PARP in Cardiac Arrest

尽管关于心脏骤停/心肺复苏(CPR)的研究已有40年，但临床结果仍然很差。这项研究的大部分形式是开发新的心肺复苏方法学或药理学方法。然而，现在需要新的基础科学和分子遗传学方法来提高我们对心脏骤停/心肺复苏过程中损伤的关键机制的理解，以开发新的治疗干预措施。多聚(ADP-核糖)聚合酶(PARP)是一种丰富的核酶，有助于维持 神经元和许多其他类型细胞的基因组完整性。用药物抑制PARP或通过PARP基因敲除动物使PARP缺乏可减轻局灶性脑缺血后的损伤，但有关PARP和心脏骤停/心肺复苏的信息很少。我们的初步研究表明，与野生型小鼠相比，PARP缺失突变体(PARP-/-)在心脏骤停/心肺复苏后的细胞损伤显著减少。目的1将研究在PARP缺陷小鼠中观察到的这种明显的神经保护，描述在进化损伤的几天内完整动物的保护范围和寿命。我们将结合使用随时间推移的功能行为评估和终末组织病理学。目的2将确定一氧化氮的产生对活体PARP激活和心脏骤停/心肺复苏过程中随后的损伤的重要性。抑制PARP与神经元或诱导型一氧化氮合酶的相对后果将在心脏骤停/CPR小鼠模型中进行检验。最后，使用一种在两个不同的催化和切割位点上转染携带PARP突变体的病毒的策略，我们将 开始研究PARP在心脏骤停/心肺复苏中的重要作用的分子机制。C端NAD结合域在PARP介导的神经保护中的作用将通过比较在PARP-/-中心脏骤停/心肺复苏的结果而确定，无论是否转染具有非活性催化结构域的突变形式的PARP。此外，我们还将通过比较在PARP-/-中是否转染缺乏caspase 3裂解位点的突变形式的PARP(D214G)来检查半胱氨酸天冬氨酸酶(Caspase)-PARP之间的相互作用 突变体)。这些实验将进一步加深我们对PARP介导的体内缺血性脑损伤机制的理解，并可能为开发有效的、新的治疗策略以保护心脏骤停/心肺复苏后的神经功能提供合理的基础。

Despite four decades of research concerning cardiac arrest/cardiopulmonary resuscitation (CPR), clinical outcome remains poor. Much of this research has taken the form of developing new methodologic or pharmacologic approaches to CPR. However, now, novel basic science and molecular genetic approaches to CPR are needed to improve our understanding of critical mechanisms of injury during cardiac arrest/CPR to develop new therapeutic interventions. Poly (ADP-ribose) polymerase (PARP) is an abundant nuclear enzyme which helps maintain genomic integrity in neurons and numerous other cell types. Inhibition of PARP with pharmacologic agents or PARP deficiency via PARP knockout animals reduces injury after focal cerebral ischemia, but there is little information concerning PARP and cardiac arrest/CPR. Our preliminary studies show that cell injury after cardiac arrest/CPR in PARP null mutants (PARP-/-) is markedly reduced relative to wild type mice. Aim 1 will investigate this apparent neuroprotection observed in PARP deficient mice, characterizing extent and longevity of protection in intact animals over days of the evolving injury. We will use a combination of functional behavioral evaluation over time and terminal histopathology. Aim 2 will establish the importance of nitric oxide generation to PARP activation in vivo and subsequent injury during cardiac arrest/CPR. The relative consequences of inhibiting PARP vs neuronal or inducible nitric oxide synthase will be examined in the cardiac arrest/CPR mouse model. Lastly, using a strategy of transfecting virus carrying PARP mutant at two distinct catalytic and cleavage sites, we will begin to examine the molecular mechanism of PARP's importance in cardiac arrest/CPR. The role of the C-terminal NAD binding domain in PARP-mediated neuroprotection will be determined by comparison of cardiac arrest/CPR outcome in PARP-/- with or without transfection of a mutant form of PARP with inactive catalytic domain. Also, we will examine cysteine aspartase (CASPASE)-PARP interactions by comparison of cardiac arrest/CPR outcome in PARP-/- with and without transfection of a mutant form of PARP lacking in CASPASE 3 cleavage site (D214G mutant). These experiments will further our understanding of PARP-mediated mechanisms of in vivo ischemic brain injury and may provide a rational basis for development of effective, new therapeutic strategies to preserve neurological function atter cardiac arrest/CPR.