CARDIOPROTECTIVE EFFECTS OF PDE-5 INHIBITORS

描述(申请人提供)：磷酸二酯酶-5(PDE-5)抑制剂西地那非(伟哥)，已知可增强男性勃起功能，对心脏缺血-再灌注损伤(I/R)具有强大的心脏保护作用。我们建议对西地那非和其他临床使用的PDE-5抑制剂触发导致心脏保护的信号通路的机制进行进一步的深入研究。假设1：新型PDE-5抑制剂在心脏和心肌细胞中触发对I/R的保护。我们将研究短效(西地那非、异地那非)或长效(他达拉非)PDE-5抑制剂对缺血再灌注和细胞凋亡的保护作用。假设2：包括腺苷、缓激肽和一氧化氮在内的预适应介质的释放介导了PDE-5抑制剂诱导的心脏保护。使用药物抑制剂和Gone基因敲除小鼠的腺苷A1、缓激肽B2受体、eNOS、nNOS和iNOS。我们将确定这些介质在PDE-5抑制剂的心脏保护中的直接作用。假设3：PDE-5抑制剂选择性地激活在eNOS/iNOS/nNOS基因转录中起重要作用的PKC亚型，从而诱导心肌保护作用。我们将研究PDE-5抑制剂处理后PKC同工酶的易位。使用PKC同工酶的选择性多肽阻断剂，我们将确定它们在eNOS、iNOS或nNOS转录中的作用。假设4：PDE-5抑制剂刺激鸟苷环化酶并升高cGMP，导致蛋白激酶G(PKG)激活，这是打开mitoKATP通道的前奏。通过腺病毒基因转移PKG的剪接变异体或其零突变体，我们将确定它们在心肌细胞和完整心脏中对I/R的保护作用。假设5：PDE-5抑制剂通过激活转录因子HSF-1过表达HSPs来保护I/R。利用siRNAs，我们将确定HSF-1在PDE-5抑制剂对小鼠心脏保护中的因果关系。这些研究将为PDE-5抑制剂对I/R损伤的保护作用提供大量新的信息。预计这些研究将对将长期研究的缺血和药物预适应现象带入临床前沿产生巨大影响。

DESCRIPTION (provided by applicant): Phosphodiesterase-5 (PDE-5) inhibitor, sildenafil (Viagra), which is known to enhance erectile function in men, induces powerful cardioprotective effect against ischemia-reperfusion injury (I/R) in heart. We propose to perform further in depth studies on the mechanisms by which sildenafil and other clinically used PDE-5 inhibitors trigger signaling pathways that lead to cardioprotection. Hypothesis 1: Novel class of PDE-5 inhibitors trigger protection against I/R in heart and myocytes. We will study the effect of short acting (sildenafil, verdenafil) or long-acting (tadalafil) PDE-5 inhibitors on protective effect against I/R and apoptosis. Hypothesis 2: Release of preconditioning mediators, including adenosine, bradykinin and nitric oxide mediate cardioprotection induced by PDE-5 inhibitors. Using pharmacological inhibitors and gone knockout mice of adenosine A1, bradykinin B2 receptors, eNOS, nNOS and iNOS. we will determine the direct role of these mediators in cardioprotection by PDE-5 inhibitors. Hypothesis 3: PDE-5 inhibitors selectively activate PKC isoforms that play an essential role in the gene transcription of eNOS/iNOS/nNOS to induce cardioprotective effect. We will study translocation of PKC isozymes after treatment with PDE-5 inhibitors. Using selective peptide blockers of PKC isozymes, we will determine their role in gone transcription of eNOS, iNOS or nNOS. Hypothesis 4:PDE-5 inhibitors stimulate guanylate cyclase and elevate cGMP causing activation of protein kinase G (PKG) as a prelude to the opening of mitoKATP channels. Using adenoviral gene transfer of splice variants of PKG or their null mutants, we will determine their role in protection against I/R in cardiomyocytes and intact heart. Hypothesis 5:PDE-5 inhibitors protect against I/R by overexpression of HSPs through activation of transcription factor, HSF-1. Using siRNAs, we will determine the cause and effect relationship of HSF-1 in cardioprotection by PDE-5 inhibitors in mice. These studies, first of its kind, will provide tremendous amount of new information on protective effect of PDE-5 inhibitors against I/R injury. It is expected that the studies will have enormous impact on bringing the long-studied phenomenon of ischemic and pharmacologic preconditioning to the clinical forefront.