Upregulation of Na+ and Ca2+ transporters in arterial smooth muscle from ouabain-induced hypertensive rats.
Upregulation of Na+ and Ca2+ transporters in arterial smooth muscle from ouabain-induced hypertensive rats.
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哇巴因诱导的高血压大鼠动脉平滑肌中 Na 和 Ca2 转运蛋白的上调。
DOI：
10.1152/ajpheart.00784.2009
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发表时间：
2010
期刊：
影响因子：
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通讯作者：
Golovina,VeraA
中科院分区：
文献类型：
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作者：
Pulina,MariaV;Zulian,Alessandra;Berra-Romani,Roberto;Beskina,Olga;Mazzocco-Spezzia,Amparo;Baryshnikov,SergeyG;Papparella,Italia;Hamlyn,JohnM;Blaustein,MordecaiP;Golovina,VeraA
长期给药（25 μg·kg−1·day−1，持续5周）可诱导大鼠出现“沃阿因高血压”（OH），但沃阿因升高血压的分子机制尚不清楚。在这里，我们比较了正常血压大鼠（NT）和高血压大鼠的肠系膜动脉平滑肌细胞（ASMCs）中的Ca2+信号。静息胞质游离Ca2+浓度（[Ca2+]cyt；用fura-2测量）和苯肾上腺素诱导的Ca2+瞬态在新解离的OH asmc中增加。免疫印迹结果显示，OH动脉中Na+/Ca2+交换器-1 （NCX1，比NT增加6倍）和Na+/Ca2+交换器-1的表达均增加（与NT相比增加2.5倍），而不是主要的耐瓦巴因α -1亚基。Ca2+的进入，由肌浆网（SR） Ca2+储存消耗与环皮唑酸（SR Ca2+- atp酶抑制剂）或咖啡因激活，在OH ASMCs中增加。这反映了c型瞬时受体电位TRPC1在OH ASMCs中的表达增加了2.5倍，TRPC1是储存操作通道（soc）的重要组成部分；一些soc的另外两种成分不表达（TRPC4）或不上调（TRPC5）。由二酰基甘油类似物1-油基-2-乙酰基-sn-甘油激活的Ba2+进入[受体操作通道（ROC）活性的测量]在OH中比NT ASMCs大得多。这与TRPC6蛋白（ROC家族成员）的六倍上调相关。重要的是，在原代培养的正常大鼠肠系膜ASMCs中，100 nM沃巴因处理72小时可显著增强NCX1和TRPC6蛋白表达，并增加静息[Ca2+]细胞和ROC活性。有机碳活性也有所增加。NCX1沉默RNA敲低可显著下调TRPC6，消除瓦他因诱导的增强；TRPC6沉默RNA敲除不影响NCX1的表达，但大大减弱了沃巴因对NCX1表达的上调。显然，NCX1和TRPC6的表达是相互关联的。因此，在体外和体内，长时间的乌巴因治疗上调了动脉肌细胞中Na+泵α2-亚单位- ncx1 - trpc6 (ROC) Ca2+信号通路。这可以解释OH大鼠肌源性反应增强、苯肾上腺素诱导的血管收缩增强以及高血压。
Prolonged ouabain administration (25 μg·kg−1·day−1for 5 wk) induces “ouabain hypertension” (OH) in rats, but the molecular mechanisms by which ouabain elevates blood pressure are unknown. Here, we compared Ca2+signaling in mesenteric artery smooth muscle cells (ASMCs) from normotensive (NT) and OH rats. Resting cytosolic free Ca2+concentration ([Ca2+]cyt; measured with fura-2) and phenylephrine-induced Ca2+transients were augmented in freshly dissociated OH ASMCs. Immunoblots revealed that the expression of the ouabain-sensitive α2-subunit of Na+pumps, but not the predominant, ouabain-resistant α1-subunit, was increased (2.5-fold vs. NT ASMCs) as was Na+/Ca2+exchanger-1 (NCX1; 6-fold vs. NT) in OH arteries. Ca2+entry, activated by sarcoplasmic reticulum (SR) Ca2+store depletion with cyclopiazonic acid (SR Ca2+-ATPase inhibitor) or caffeine, was augmented in OH ASMCs. This reflected an augmented expression of 2.5-fold in OH ASMCs of C-type transient receptor potential TRPC1, an essential component of store-operated channels (SOCs); two other components of some SOCs were not expressed (TRPC4) or were not upregulated (TRPC5). Ba2+entry activated by the diacylglycerol analog 1-oleoyl-2-acetyl-sn-glycerol [a measure of receptor-operated channel (ROC) activity] was much greater in OH than NT ASMCs. This correlated with a sixfold upregulation of TRPC6 protein, a ROC family member. Importantly, in primary cultured mesenteric ASMCs from normal rats, 72-h treatment with 100 nM ouabain significantly augmented NCX1 and TRPC6 protein expression and increased resting [Ca2+]cytand ROC activity. SOC activity was also increased. Silencer RNA knockdown of NCX1 markedly downregulated TRPC6 and eliminated the ouabain-induced augmentation; silencer RNA knockdown of TRPC6 did not affect NCX1 expression but greatly attenuated its upregulation by ouabain. Clearly, NCX1 and TRPC6 expression are interrelated. Thus, prolonged ouabain treatment upregulates the Na+pump α2-subunit-NCX1-TRPC6 (ROC) Ca2+signaling pathway in arterial myocytes in vitro as well as in vivo. This may explain the augmented myogenic responses and enhanced phenylephrine-induced vasoconstriction in OH arteries as well as the high blood pressure in OH rats.