Mechanisms of enhancement of cell proliferation caused by the mechanical stress in osteoblast

为了分析细胞对机械应力的反应机制，我们检测了在不同机械应力条件下培养的小鼠成骨细胞来源的细胞生长、增殖相关基因表达和细胞粘附分子定位的变化。结果表明，重力矢量平均旋转培养的细胞S期进程略有延迟，而离心力培养的细胞S期进程加快。细胞粘附和/或细胞骨架与细胞增殖密切相关。尽管我们通过RT-PCR检测了在机械应力（60 G离心力）下培养的细胞的细胞粘附分子（整合素）的基因表达，但整合素α1、α2、α5和β1的基因表达没有改变。为了检查当在静压下培养细胞时细胞生长是否也升高，代替离心力，对接种在盖玻片上的细胞施加压力（约100 μ g/ml）。结果，成骨细胞来源的MC 3 T3-E1细胞的生长速度略有提高，但在实验组和对照组之间没有显著差异。因此，我们使用共聚焦显微镜分析了细胞粘附分子之一整合素β1和细胞骨架如微管和微丝的定位和形成的变化。而在静压力培养、离心培养和静止对照组之间无显著性差异。另一方面，在旋转条件下培养的细胞似乎略微减少了微丝的形成。

To analyze the mechanisms of cell response against the mechanical stress, we examined the changes of cell growth, proliferation-related gene expressions and localization of cell adhesion molecules when the mouse osteoblast derived cells were cultured under different mechanical stress conditions. It had revealed that the progression of S phase in cell cycle showed a little delay when they were cultured under gravity vector-averaged clinorotation and, in contrast, it was hastened when the cells were cultured under centrifugation force. It is widely accepted that cell adhesion and/or cytoskeletons are closely related to the cell proliferation. Although we examined the gene expression of cell adhesion molecules, integrins, of the cells cultured under mechanical stress (60 G of centrifugation force) by RT-PCR, the gene expressions of integrin α1, α2, α5 and β1 did not changed. To examine whether the cell growth was also elevated when they were cultured under static pressure, instead of the centrifugation force, the cells seeded on a cover slip were inflicted the pressure (ca. 100 kPa) in a syringe for 24 hr. As the result, the growth rate of osteoblast-derived MC3T3-E1 cells was slightly elevated but it was not significant difference between the experimental group and the control group. Therefore, we analyzed the changes of localization and formation of one of the cell adhesion molecules, integrin β1, and cytoskeletons, such as microtubules and microfilaments using a confocal microscope. But no significant difference was observed among the cells cultured under the static pressure, centrifuged culture and the stationaly control. On the other hand, the cells cultured under clinorotaion seemed to slightly decrease the formation of microfilament.