Oxidant stress inhibits the store-dependent Ca(2+)-influx pathway of vascular endothelial cells.
Oxidant stress inhibits the store-dependent Ca(2+)-influx pathway of vascular endothelial cells.
复制标题
氧化应激抑制血管内皮细胞的储存依赖性Ca(2)流入途径。
DOI：
10.1042/bj2920385
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Doan,TN
中科院分区：
文献类型：
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作者：
Elliott,SJ;Doan,TN
叔丁基过氧化氢（t-BuOOH）诱导的氧化应激抑制血管内皮细胞中缓激肽刺激的Ca 2+信号传导。通过向悬浮在无Ca（2+）/EGTA缓冲液中的负载Fura-2的细胞中加入Ca（2+）释放剂来测定t-BuOOH对细胞内Ca（2+）池的影响。在对照细胞中，依次添加缓激肽和离子霉素产生类似的增加胞浆游离[Ca 2 +]（[Ca 2 +]i）。相比之下，与t-BuOOH孵育逐步降低的[Ca 2 +]i缓激肽的反应，并增加离子霉素，这表明总（离子霉素释放）的Ca 2+池保持充足的氧化应激。通过2，5-二叔丁基对苯二酚（BHQ）刺激的[Ca ~（2+）]i或~（45）Ca ~（2+）外排的增加来测量t-BuOOH对InsP_3敏感性Ca ~（2+）库的影响。用t-BuOOH孵育不抑制BHQ刺激的[Ca 2 +]i或45 Ca 2+流出的增加，表明InsP 3敏感的Ca 2+池保持充足和可释放。通过向悬浮在无Ca（2+）/EGTA缓冲液中的BHQ刺激的细胞中再加入Ca（2+）和通过BHQ刺激的45 Ca（2+）摄取来测定由内部Ca（2+）储存的释放刺激的Ca（2+）内流途径的活性。用t-BuOOH孵育细胞1小时显著抑制内流途径。在随后的时间点，t-BuOOH增加了基础[Ca 2 +]i，并增强了[Ca 2 +]i对BHQ的响应。用毒胡萝卜素证明了类似的结果。总之，这些发现表明：（1）t-BuOOH对缓激肽刺激的Ca 2+从内部钙库释放的抑制作用与这些钙库的耗竭无关，（2）钙库依赖性Ca（2+）内流途径的抑制是通过内流途径的直接作用或通过抑制连接内部钙库与质膜Ca（2+）内流的机制而发生的。
Oxidant stress induced by t-butyl hydroperoxide (t-BuOOH) inhibits bradykinin-stimulated Ca2+ signalling in vascular endothelial cells. The effect of t-BuOOH on intracellular Ca2+ pools was determined by addition of Ca(2+)-releasing agents to fura-2-loaded cells suspended in Ca(2+)-free/EGTA buffer. In control cells, sequential additions of bradykinin and ionomycin produced similar increases in cytosolic free [Ca2+] ([Ca2+]i). By contrast, incubation with t-BuOOH progressively decreased the response of [Ca2+]i to bradykinin and increased that to ionomycin, suggesting that the total (ionomycin-releasable) Ca2+ pool remains replete during oxidant stress. The effect of t-BuOOH on the InsP3-sensitive Ca2+ pool was measured by the increase in [Ca2+]i or efflux of 45Ca2+ stimulated by 2,5-di-t-butylhydroquinone (BHQ). Incubation with t-BuOOH did not inhibit BHQ-stimulated increases in [Ca2+]i or 45Ca2+ efflux, suggesting that the InsP3-sensitive Ca2+ pool remains replete and releasable. Activity of the Ca(2+)-influx pathway stimulated by release of internal Ca2+ stores was determined via re-addition of Ca2+ to BHQ-stimulated cells suspended in Ca(2+)-free/EGTA buffer and via BHQ-stimulated 45Ca2+ uptake. Incubation of cells with t-BuOOH for 1 h significantly inhibited the influx pathway. At later time points, t-BuOOH increased basal [Ca2+]i and potentiated the response of [Ca2+]i to BHQ. Similar results were demonstrated with thapsigargin. Together, these findings suggest that (1) the inhibitory effect of t-BuOOH on bradykinin-stimulated release of Ca2+ from internal stores is not related to depletion of these stores, and (2) inhibition of the store-dependent Ca(2+)-influx pathway occurs by a direct effect of the influx pathway or by inhibition of the mechanism which links the internal Ca2+ store to plasmalemmal Ca2+ influx.