持续及周期流体静压力对大鼠BMSCs增殖及凋亡的影响

Effects of continuous hydrostatic pressure and cyclic hydrostatic pressure to proliferation and apoptosis of rats' BMSCs

目的:通过观察持续及周期静压力刺激对骨髓间充质干细胞(BMSCs)体外增殖和凋亡的影响,以期为软骨组织工程研究中力学加载方式的选择提供实验依据。方法:全骨髓贴壁分离法分离大鼠BMSCs,经表面标志物检测后,分为对照组(control)、持续流体静压力45Kpa组(45Kpa)、周期流体静压力45Kpa组(0～45Kpa)、持续流体静压力90Kpa组(90Kpa)及周期流体静压力90Kpa组(0～90Kpa),压力加载时间1h,连续加压2天,周期流体静压力的频率为0.1Hz。流式细胞术检测细胞周期和凋亡,激光共聚焦显微镜下观察细胞骨架。结果:90Kpa及0～90Kpa流体静压力作用下细胞增殖指数显著高于对照组(P<0.05);BMSCs加载45Kpa、0～45Kpa、90Kpa及0～90Kpa流体静压力后,细胞骨架光密度值显著高于对照组(P<0.05);90Kpa及0～90Kpa流体静压力可导致细胞凋亡(P<0.05),其中90Kpa持续流体静压力中细胞凋亡率与对照组相比差异极显著(P<0.01)。结论:高强度周期流体静压力可促进细胞增殖,而高强度持续压力则导致细胞凋亡,低强度周期压力可促使细胞骨架重组。

Objective To investigate the effects of continuous hydrostatic pressure and cyclic hydrostatic pressure on proliferation and apoptosis of bone marrow mesenchymal stem cells （BMSCs） and further to provides experimental evidence for the selection of the mechanical load for cartilage tissue engineering. Methods High purity of BMSCs was obtained from SD rats by the whole bone marrow adherent separation culture method. Detecting cell surface markers.Then they were divided into control group,continuous hydrostatic pressure group of 45kpa （45kpa）,continuous hydrostatic pressure group of 90kpa （90kpa）, cyclic hydrostatic pressure group of 45kpa （0-45kpa）, and cyclic hydrostatic pressure group of 90 kpa （0-90kpa）.The above 5 groups of BMSCs were treated with pressure l h/d for consecutive 2 days. The frequency of cyclic hydrostatic pressure groups set to 0.1Hz.The apoptosis and cell cycle was determined by flow cytometry.Cell cytoskeleton was observe by Laser confocal microscope. Results Cell proliferation index of continuous hydrostatic pressure group and cyclic hydrostatic pressure group of 90kpa were significantly higher than the control group （P〈0.05）,the mean optical density value of the continuous hydrostatic pressure group （45Kpa,90kpa）and cyclic hydrostatic pressure group （0N45KPa, 0N90kpa）were higher than the control group （P〈 0.05）, the apoptosis rate of the 90kpa, both of continuous hydrostatic pressure group and cyclic hydrostatic pressure group were higher than the control group （P〈0.05）,the mean optical density value of the continuous hydrostatic pressure group （90kpa） was extremely significant difference compared with control group （P 〈0.01）. Conclusion High strength cycle hydrostatic pressure can promote cell proliferation,While high strength continuous pressure lead cells to apoptosis,and low intensity cycle stress can induce cytoskeleton recombined.