Endothelin-1 and PKC Induce Positive Inotropy Without Affecting pHi in Ventricular Myocytes
Endothelin-1 and PKC Induce Positive Inotropy Without Affecting pHi in Ventricular Myocytes
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Endothelin-1 和 PKC 在心室肌细胞中诱导正性肌力而不影响 pHi
DOI：
10.3181/00379727-231-2310865
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发表时间：
2006
影响因子：
3.2
通讯作者：
Jeffery W. Walker
中科院分区：
文献类型：
--
作者：
Misuk Kang;Jeffery W. Walker
已有研究认为，细胞内碱化是内皮素(ET)-1引起的心肌细胞收缩能力增强的基础。用pH敏感的荧光染料snarf-1观察ET-1对培养的成年大鼠心室肌细胞收缩性能和细胞内pH的影响。在ET-1刺激下，观察到不同的PHI变化。大多数心肌细胞(n=20/32)没有碱化，但对ET-1的反应表现出大约60%的收缩幅度增加。在剩余的心肌细胞中(12/32)，ET-1引起pHi升高0.05±0.02个pH单位，其收缩幅度也增加了约60%。因此，ET-1介导的正性肌力(增强的肌力)与细胞内碱化之间没有很强的相关性。为了确定ET-1的收缩和PHI反应是否由蛋白激酶C(PKC)介导，将黄色荧光蛋白(YFP)-显性负性(DN)PKC构建物作为异构体特异性抑制物。在表达dN-PKC-ε-yfp的心肌细胞中，ET-1介导的正性变力反应显著减弱至13±15%，但仍可观察到碱化现象。DN-PKC-δ-yfp的表达也不能阻断碱化，但在这种情况下仍可观察到正性变力反应。在先前的研究中，我们发现PKC-δ和PKC-ε的表达对佛波醇12，13-二丁酸酯(PDBu)的刺激有很强的正性肌力作用。使用这个系统，PDBu不能影响大多数表达PKC的心肌细胞的phi，尽管>；60%的抽动幅度增加了。总体而言，对于ET-1和PDBu，正性变力反应和碱化的相关性很差，无论是否有dN-PKC结构，以及是否有野生型PKC结构。这些结果表明，ET-1通过PKC-ε产生正性肌力作用，而不是通过细胞内碱化作用。
It has been proposed that intracellular alkalinization underlies the enhanced contractility of ventricular myocytes exposed to endothelin (ET)-1. The effects of ET-1 on the contractility and intracellular pH (pHi) were examined here in cultured adult rat ventricular myocytes by employing the pH-sensitive fluorescent dye SNARF-1. Variable pHi changes were observed on ET-1 stimulation. Most myocytes (n = 20 of 32) did not alkalinize, but showed an approximate 60% increase in twitch amplitude in response to ET-1. In the remaining myocytes (12 of 32), ET-1 induced an increase in pHi by 0.05 ± 0.02 pH units with a similar approximate 60% increase in twitch amplitude. Therefore, there was no strong correlation between ET-1–mediated positive inotropy (enhanced contractility) and intracellular alkalinization. To determine whether ET-1 contractile and pHi responses were mediated by protein kinase C (PKC), yellow fluorescent protein (YFP)–fused dominant negative (dn) PKC constructs were used as isoform specific inhibitors. In dn-PKC-ε-YFP–expressing myocytes, the ET-1–mediated positive inotropic response was greatly diminished to 13 ± 15%, but alkalinization was still observed. Expression of dn-PKC-δ-YFP also did not block alkalinization, but in this case the positive inotropic response was still observed. In a previous study, we showed that expression of PKC-δ and PKC-ε caused a strong positive inotropy on stimulation with phorbol 12,13-dibutyrate (PDBu). Using this system, PDBu failed to affect pHi in the majority of PKC expressing myocytes despite increases in twitch amplitudes of >60%. Overall, the poor correlation of positive inotropic responses and alkalinization was observed for ET-1 with and without dn-PKC constructs and for PDBu with and without wild-type PKC constructs. These results suggest that ET-1 produces positive inotropy via PKC-ε by mechanisms other than intracellular alkalinization.
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DOI：
10.1152/ajpheart.1998.275.4.h1473
发表时间：
1998
期刊：
The American journal of physiology
影响因子：
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作者：
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通讯作者：
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影响因子：
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作者：
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影响因子：
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作者：
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影响因子：
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作者：
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