Anesthetic-induced preconditioning delays opening of mitochondrial permeability transition pore via protein Kinase C-epsilon-mediated pathway.
Anesthetic-induced preconditioning delays opening of mitochondrial permeability transition pore via protein Kinase C-epsilon-mediated pathway.
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DOI：
10.1097/aln.0b013e3181a91957
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发表时间：
2009-08
期刊：
影响因子：
8.8
通讯作者：
Bosnjak ZJ
中科院分区：
文献类型：
--
作者：
Pravdic D;Sedlic F;Mio Y;Vladic N;Bienengraeber M;Bosnjak ZJ
挥发性麻醉剂诱导的预适应(APC)对心脏的保护作用涉及蛋白激酶C(PKC)的激活。本研究探讨了APC激活的PKC在延缓线粒体通透性转换孔(MPTP)开放中的重要性。大鼠心肌细胞暴露于异氟烷中，加入或不加入非选择性蛋白激酶C抑制剂白屈菜红碱或蛋白激酶C-δ(Rotlerin)和蛋白激酶C-ε(Myristoylated PKC-εV1-2)的同种异构体特异性抑制剂，用激光共聚焦显微镜测定其开放时间。在有无白屈菜红碱存在和不存在的情况下，观察了异氟烷对大鼠线粒体的开放作用，以及分离后直接用异氟烷处理的线粒体。用Western blotting检测pKC-ε在APC和对照心肌细胞中的转位。在心肌细胞中，APC延长了诱导MPTP开放所需的时间(2 61±2 6 S比2 16±2 7 S对照组；P<；0.0 5)，而白屈菜红碱使这一延迟消失为2 13±2 2 S。当应用PKC-ε抑制剂时，异氟醚的作用也被取消(2 10±2 2 S)，但在PKC-δ抑制剂的存在下(2 6 9±31 S)。Western blotting显示蛋白激酶C-ε在APC细胞中向线粒体易位。APC大鼠线粒体开放MPTP所需的钙离子浓度显著高于对照组(45±8μM mg-1比±8μM mg-1APC)，白屈菜红能逆转APC的作用。相比之下，异氟醚不能保护直接治疗的线粒体。APC通过PKC-ε介导抑制MPTP开放，而不是通过PKC-δ诱导MPTP延迟开放。这些结果表明异氟醚保护心脏的胞浆成分和线粒体成分之间存在联系。
Cardioprotection by volatile anesthetic-induced preconditioning (APC) involves activation of protein kinase C (PKC). The current study investigated the importance of APC-activated PKC in delaying mitochondrial permeability transition pore (mPTP) opening. Rat ventricular myocytes were exposed to isoflurane in the presence or absence of nonselective PKC inhibitor chelerythrine or isoform-specific inhibitors of PKC-δ (rottlerin) and PKC-ε (myristoylated PKC-ε V1-2 peptide), and the mPTP opening time was measured using confocal microscopy. Ca2+-induced mPTP opening was measured in mitochondria isolated from rats exposed to isoflurane in the presence and absence of chelerythrine, or in mitochondria directly treated with isoflurane after isolation. Translocation of PKC-ε was assessed in APC and control cardiomyocytes by Western blotting. In cardiomyocytes, APC prolonged time necessary to induce mPTP opening (261±26 s APC vs. 216±27 s control; P<0.05), while chelerythrine abolished this delay to 213±22 s. The effect of isoflurane was also abolished when PKC-ε inhibitor was applied (210±22 s), but not in the presence of PKC-δ inhibitor (269±31 s). Western blotting revealed translocation of PKC-ε toward mitochondria in APC cells. The Ca2+ concentration required for mPTP opening was significantly higher in mitochondria from APC rats (45±8 μM mg-1 control vs. 64±8 μM mg-1 APC), and APC effect was reversed with chelerythrine. In contrast, isoflurane did not protect directly treated mitochondria. APC induces delay of mPTP opening through PKC-ε-mediated inhibition of mPTP opening, but not through PKC-δ. These results point to the connection between cytosolic and mitochondrial components of cardioprotection by isoflurane.