TNFα、AII与mmLDL致血管内皮细胞膜钙激活钾通道异常
摘要
血管内皮细胞能合成合释放多种血管活性物质，对维持血管的舒缩，抑制血小板的聚集及平滑肌的增殖均具有重要意义。内皮细胞功能障碍是许多严重心血管病如高血压、动脉粥样硬化等发病的共同环节和关键，而且内皮细胞也是大多数心血管病因子的主要作用部位。认识致病因子攻击内皮细胞的早期反应的种种机制至关重要。
肿瘤坏死因子-α(TNFα)、血管紧张素Ⅱ(AⅡ)与轻度氧化低密度蛋白(mmLDL)是动脉粥样硬化和高血压等主要心血管病的重要致病因子。它们攻击内皮细胞以致损害的一个重要环节是使胞内钙浓度升高。胞内钙的升高依赖外钙内流和胞内钙库释放。促进外钙内流的电化学驱动力则受控于膜电位。所以，参与膜电位形成和调控膜电位变化的钾通道，尤其是电导颇大的钙激活钾通道(KCa)在维持膜的稳定性和调节胞内钙浓度，影响内皮细胞功能方面的作用甚为重要。由此，对内皮细胞膜上KCa的动力学特征及其在致病因子作用下的改变进行探讨从而认识受害的内皮细胞在疾病早期反应中的离子通道机制，以寻找更有效的早期的防治措施实属首要。然而，目前关于血管内皮细胞离子通道的研究鲜见报道，上述致病因子作用下的报告更加缺乏。为此，本文设计实验，以人脐静脉内皮细胞株ECV304为研究对象，应用膜片箝技术探讨以上三个致病因子对大电导钙激活钾通道(BKCa)活动的影响，以期逐步阐明内皮细胞受损早期离子通道机制。
结果发现：
1、ECV304细胞膜存在BKCa，其电导为202.54±16.62pS，反转电位在0mV左右。
2、200U／ml TNFα可强烈而迅速地激活BKCa，使其电流幅值增大，开放时延长，关闭时缩短，开放概率增加。激活G蛋白可加强TNFα的增强效应。
Introductionratherosclerosis and hypertensin are associated with adherence of blood cell, thrombosis, vasoconstriction and proliferation of vascular smooth muscle. A lot of cardiovascular risk factors act on vascular endothelium leading to endothelial cells dysfunction which induced the above pathological changes through different mechanisms.
Stimulation of endothelial cells with tumor necrosis factor-a(TNFa), generates two types of signals rone that initiates programmed cell death, and the other one leads to activation of the transcription factor NF-kB and subsequently to the inflammatory response. Endothelial cells are exposed to angiontensinII(AII), which also induces apoptosis. In addition,AII activates NOS and increases the synthesis of NO. The interaction between NO and superoxide generates peroxynitrite. The latter is well known as a mediator of NO toxicity. Minimally modified low density lipoprotein(mmLDL)plays a significant role in early stage of atherogenesis. It promots adherence of monocyte to endothelial and produce foma cells.
