Effects of heat shock protein 70 activation by metabolic inhibition preconditioning or κ-opioid receptor stimulation on Ca2+ homeostasis in rat ventricular myocytes subjected to ischemic insults
Effects of heat shock protein 70 activation by metabolic inhibition preconditioning or κ-opioid receptor stimulation on Ca2+ homeostasis in rat ventricular myocytes subjected to ischemic insults
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DOI：
10.1124/jpet.104.067926
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发表时间：
2004-08-01
影响因子：
3.5
通讯作者：
Wong, TM
中科院分区：
文献类型：
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作者：
Liu, J;Kam, KWL;Wong, TM
热休克蛋白70(HSP70)介导预适应的延迟心脏保护作用。细胞内钙超载([Ca~(2+)](I))引起损伤，而[Ca~(2+)](I)超载的减弱被认为是心肌保护的机制。有证据表明，HSP70与[Ca~(2+)](I)动态平衡有关。我们假设，通过预适应激活HSP70可以恢复因缺血损伤而改变的[Ca~(2+)](I)稳态。为了验证这一假设，我们确定了代谢抑制的预适应或U50,488H[trans-(+)-3，4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)[环己基]-苯乙酰胺(一种kappa阿片受体激动剂)对代谢抑制和缺氧(MI/A)并阻断热休克蛋白70合成的心肌细胞存活率和损伤、热休克蛋白70表达和[Ca+](I)的影响。用10 mM 2-脱氧-D-葡萄糖和10 mM二亚硫酸钠处理心肌细胞10分钟后，心肌细胞死亡细胞百分率、乳酸脱氢酶释放所反映的损伤程度和荧光分光光度法测定的静息[Ca~(2+)](I)显著增加，而电诱导的[Ca~(2+)](I)瞬变幅度降低。然而，在MI/A前20 h，心肌细胞在20 mM乳酸和10 mM 2-脱氧-D-葡萄糖(MIP)或30 um U_(50,488H)(U50,488H，UP)作用30分钟后，存活和损伤的变化。[Ca~(2+)](I)反应明显减弱。伴随而来的是HSP70表达的显著增加。此外，用选择性反义寡核苷酸阻断HSP70的合成可消除MIP或UP的有利作用。本研究首次证明，通过预适应诱导的热休克蛋白70的激活，部分恢复了因缺血损伤而改变的[Ca~(2+)](I)稳态。
Heat shock protein 70 (HSP70) mediates delayed cardioprotection of preconditioning. Cytosolic calcium ([Ca2+](i)) overload precipitates injury, whereas attenuation of [Ca2+](i) overload is believed to be responsible for cardioprotection. There is evidence suggesting a link between HSP70 and [Ca2+] (i) homeostasis. We hypothesize that activation of HSP70 by preconditioning may restore [Ca2+](i) homeostasis altered by ischemic insults. To test the hypothesis, we determined the effects of preconditioning with metabolic inhibition or pretreating with U50,488H [trans-(+)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl]-benzeneacetamide ( a kappa-opioid receptor agonist)] on viability and injury, HSP70 expression, and [Ca2+] (i) in ventricular myocytes subjected to metabolic inhibition and anoxia (MI/A), with blockade of HSP70 synthesis. In myocytes with vehicle pretreatment, the percentage of dead cells determined by trypan blue exclusion, the injury reflected by release of lactate dehydrogenase, and the resting [Ca2+](i) measured by spectrofluorometry significantly increased, whereas the amplitude of electrically induced [Ca2+](i) transient decreased, after 10 min with 10 mM 2-deoxy-D-glucose and 10 mM sodium dithionite, known to cause MI/A. However, when myocytes were subjected for 30 min to either 20 mM lactate and 10 mM 2-deoxy-D-glucose (MIP) or 30 muM U50,488H ( UP) 20 h before MI/A, the changes in viability and injury, and [Ca2+](i) responses were significantly attenuated. These were accompanied by a significantly increased HSP70 expression. Furthermore, blockade of HSP70 synthesis with selective antisense oligonucleotides abolished the beneficial effects of MIP or UP. This study provides first evidence that activation of HSP70 induced by preconditioning, which conferred delayed cardioprotection, restored partially the [Ca2+](i) homeostasis altered by ischemic insults.