Division of Cell Biology, The Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada.
J Mol Cell Cardiol. 2013 Jul;60:142-50. doi: 10.1016/j.yjmcc.2013.04.010. Epub 2013 Apr 19.
Multiple initiatives are underway to harness the clinical benefits of remote ischemic preconditioning (rIPC) based on applying non-invasive, brief, intermittent limb ischemia/reperfusion using an external occluder. However, little is known about how rIPC induces protection in cardiomyocytes, particularly through G-protein coupled receptors. In these studies, we determined the role of opioid and adenosine receptors and their functional interactions in rIPC cardioprotection. In freshly isolated cardiomyocytes subjected to 45-min simulated ischemia followed by 60-min simulated reperfusion, we examined the ability of plasma dialysate (derived from blood obtained from rabbits remotely preconditioned by application of brief cycles of hind limb ischemia/reperfusion, rIPC dialysate) to protect cells against necrosis. rIPC dialysate and selective activation of either δ-opioid receptors or κ-opioid receptors significantly reduced the % of dead cells after simulated ischemia and simulated reperfusion. Inhibition of adenosine A1 receptors, but not adenosine A3 receptors, blocked the protection by rIPC dialysate, δ-opioid receptor and κ-opioid receptor activation. In HEK293 cells expressing either hemagglutinin A-tagged δ-opioid receptors or hemagglutinin A-tagged κ-opioid receptors, selective immunoprecipitation of adenosine A1 receptors pulled down both δ-opioid and κ-opioid receptors. This molecular association of adenosine A1 receptors with δ-opioid and κ-opioid receptors was confirmed by reverse pull-down assays. These findings strongly suggest that rIPC cardioprotection requires the activation of δ-opioid and κ-opioid receptors and relies on these receptors functionally interacting with adenosine A1 receptors.
多项计划正在进行中，旨在利用远程缺血预处理（rIPC）的临床益处，方法是使用外部闭塞器对非侵入性、短暂、间歇性肢体缺血/再灌注。然而，对于 rIPC 如何在心肌细胞中诱导保护作用，特别是通过 G 蛋白偶联受体，知之甚少。在这些研究中，我们确定了阿片类和腺苷受体及其在 rIPC 心脏保护中的功能相互作用的作用。在经历 45 分钟模拟缺血随后 60 分钟模拟再灌注的新鲜分离的心肌细胞中，我们检查了血浆透析液（源自通过短暂循环应用后肢缺血/再灌注远程预处理的兔子的血液获得）保护细胞免受坏死的能力。rIPC 透析液和选择性激活 δ-阿片受体或 κ-阿片受体显着降低模拟缺血和模拟再灌注后死亡细胞的百分比。抑制腺苷 A1 受体，但不抑制腺苷 A3 受体，阻断 rIPC 透析液、δ-阿片受体和 κ-阿片受体激活的保护作用。在表达血凝素 A 标记的 δ-阿片受体或血凝素 A 标记的 κ-阿片受体的 HEK293 细胞中，选择性免疫沉淀腺苷 A1 受体下拉了 δ-阿片和 κ-阿片受体。通过反向下拉测定证实了腺苷 A1 受体与 δ-阿片受体和 κ-阿片受体的这种分子关联。这些发现强烈表明，rIPC 心脏保护需要激活 δ-阿片受体和 κ-阿片受体，并且依赖于这些受体与腺苷 A1 受体的功能相互作用。