The increasing of cytoplasmic calcium concentration is companied by endothelial cell dysfunction. Cytoplamic calcium elavation depends on calcium influx from extracellular space and intracellular calcium releasing from calcium store. Electrochemical gradient for calcium influx is affected by membrane potential. So the postassium channel, forming and regulating the membrane potential, especially maxi-conductance calcium activated potassium channel is very important in the regulation of
肿瘤坏死因子-α(TNFα)、血管紧张素Ⅱ(AⅡ)与轻度氧化低密度蛋白(mmLDL)是动脉粥样硬化和高血压等主要心血管病的重要致病因子。它们攻击内皮细胞以致损害的一个重要环节是使胞内钙浓度升高。胞内钙的升高依赖外钙内流和胞内钙库释放。促进外钙内流的电化学驱动力则受控于膜电位。所以，参与膜电位形成和调控膜电位变化的钾通道，尤其是电导颇大的钙激活钾通道(KCa)在维持膜的稳定性和调节胞内钙浓度，影响内皮细胞功能方面的作用甚为重要。由此，对内皮细胞膜上KCa的动力学特征及其在致病因子作用下的改变进行探讨从而认识受害的内皮细胞在疾病早期反应中的离子通道机制，以寻找更有效的早期的防治措施实属首要。然而，目前关于血管内皮细胞离子通道的研究鲜见报道，上述致病因子作用下的报告更加缺乏。为此，本文设计实验，以人脐静脉内皮细胞株ECV304为研究对象，应用膜片箝技术探讨以上三个致病因子对大电导钙激活钾通道(BKCa)活动的影响，以期逐步阐明内皮细胞受损早期离子通道机制。
结果发现：
1、ECV304细胞膜存在BKCa，其电导为202.54±16.62pS，反转电位在0mV左右。
2、200U／ml TNFα可强烈而迅速地激活BKCa，使其电流幅值增大，开放时延长，关闭时缩短，开放概率增加。激活G蛋白可加强TNFα的增强效应。
Introductionratherosclerosis and hypertensin are associated with adherence of blood cell, thrombosis, vasoconstriction and proliferation of vascular smooth muscle. A lot of cardiovascular risk factors act on vascular endothelium leading to endothelial cells dysfunction which induced the above pathological changes through different mechanisms.
Stimulation of endothelial cells with tumor necrosis factor-a(TNFa), generates two types of signals rone that initiates programmed cell death, and the other one leads to activation of the transcription factor NF-kB and subsequently to the inflammatory response. Endothelial cells are exposed to angiontensinII(AII), which also induces apoptosis. In addition,AII activates NOS and increases the synthesis of NO. The interaction between NO and superoxide generates peroxynitrite. The latter is well known as a mediator of NO toxicity. Minimally modified low density lipoprotein(mmLDL)plays a significant role in early stage of atherogenesis. It promots adherence of monocyte to endothelial and produce foma cells.
The increasing of cytoplasmic calcium concentration is companied by endothelial cell dysfunction. Cytoplamic calcium elavation depends on calcium influx from extracellular space and intracellular calcium releasing from calcium store. Electrochemical gradient for calcium influx is affected by membrane potential. So the postassium channel, forming and regulating the membrane potential, especially maxi-conductance calcium activated potassium channel is very important in the regulation of
引文
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7 郑永芳，张翠华，王晓芳．L-NNA和还原型Hb对SHR_(SP)肠系膜动脉内皮细胞依赖性舒张的影响．生理学报 1994，46 (6)：568-574
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2 Drexler H, Hornig B. Endothelial dysfunction in human disease. J Mol Cell Cardiol 1999, 31: 51-60
3 Shimokawa H. Primary endothelial dysfunction: atherosclerosis. J Mol Cell Cardiol 1999, 31: 23-37
4 Mombouli JV, Vanhoutte PM. Endothelial dysfunction: from physiology to therapy J Mol Cell Cardiol 1999, 31: 61-74
5 Boulanger CM. Secondary endothelial dysfunction: hypertension and heart failure J Mol Cell Cardiol 1999, 31: 39-49
6 张翠华，郑永芳，王晓芳．美蓝与消炎痛对自发性高血压大鼠内脏血管内皮依赖性舒张减弱的影响．基础医学与临床 1994，14 (4)：42-45
7 郑永芳，张翠华，王晓芳．L-NNA和还原型Hb对SHR_(SP)肠系膜动脉内皮细胞依赖性舒张的影响．生理学报 1994，46 (6)：568-574
8 Camussi G, Alaano E, Tetta C et al. The molecular action of tumor necrosis factor-α Eur J Biochem 1991, 202: 3-14
9 Modur V, Zimmerman GA, Prescott SM et al. Endothelial cell cell inflammatory responses to tumor necrosis factor-α. J Biochem 1996, 271(22): 13094-13102
10 Peiretti F, Alessi MC, Henry M. Intracellular calcium mobilization suppresses the TNFα stimulated synthesis of PA-1 in human endothelial cells. Arterioscler Thromb Vasc Biol 1997, 17: 1550-1560
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