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，我们检测了不同机械应力条件下培养小鼠成骨细胞时细胞生长、增殖相关基因表达和细胞粘附分子定位的变化。结果表明，在重力载体平均旋转条件下，细胞周期S期的进展有一定的延迟，而在离心力条件下，细胞周期S期的进展有一定的加快。人们普遍认为细胞粘附和/或细胞骨架与细胞增殖密切相关。通过RT-PCR检测机械应力（60 G离心力）培养细胞的细胞粘附分子整合素基因表达，整合素α1、α2、α5和β1基因表达没有变化。为了检验细胞在静压下而不是在离心力下培养时是否也能提高细胞的生长，将细胞接种在盖片上，在注射器中施加压力（约100 kPa） 24小时。结果，成骨细胞来源的MC3T3-E1细胞的生长速度略有升高，但实验组与对照组之间差异不显著。因此，我们利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。但静压、离心和静对照培养的细胞无显著差异。另一方面，恒温培养的细胞微丝的形成似乎略有减少。

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.