NO mobilizes intracellular Zn2+ via cGMP/PKG signaling pathway and prevents mitochondrial oxidant damage in cardiomyocytes
NO mobilizes intracellular Zn2+ via cGMP/PKG signaling pathway and prevents mitochondrial oxidant damage in cardiomyocytes
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DOI：
10.1016/j.cardiores.2007.05.015
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发表时间：
2007-07-15
影响因子：
10.8
通讯作者：
Xu, Zhelong
中科院分区：
文献类型：
--
作者：
Jang, Youngho;Wang, Huihua;Xu, Zhelong
目的：我们的目的是确定 NO 是否通过动员细胞内游离锌 (Zn2+) 来预防线粒体氧化损伤。方法：通过对负载 Newport Green DCF 的酶法分离成年大鼠心肌细胞进行成像来测定 Zn2+ 水平。通过负载四甲基罗丹明乙酯 (TMRE) 的心肌细胞成像来评估线粒体膜电位 (Delta Psi(m))。结果：S-亚硝基-N-乙酰青霉胺 (SNAP) 显着增加 Zn2+，而 ODQ 和 NS2028（鸟苷酸的两种特异性抑制剂）均能阻断 Zn2+ 环化酶。蛋白激酶 G (PKG) 抑制剂 KT5823 阻断 SNAP 的作用，而 PKG 激活剂 8-Br-cGNIP 模仿 SNAP 的作用，表明 cGMP/PKG 途径负责 SNAP 的作用。 5-羟基癸酸 (5HD) 可以阻止 Zn2+ 的增加，但二氮嗪可以模拟这种增加，这意味着线粒体 K-ATP 通道的开放可能是造成这种效应的原因。由于 Zn2+ 的螯合阻碍了 SNAP 对 H2O2 引起的 Delta Psi(m) 损失的预防作用，而外源锌 (1 μM ZnCl2) 阻止了 A Delta Psi(m) 的消散，因此 Zn2+ 可能在 NO 的保护作用中发挥关键作用。 MEK（丝裂原激活蛋白激酶或细胞外信号调节激酶）抑制剂 PD98059 阻断了 SNAP 和锌对 Delta Psi(m) 的预防作用，表明细胞外信号调节激酶 (ERK) 介导了这两种化合物对线粒体氧化损伤的保护作用。 Western blot分析进一步表明，ZnCl2显着增强ERK的磷酸化，证实ERK参与Zn2+的作用。结论：在离体心肌细胞中，NO通过cGMP/PKG途径打开线粒体K-ATP通道来动员内源性锌。在这些细胞中，Zn2+ 可能是 NO 对线粒体死亡途径作用的重要介质。 (c) 2007 年欧洲心脏病学会。由 Elsevier B.V. 出版。保留所有权利。
Objective: Our aim was to determine if NO prevents mitochondrial oxidant damage by mobilizing intracellular free zinc (Zn2+).Methods: Zn2+ levels were determined by imaging enzymatically isolated adult rat cardiomyocytes loaded with Newport Green DCF. Mitochondrial membrane potential (Delta Psi(m)) was assessed by imaging cardiomyocytes loaded with tetramethylrhodamine ethyl ester (TMRE).Results: S-nitroso-N-acetylpenicillamine (SNAP) dramatically increased Zn2+, which was blocked by both ODQ and NS2028, two specific inhibitors of guanylyl cyclase. The protein kinase G (PKG) inhibitor KT5823 blocked the effect of SNAP while the PKG activator 8-Br-cGNIP mimicked the action of SNAP, indicating that the cGMP/PKG pathway is responsible for the effect of SNAP. The increased Zn2+ was prevented by 5-hydroxydecanoate (5HD) but was mimicked by diazoxide, implying that mitochondrial K-ATP channel opening may account for this effect. Since chelation of Zn2+ blocked the preventive effect of SNAP on H2O2-induced loss of Delta Psi(m) and exogenous zinc (1 mu M ZnCl2) prevented dissipation of A Delta Psi(m), Zn2+ may play a critical role in the protective effect of NO. The MEK (mitogen-activated protein kinase or extracellular signal-regulated kinase) inhibitor PD98059 blocked the preventive effects of SNAP and zinc on Delta Psi(m), indicating that extracellular signal-regulated kinase (ERK) mediates the protective effect of both these compounds on mitochondrial oxidant damage. A Western blot analysis further showed that ZnCl2 significantly enhances phosphorylation of ERK, confirming the involvement of ERK in the action of Zn2+.Conclusions: In isolated cardiomyocytes, NO mobilizes endogenous zinc by opening mitochondrial K-ATP channels through the cGMP/PKG pathway. In these cells, Zn2+ may be an important mediator of the action of NO on the mitochondrial death pathway. (c) 2007 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.