17β-雌二醇对人内皮细胞细胞内钙稳态的影响。
The effect of 17 beta-estradiol on intracellular calcium homeostasis in human endothelial cells.
机构信息
Department of Physiology & Pharmacology, Thomas J. Long School of Pharmacy and Health Sciences, 751 Brookside Rd, University of the Pacific, Stockton, CA 95211, USA.
出版信息
Eur J Pharmacol. 2010 Mar 25;630(1-3):92-9. doi: 10.1016/j.ejphar.2009.12.030. Epub 2010 Jan 4.
The cardiovascular effects of estrogen are mediated in part by augmenting the function of endothelial nitric oxide synthase. Endothelial nitric oxide synthase activity is dependent on many cofactors including Ca(2+). Hence, we investigated the effect of chronic 17 beta-estradiol treatment on the intracellular Ca(2+) concentration and endothelial nitric oxide synthase protein expression in the human endothelial cell line, EA.hy926, using spectrofluorometry and Western blot, respectively. Inhibiting the sarco(endo)plasmic reticulum Ca(2+) ATPase with thapsigargin caused an increase in the intracellular Ca(2+) concentration, which was higher in chronically 17 beta-estradiol-treated (1muM, 24h) cells loaded with Fura-2-acetoxymethyl ester compared to vehicle-treated cells, suggesting a higher endoplasmic reticulum Ca(2+) content in 17 beta-estradiol-treated cells. An enhanced Ca(2+) influx pathway in chronically 17 beta-estradiol-treated cells was also observed. In addition, 17 beta-estradiol-treated cells expressed higher levels of endothelial nitric oxide synthase protein in comparison to vehicle-treated cells. The chronic effect of 17 beta-estradiol on Ca(2+) homeostasis and endothelial nitric oxide synthase expression was attenuated with the nonselective estrogen receptor inhibitor, ICI 182,780 (10muM, 7alpha, 17beta-[9-[(4,4,5,5,5-Pentafluoropentyl)sulfinyl]nonyl] estra-1,3,5(10)-triene-3,17-diol). Furthermore, analysis of the thapsigargin-evoked Ca(2+) response in chronically 17 beta-estradiol-treated estrogen receptor alpha-knockdown cells showed no significant difference in Ca(2+) response compared to vehicle-treated estrogen receptor alpha-knockdown cells, indicating that the regulation of Ca(2+) homeostasis by 17 beta-estradiol is mediated through an estrogen receptor alpha-dependent pathway. These data revealed an estrogen receptor alpha-dependent modulation of Ca(2+) homeostasis accompanying the enhancement of endothelial nitric oxide synthase expression in 17 beta-estradiol-treated human endothelial cells.
雌激素的心血管作用部分通过增强内皮型一氧化氮合酶的功能来介导。内皮型一氧化氮合酶的活性依赖于许多辅助因子，包括 Ca(2+)。因此，我们使用荧光光谱法和 Western blot 分别研究了慢性 17β-雌二醇处理对人内皮细胞系 EA.hy926 细胞内 Ca(2+)浓度和内皮型一氧化氮合酶蛋白表达的影响。用 thapsigargin 抑制肌浆内质网 Ca(2+)ATP 酶会导致细胞内 Ca(2+)浓度增加，与用载体处理的细胞相比，用 Fura-2-乙氧甲酯负载的慢性 17β-雌二醇处理（1μM，24h）的细胞中 Ca(2+)浓度更高，这表明 17β-雌二醇处理的细胞内质网 Ca(2+)含量更高。还观察到慢性 17β-雌二醇处理的细胞中存在增强的 Ca(2+)内流途径。此外，与用载体处理的细胞相比，17β-雌二醇处理的细胞表达更高水平的内皮型一氧化氮合酶蛋白。用非选择性雌激素受体抑制剂 ICI 182,780（10μM，7α，17β-[9-[(4,4,5,5,5-五氟戊基)亚磺酰基]壬基]estra-1,3,5(10)-三烯-3,17-二醇）减弱了 17β-雌二醇对 Ca(2+)稳态和内皮型一氧化氮合酶表达的慢性作用。此外，在慢性 17β-雌二醇处理的雌激素受体α敲低细胞中分析 thapsigargin 诱发的 Ca(2+)反应，与用载体处理的雌激素受体α敲低细胞相比，Ca(2+)反应没有明显差异，表明 17β-雌二醇对 Ca(2+)稳态的调节是通过雌激素受体α依赖性途径介导的。这些数据表明，在 17β-雌二醇处理的人内皮细胞中，Ca(2+)稳态的雌激素受体α依赖性调节伴随着内皮型一氧化氮合酶表达的增强。
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