中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (46): 8609-8612.doi: 10.3969/j.issn.1673-8225.2011.46.014

• 血管组织构建 vascular tissue construction • 上一篇    下一篇

人脐静脉内皮细胞单层通透性改变的效应因子

闫承慧，于海波，张效林，康  建，韩雅玲   

  1. 解放军沈阳军区总医院全军心血管病研究所心内科，辽宁省沈阳市  110016
  • 收稿日期:2011-05-08 修回日期:2011-08-11 出版日期:2011-11-12 发布日期:2011-11-12
  • 通讯作者: 韩雅玲，博士，教授，博士生导师，解放军沈阳军区总医院全军心血管病研究所心内科，辽宁省沈阳市 110016 hanyaling@263.net
  • 作者简介:闫承慧☆，女，1973年生，黑龙江省哈尔滨市人，汉族，2003年哈尔滨医科大学毕业，博士，副主任技师，主要从事心血管病基础研究。 yanch1029@163.com
  • 基金资助:

    国家自然科学基金面上项目(30770793, 30971218，81070097)；国家自然科学基金青年基金项目(30800465)资助；辽宁省自然科学基金(20092088)和辽宁省科技攻关项目(2009225009-9)资助。

Effective factors for regulating monolayer permeability of human umbilical vein endothelial cells

Yan Cheng hui, Yu Hai-bo, Zhang Xiao-lin, Kang Jian, Han Ya-ling   

  1. Department of Cardiology, Institute of Cardiovascular Diseases, General Hospital of Shenyang Military Area Command of Chinese PLA, Shenyang  110016, Liaoning Province, China
  • Received:2011-05-08 Revised:2011-08-11 Online:2011-11-12 Published:2011-11-12
  • Contact: Han Ya-ling, Doctor, Professor, Doctoral supervisor, Department of Cardiology, Institute of Cardiovascular Diseases, General Hospital of Shenyang Military Area Command of Chinese PLA, Shenyang 110016, Liaoning Province, China hanyaling@263.net
  • About author:Yan Cheng-hui☆, Doctor, Associate chief technician, Department of Cardiology, Institute of Cardiovascular Diseases, General Hospital of Shenyang Military Area Command of Chinese PLA, Shenyang 110016, Liaoning Province, China yanch1029@163.com
  • Supported by:

    the National Natural Science Foundation of China (General Program), No. 30770793*, 30971218*，81070097*; the National Natural Science Foundation of China (Youth Foundation Program), No. 30800465*; the Natural Science Foundation of Liaoning Province, No. 20092088*; High-Tech Science and Technology Program of Liaoning Province, No. 2009225009-9*

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373

被引次数

9

摘要：

背景：外源性刺激引起血管屏障功能损伤的分子机制是血管病理生理学尚未阐明的热点问题之一。
目的：探讨炎症递质脂多糖诱导的人脐静脉内皮细胞单层通透性改变的效应分子，寻找有效治疗靶点。
方法：应用脂多糖刺激并观察人脐静脉内皮细胞骨架蛋白F-actin和细胞单层通透性的改变。应用荧光免疫组化和Western blot方法检测脂多糖刺激前后细胞中RhoA和SRF等信号分子的改变。并通过阻断实验证实RhoA-SRF信号通路的作用。
结果与结论：100 µg/L脂多糖刺激6 h可引起人脐静脉内皮细胞中F-actin快速重构并形成大量应力纤维，细胞单层通透性明显增强。细胞中活化RhoA的表达明显增加，SRF发生明显的入核转位现象。应用特异性分子抑制剂Y27632抑制RhoA的活化后，细胞中F-actin重构现象消失，细胞单层通透性增加也受到明显抑制，SRF蛋白发生明显的出现转位。而应用Latrunculin B抑制脂多糖刺激的人脐静脉内皮细胞中F-actin应力纤维形成，对抗通透性增加，但RhoA活化未受到干扰，SRF入核现象则受到抑制。提示RhoA-SRF通路的活化介导了脂多糖诱导的人脐静脉内皮细胞中F-actin重构和内皮单层通透性增加，特异性抑制F-actin也可以阻断脂多糖引起的血管内皮单层通透性增加，同时反馈抑制SRF的入核活化现象。

关键词: 人脐静脉内皮细胞, 通透性, 细胞骨架蛋白, RhoA, ERK

Abstract:

BACKGROUND: The molecular mechanism underlying exogenous stimulation-caused vascular barrier dysfunction remains poorly understood in the field of vascular pathophysiology.
OBJECTIVE: To investigate the effective factors for regulating monolayer permeability of human umbilical vein endothelial cells and search for the effective treatment target.
METHODS: Human umbilical vein endothelial cells (HUVECs) were exposed to lipopolysaccharide (LPS) with and without specific inhibitors of RhoA and F-actin stress fibre. Changes in RhoA/SRF/F-actin activity in association with cell permeability and F-actin rearrangement signaling were investigated by immunostaining, transwell assay and western blot analysis.
RESULTS AND CONCLUSIONS: When HUVECs were treated with LPS, F-actin cytoskeleton rearrangement was detected by rhodamine-phalloidin staining in a dose and time-dependent manner, meanwhile the HUVEC hyperpermeability was investigated. Furthermore, the RhoA activity and SRF translocalization into nuclei were identified to be associated with elevation of permeability in HUVECs. The effects were blocked when cells were pretreated with Y27632 or Latrunculin B, respectively. Moreover, Latrunculin B added also inhibited the translocalization of SRF into the nuclei mediated by RhoA activity. Our results have identified the RhoA/SRF and their substrates F-actin rearrangement mediated LPS-induced changes in HUVECs permeability.

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