模拟失重对流体剪切应力诱导MG-63细胞ERK2激活的影响

目的 研究成骨细胞力学信号转导功能中酪氨酸磷酸结合结构域适应蛋白(Shc)和细胞外信号调节蛋白激酶(ERK2)的关系,并观察模拟失重条件下流体剪切应力(FSS)诱导ERK2活性的变化,以期从信号转导水平探讨模拟失重对成骨细胞力学信号转导功能的影响.方法 将负性缺失突变体质粒Shc-SH2和活性ERK质粒Myc-ERK2共同转染入MG-63细胞中,同时将真核表达载体pcDNA3和Myc-ERK2也共同转染入MG-63细胞中作为转染平行对照组,对转染细胞进行1.5 Pa流体剪切应力处理15 min.利用回转器模拟失重60 h,对转染细胞进行1.5 Pa FSS处理15 min.结果 在转染平行对照组中,FSS组中的Myc-ERK2的活性显著升高;在Shc-SH2转染组中,Myc-ERK2的活性与转染平行对照组相比显著降低(P＜0.05),在FSS组中未见Myc-ERK2活性的变化.转染了真核表达载体pcDNA3和Myc-ERK2的细胞在模拟失重条件培养及FSS作用15 min后,Myc-ERK2的活性升高,但与未进行模拟失重的对照组相比,Myc-ERK2的活性显著降低(P＜0.05).结论 在力学信号转导通路中,Shc可以介导力学信号以激活ERK2.模拟失重对细胞的力学信号转导功能具有不良影响.

Effects of Simulated Weightlessness on the Induction of ERK2 by Fluid Shear Stress in MG-63 Osteosarcoma Cells

Objective To understand the effect of simulated weightlessness on the osteoblasic mechano-signal transduction with the level of signal transduction,to explore the relationship between Shc and ERK2 in the mechanotransduction of MG-63 osteosarcoma cells,and to survey the effect of simulated weightlessness on the induction of flow shear stress(FSS) to the activation of ERK2 in MG-63 osteosarcoma cells. Methods The MG-63 osteosarcoma cells of one group were co-transfected with negative mutant of Shc (Shc-SH2) and Myc-ERK2. Another group cells were co-transfected with pcDNA3 parental vector together and Myc-ERK2 as control group. Cells of both groups were treated by 1.5 Pa FSS and no FSS for 15 min. The effect of FSS on the activation of Myc-ERK2 was examined. The control group cells were cultured for 60 h in 2 different gravitational conditions, 1 G and simulated weightlessness with clinostat. Then, 1.5 Pa FSS and no FSS were set to the cells of two gravitational conditions for 15 min. Results In 1 G terrestrial gravitational condition, FSS caused significantly activating Myc-ERK2 in cells transfected with pcDNA3, and this FSS induction to Myc-ERK2 activity was drastically reduced in cells transfected with Shc-SH2 (P<0.05). There was no measurable value in the amount of Myc-ERK2 activities in MG-63 cells without FSS treatment. After the MG-63 cells of control group, transfected with pcDNA3 and Myc-ERK2, were cultured in simulated weightlessness condition with clinostat for 60 h and were treated with mechanical stimulation,1.5 Pa FSS for 15 min, the activity of Myc-ERK2 was increased, but it was significantly decreased as compared with cultured in 1 G, terrestrial gravitational condition (P<0.05). Conclusion It is suggested that in the pathway of signal transduction Shc may be involved in the upstream signaling for the flow shear stress induction to activate ERK2,and simulated weightlessness condition has an adverse effect on mechanotransduction in MG-63 cells.