PKCα mediates acetylcholine-induced activation of TRPV4-dependent calcium influx in endothelial cells
PKCα mediates acetylcholine-induced activation of TRPV4-dependent calcium influx in endothelial cells
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DOI：
10.1152/ajpheart.00142.2011
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发表时间：
2011-09-01
影响因子：
4.8
通讯作者：
Thodeti, Charles K.
中科院分区：
文献类型：
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作者：
Adapala, Ravi K.;Talasila, Phani K.;Thodeti, Charles K.
Adapala RK，Talasila PK，Bratz IN，Zhang DX，Suzuki M，Meszaros JG，Thodeti CK. PKC α介导乙酰胆碱诱导的内皮细胞TRPV 4依赖性钙内流的激活Am J Physiol Heart Circ Physiol 301：H757-H765，2011.首次发表于2011年6月24日; doi：10.1152/ajpheart.00142.2011。瞬时受体电位香草酸通道4（TRPV 4）是一种多模态激活的非选择性阳离子通道，参与血管舒张和高血压的调节。我们和其他人最近表明，周期性拉伸和剪切应力激活TRPV 4介导的内皮细胞（EC）中的钙内流。除了机械力，乙酰胆碱（ACh）被证明激活TRPV 4介导的钙内流内皮细胞，这是重要的一氧化氮依赖性血管舒张。然而，ACh激活TRPV 4的分子机制尚不清楚。在这里，我们表明特定的TRPV 4拮抗剂AB-159908抑制了ACh诱导的钙内流和内皮一氧化氮合酶（eNOS）磷酸化，但不抑制细胞内储存的钙释放。重要的是，在TRPV 4无效EC中，钙池操作的钙内流的激活没有改变，表明TRPV 4依赖性钙内流是通过受体操作的途径介导的。此外，我们发现ACh处理激活蛋白激酶C（PKC）α，并通过特异性抑制剂Go-6976抑制PKC α活性，或通过慢性12-O-十四烷酰佛波醇-13-乙酸酯处理表达PKC α的激酶死亡突变体，但不表达PKC β或下调PKC α表达，完全消除ACh诱导的钙内流。最后，我们发现，乙酰胆碱诱导的血管舒张抑制PKC α抑制剂Go-6976在小肠系膜动脉从野生型小鼠，但不是在TRPV 4无效的小鼠。综上所述，这些研究结果首次证明，PKC的一种特异性亚型PKC α介导内皮细胞中激动剂诱导的受体介导的TRPV 4活化。
Adapala RK, Talasila PK, Bratz IN, Zhang DX, Suzuki M, Meszaros JG, Thodeti CK. PKC alpha mediates acetylcholine-induced activation of TRPV4-dependent calcium influx in endothelial cells. Am J Physiol Heart Circ Physiol 301: H757-H765, 2011. First published June 24, 2011; doi:10.1152/ajpheart.00142.2011.-Transient receptor potential vanilloid channel 4 (TRPV4) is a polymodally activated nonselective cationic channel implicated in the regulation of vasodilation and hypertension. We and others have recently shown that cyclic stretch and shear stress activate TRPV4-mediated calcium influx in endothelial cells (EC). In addition to the mechanical forces, acetylcholine (ACh) was shown to activate TRPV4-mediated calcium influx in endothelial cells, which is important for nitric oxide-dependent vasodilation. However, the molecular mechanism through which ACh activates TRPV4 is not known. Here, we show that ACh-induced calcium influx and endothelial nitric oxide synthase (eNOS) phosphorylation but not calcium release from intracellular stores is inhibited by a specific TRPV4 antagonist, AB-159908. Importantly, activation of store-operated calcium influx was not altered in the TRPV4 null EC, suggesting that TRPV4-dependent calcium influx is mediated through a receptor-operated pathway. Furthermore, we found that ACh treatment activated protein kinase C (PKC) alpha, and inhibition of PKC alpha activity by the specific inhibitor Go-6976, or expression of a kinase-dead mutant of PKC alpha but not PKC epsilon or downregulation of PKC alpha expression by chronic 12-O-tetradecanoylphorbol-13-acetate treatment, completely abolished ACh-induced calcium influx. Finally, we found that ACh-induced vasodilation was inhibited by the PKC alpha inhibitor Go-6976 in small mesenteric arteries from wild-type mice, but not in TRPV4 null mice. Taken together, these findings demonstrate, for the first time, that a specific isoform of PKC, PKC alpha, mediates agonist-induced receptor-mediated TRPV4 activation in endothelial cells.