正常生理和尿毒症状态下剪切力对静脉内皮细胞中KLF2及eNOS表达的影响

Effect of shear stress on KLF2 and eNOS expression in vein endothelial cells under normal physiology and uremia conditions

目的研究生理及尿毒症状态下剪切力对静脉内皮细胞中,KLF2和eNOS表达的影响,探讨导致静脉内皮细胞功能紊乱的机制。方法分别在生理状态和尿毒症状态下,在平行板流动腔内模拟不同剪切力,用剪切力作用各组静脉内皮细胞4、12、24 h。采用免疫组化荧光染色技术和实时荧光定量PCR技术检测KLF2及eNOS的表达情况。结果在生理状态下,KLF2明显受剪切力作用调节,高强度剪切力和生理强度剪切力会上调KLF2的表达,而低强度剪切力和振荡剪切力会下调KLF2的表达,但随作用时间的延长,KLF2的表达也会有所上调。在尿毒症状态下,KLF2表达明显被抑制,即使再给予高剪切力作用,KLF2水平也不及正常生理状态;在低剪切力及振荡剪切力作用下,KLF2表达更明显地被抑制。KLF2主要在细胞核内表达,随着剪切力的作用,KLF2也在细胞质中表达,而eNOS主要在细胞质和细胞核内呈颗粒样表达。结论动静脉内瘘术后在尿毒症环境及振荡剪切力的多重因素作用下,KLF2和eNOS的表达受到抑制,可能是导致静脉内皮细胞功能紊乱、内膜增生、自体动静脉内瘘失功发生和发展的主要机制。

Objective To study the effect of shear stress on the expression of KLF2 and eNOS in venous endothelial cells under physiological and uremic conditions,and to explore the mechanism leading to dysfunction of venous endothelial cells.Methods Under physiological conditions and uremia conditions,different shear stresses were simulated in the parallel plate flow cavity,and the shear stresses were applied to the venous endothelial cells of each group for 4,12,and 24 hours.The expression of KLF2 and eNOS was detected by immunohistochemical fluorescent staining technique and real-time fluorescent quantitative PCR technique.Results Under physiological conditions,KLF2 is obviously regulated by shear stresses.High-intensity shear stresses and physiological shear stresses will up-regulate the expression of KLF2,while low-intensity shear stresses and oscillating shear stresses will down-regulate the expression of KLF2.As the duration of action increases,the expression of KLF2 will also increase.In the state of uremia,the expression of KLF2 is significantly inhibited.Even if high shear stresses is applied,the level of KLF2 is not high-expressed as the physiological state.And under the action of low shear stresses and oscillating shear stresses,KLF2 expression is more significantly inhibited.KLF2 is mainly expressed in the nucleus.With the action of shear stresses,KLF2 is also expressed in the cytoplasm,while eNOS is mainly expressed in granular form in the cytoplasm and nucleus.Conclusions After arteriovenous fistula operation,the expression of KLF2 and eNOS is inhibited under the action of multiple factors of uremia environment and oscillating shear stresses,which may be the main cause of the occurrence and development of venous endothelial cell dysfunction,intimal hyperplasia,and AVF failure.