 80. 634-634 (2001)

K.Onodera、I.Takahashi、Y.Sasano 等人：“机械张力抑制胚胎大鼠肢芽细胞的软骨形成”牙科研究杂志。