PARP in focal ischemia

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细胞类型的基因组完整性。用药物抑制该酶或通过转基因范式产生PARP缺乏可减轻脑缺血后的损伤。我们的初步研究表明，PARP零突变体（PARP-/-）的大脑中动脉闭塞后梗死（MCAO）相对于野生型小鼠显著减少。目的1将充分研究在PARP缺陷小鼠中观察到的这种明显的神经保护作用，描述在进化成梗死的日子里，完整动物的保护程度和寿命。我们将使用磁共振成像（MRI）对受伤小鼠的大脑进行无创检查，并将这些数据与功能行为评估和终末期组织病理学相结合。目的2将确定一氧化氮的产生对体内PARP激活和MCAO过程中随后的损伤的重要性。在小鼠MCAO模型中，研究了抑制PARP对神经元型或诱导型一氧化氮合酶的相对影响。最后，使用一种新的策略或在两个不同的催化和切割位点转染携带PARP突变体的病毒，我们将开始研究PARP在中风中的重要分子机制。c端NAD结合域在PARP介导的神经保护中的作用将通过比较PARP无效小鼠的卒中结果来确定，转染或不转染具有失活催化结构域的PARP突变形式。此外，我们将检测半胱氨酸分离酶(CASPASE)-PARP相互作用，比较PARP缺失小鼠卒中结果，转染和不转染缺乏活性CASPASE 3切割位点的PARP突变形式（D214G）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。

Poly(ADP-ribose) polymerase (PARP) is an abundant nuclear enzyme which helps to maintain genomic integrity in neurons and numerous other cell types. Inhibition of the enzyme with pharmacological agents or PARP deficiency generated via transgenic paradigms result in reduced injury after cerebral ischemia. Our preliminary studies show that infarction after middle cerebral artery occlusion (MCAO) in PARP null mutants (PARP-/-) is strikingly reduced relative to wild type mice. Aim 1 will fully examine this apparent neuroprotection observed in PARP deficient mice, characterizing extent and longevity of protection in the intact animal over the days of evolving infarction. We will use magnetic resonance imaging (MRI) to non-invasively examine the injured mouse brain and combine these data with functional behavior 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 MCAO. The relative consequences of inhibiting PARP versus neuronal or inducible nitric oxide synthase are examined in the mouse MCAO model. Lastly, using a novel strategy or transfecting virus carrying PARP mutant at two distinct catalytic and cleavage sites, we will begin to examine the molecular mechanism of PARP's important in stroke. The role of the C-terminal NAD binding domain in PARP- mediated neuroprotection will be determined by comparison of stroke outcomes in PARP null mice with or without transfection of a mutant form of PARP with inactive catalytic domain. In addition, we will examine cysteine apartase (CASPASE)-PARP interactions of comparison of stroke outcomes in PARP null mice with and without transfection of a mutant form of PARP lacking in active CASPASE 3 cleavage site (D214G) mutant. These experiments will further our understanding of PARP-mediated mechanisms of in vivo ischemic brain injury.