基础医学与临床 ›› 2011, Vol. 31 ›› Issue (8): 864-868.

• 研究论文 • 上一篇    下一篇

同型半胱氨酸抑制人脐静脉内皮细胞eNOS活性

钟华1,胡清华1,张会敏2,赵丹3,孙志萍1,李增春1,何芳1   

  1. 1. 石河子大学医学院
    2. 新乡医学院
    3. 郧阳医学院
  • 收稿日期:2010-09-10 修回日期:2010-10-14 出版日期:2011-08-05 发布日期:2011-07-13
  • 通讯作者: 何芳 E-mail:FangF2002shz@126.com
  • 基金资助:
    国家自然科学基金

Homocysteine inhibits activity of eNOS in cultured human umbilical vein endothelial cells

Hua ZHONG1,Qing-hua HU2,Hui-min ZHANG2,Dan ZHAO2,Zhi-ping SUN2,Zeng-chun LI2,Fang HE1   

  1. 1. Medical College of Shihezi University
    2.
  • Received:2010-09-10 Revised:2010-10-14 Online:2011-08-05 Published:2011-07-13
  • Contact: Fang HE E-mail:FangF2002shz@126.com

摘要: 目的 研究不同浓度同型半胱氨酸(Hcy),作用不同的时间,对人脐静脉内皮细胞中内皮型一氧化氮合酶(eNOS)活性的影响机制。方法 取2~3代人脐静脉内皮细胞(HUVECs)与不同浓度Hcy孵育以及孵育不同时间。用Western blot和实时荧光定量RT-PCR法检测热休克蛋白90(HSP90)、蛋白激酶B(Akt) 蛋白表达,小凹蛋白(Caveolin-1)、eNOS、P-eNOS蛋白及mRNA的表达,二甲基精氨酸二甲胺水解酶(DDAH)mRNA表达。结果:(1)与使静息和激活状态下的对照组相比,在Hcy作用下,Caveolin-1蛋白的表达上调(p<0.05),而Akt蛋白的表达下调(p<0.05),且均呈浓度、时间依赖性。eNOS蛋白的表达无明显改变(p﹥0.05);(2)Hcy对其他检测指标无影响。结论 Hcy引起的eNOS活性降低与增强Caveolin-1蛋白表达使其与eNOS藕联增加,减少HSP90、Akt和eNOS复合物的形成,进而抑制了eNOS的活性有关。

关键词: 同型半胱氨酸, eNOS, 人脐静脉内皮细胞, A23187

Abstract: Objective To study the mechanism of homocysteine(Hcy)at different concentrations and action times on eNOS activity in cultured human umbilical vein endothelial cells(HUVECs), both in base and calcium ionophore activated state. METHODS HUVECs were collected and cultured to the second passage.Then the second passage of HUVECs were cultured with different concentrations Hcy and in different action times. The expression of HSP90,Akt,Caveolin-1,eNOS,P-eNOS proteins were detected by western blot and the expression of Caveolin-1,eNOS,DDAH mRNA by RT-PCR.RESULTS (1) Compared with control group, both in base and calcium ionophore activated state. The expression of Caveolin-1 group increased obviously(p<0.05) in concentration and time-dependent manner but the expression of Akt showed a decrease(p<0.05). The expression of eNOS did not change significantly(p﹥0.05);(2) There was no difference in the expression of HSP90,eNOS,P-eNOS protein and mRNA among every group. DDAH mRNA among every group too..CONCLUSIONS There was no relationship between decreasing activity of eNOS induced by Hcy and eNOS expression, Hcy probably can increase the association of Caveolin-1and eNOS by enhancing the expression of Caveolin-1,and reduce the complex formation of Caveolin-1, Akt,Caveolin-1 in HUVEC both in base and activated state, which can decrease the activity of eNOS.

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