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目的探讨高张应变调控的肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)对内皮源性微体(endothelial microparticles,EMPs)数量与表面细胞间黏附分子-1(intercellular cell adhesion molecule-1,ICAM-1)表达的作用。方法采用Flexercell细胞张应变加载系统对大鼠胸主动脉内皮细胞(endothelial cells,ECs)分别施加5%(模拟正常生理状态)和18%(模拟高血压状态)幅度的周期性张应变,加载频率均为1.25 Hz,加载持续时间为24 h,实时PCR检测不同幅度张应变条件下ECs的TNF-αmRNA表达水平。之后应用TNF-α刺激大鼠胸主动脉ECs,收集上清液,超速离心提取得到内皮源性微体(endothelial microparticles,EMPs);用亲脂性苯乙烯基(lipophilic styryl)以及透射电镜对EMPs进行形态鉴定;流式细胞术对TNF-α刺激产生的Annexin V阳性EMPs进行计数,并检测EMPs表面ICAM-1的表达。结果与5%正常张应变组相比,18%高张应变条件下ECs的TNF-α表达水平显著上升。TNF-α能够显著上调ECs产生Annexin V阳性的EMPs数量,且TNF-α刺激ECs产生的EMPs表面ICAM-1表达量显著增加。结论高张应变条件下ECs高表达TNF-α可能介导了EMPs产生和表面ICAM-1高表达。研究结果为后续探讨EMPs在血管重建力学生物学机制中的作用提供新的实验证据。
Objective To investigate the effect of hyper tensioned strain of tumor necrosis factor-α (TNF-α) on the number of endothelial microparticles (EMPs) and the expression of intercellular cell adhesion molecule- 1, ICAM-1) expression. Methods Cyclic tensile strain of 5% (simulated normal physiological state) and 18% (simulated hypertensive state) amplitude were applied to rat aortic endothelial cells (ECs) using Flexercell cell strain loading system. The loading frequency Were 1.25 Hz and the loading duration was 24 h. Real-time PCR was used to detect the expression of TNF-αmRNA in ECs under different tensile strain. After that, the ECs of rat thoracic aorta were stimulated by TNF-α. The supernatants were collected and subjected to ultracentrifugation to obtain endothelial microparticles (EMPs). EMPs were treated with lipophilic styryl and TEM Morphological identification; Annexin V positive EMPs produced by TNF-α stimulation were counted by flow cytometry and the expression of ICAM-1 on the surface of EMPs was detected. Results Compared with 5% normal tensile strain, the expression of TNF-α in ECs increased significantly under 18% high strain. TNF-α significantly up-regulated the number of Annexin V-positive EMPs produced by ECs, and the expression of ICAM-1 on EMPs stimulated by TNF-α was significantly increased. Conclusion High expression of TNF-α in ECs under high-strain condition may mediate the production of EMPs and the high expression of ICAM-1 on the surface. The results provide new experimental evidence for the follow-up study of the role of EMPs in the biological mechanism of vascular remodeling.