体外震波对骨微血管内皮细胞生理功能的影响

Impact of extracorporeal shock waves on physiological function of bone microvascular endothelial cells

背景:体外震波作为一种特殊类型的外源性物理作用信号,通过刺激、激活内皮细胞上机械力学信号感受器,激活细胞内特殊信号传导系统,调控内皮细胞基因表达,对内皮细胞生理功能产生影响。目的:观察不同能流密度及作用次数体外震波对股骨头骨微血管内皮细胞成血管能力、迁移能力及抗凋亡作用的影响。方法:从关节置换患者股骨头获取股骨头骨松质,分离、培养、传代培养股骨头骨微血管内皮细胞。通过血管性血友病因子、CD31血管内皮细胞标记物抗体进行免疫荧光鉴定。根据体外震波不同能流密度(低0.03 m J/mm2,高0.11 mJ/mm2)及不同作用次数(400次,800次)对细胞进行分组。通过细胞3D培养实验观察内皮细胞成血管能力,划痕实验观察内皮迁移能力,流式细胞仪检测内皮细胞凋亡情况。结果与结论:(1)从股骨头松质骨中分离、培养出的细胞均能表达血管性血友病因子、CD31,表明这些细胞具有血管内皮细胞的特征,阳性率接近100%;(2)体外震波作用于股骨头骨微血管内皮细胞提高其体外成血管能力及迁移能力,低能流密度组促进成血管能力和细胞迁移能力显著优于高能流密度组,高能流密度组内随着作用次数的增加促成血管能力下降;(3)对于激素诱导股骨头骨微血管内皮细胞凋亡有保护作用;(4)结果说明,体外震波对股骨头骨微血管内皮细胞生理功能的影响与作用能流密度及作用次数相关。

BACKGROUND： Extracorporeal shock wave has been shown to influence the physiological function of endothelial cells via the activation of mechanoreceptors and specific signal transduction system, and gene expression regulation.OBJECTIVE： To explore the impact of different energy flow densities and numbers of shots of extracorporeal shock waves on the new vessel formation ability, migration capability and apoptosis of bone microvascular endothelial cells. METHODS： Bone microvascular endothelial cells isolated from the femoral head of patients undergoing arthroplasty were subcultured in vitro, and then were immunofluorescently evaluated with endothelial cell marker antibodies to CD31 and von Willebrand factor（vW F）, and grouped according to different energy flow densities（low, 0.03 m J/mm2; high, 0.11 mJ /mm2） and numbers of shots（400 and 800）. Capillary-like tube formation, migration capability and apoptosis of bone microvascular endothelial cells were determined by 3-D culture in vitro, scratch test, and flow cytometry, respectively. RESULTS AND CONCLUSION： v WF and CD31 were positively expressed in approximately 100% of bone microvascular endothelial cells, which indicates the cultured cells had characterization of microvascular endothelial cells. Extracorporeal shock wave enhanced angiogenesis and migration capability of bone microvascular endothelial cells derived from the femoral head, and especially low-energy flow density of extracorporeal shock wave exerted more superior effects. Angiogenesis of bone microvascular endothelial cells was decreased with the increased shot number in the low-energy flow density group. In addition, extracorporeal shock wave inhibited bone microvascular endothelial cell apoptosis induced by steroids. Our results suggest that energy flow density and number of shots of extracorporeal shock waves impact the physiological function of bone microvascular endothelial cells derived from the femoral head.