PARP in excitotoxic and apoptotic neuronal injury

PARP 在兴奋性毒性和凋亡性神经元损伤中的作用

基本信息

批准号：
6349244
负责人：
VALINA L. DAWSON
金额：
$ 22.49万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-05-01 至 2001-04-30
项目状态：
已结题

项目摘要

Stroke is the third leading cause of death and disability in the United States. Glutamate is a major mediator of neuronal damage in focal ischemia models of stroke. This excitotoxicity is mediated in part by mitochondrial dysfunction and free radical induced oxidative damage. The culture and animal models, glutamate mediates most of its toxicity through activation of NMDA receptors. The increase in intracellular Ca2+ concentration leads to an activation of Ca2+ dependent enzymes, including neuronal nitric oxide synthase (nNOS) which produces nitric oxide (NO). Additionally the increase in intracellular Ca2+ results in increased superoxide anion which can react with NO to from peroxynitrite. Increased peroxynitrite can induce DNA nicks and breaks which activates the nuclear enzyme, poly (ADP-ribose) polymerase (PARP) which catalyzes the transfer of ADP-ribose from NAD to nuclear proteins. For every model of NAD consumed, four free energy equivalents of ATP are required to restored NAD levels. It is possible that this consumption of NAD and subsequently ATP in an injured cell leads to cell death, however, the mechanism(s) by which PARP activation mediates neuronal death are not known. Consistent with pharmacologic inhibition of PARP, primary cortical cultures from PARP knockout mutant mice are resistant to NMDA and NO neurotoxicity and combined oxygen-glucose deprivation induced neuronal damage. Consistent with these in vitro findings, PARP knockout mutant mice have remarkably reduced infarct volumes following transient focal ischemia. Protection in the PARP knockout cultures and mice exceed protection observed with nNOS knockout mice or following treatment with the NMDA receptor antagonist, MK801 suggesting that other cell death signals may converge on PARP to elicit toxicity in ischemia-reperfusion injury. PARP is a complex protein. The biochemistry and cell biology of PARP in post-mitotic neurons is not known. The mechanism of activation of PARP in post-mitotic neurons is assumed to be DNA nicks but this is not known. The mechanism of PARP mediated neurotoxicity are not known. The following specific aims are proposed to investigate PARP neurobiology and discover the actions of PARP that result in neuronal cell death: 1. What is the role of PARP in excitotoxic versus apoptotic neuronal death? 2. What is the role of reactive oxygen species and DNA damage in the activation of PARP? 3. What is the interaction between caspase activation and PARP-mediated neurotoxicity? 4. What is the role of energy impairment in PARP mediated neuronal death? 5. What is the role of protein ribosylation in PARP mediated neuronal death?

中风是美国死亡和残疾的第三大主要原因。谷氨酸是中风局灶性缺血模型中神经元损伤的主要介体。这种兴奋性毒性部分是通过线粒体功能障碍和自由基诱导的氧化损伤介导的。培养物和动物模型，谷氨酸通过激活NMDA受体介导了其大部分毒性。细胞内Ca2+浓度的增加导致Ca2+依赖性酶的激活，包括产生一氧化氮（NO）的神经元一氧化氮合酶（NNOS）。另外，细胞内Ca2+的增加导致超氧化阴离子增加，这可能与过氧亚硝酸盐反应不反应。过氧亚硝酸盐增加可以诱导DNA划痕和断裂，从而激活核酶，聚（ADP-核糖）聚合酶（PARP），从而催化ADP-核糖从NAD到核蛋白的转移。对于每种消耗的NAD模型，需要四个自由能当量的ATP恢复NAD级别。 NAD和随后在受伤的细胞中的ATP的消耗可能导致细胞死亡，但是，PARP激活介导神经元死亡的机制尚不清楚。与PARP的药理学抑制一致，来自PARP基因敲除突变小鼠的原代皮质培养物具有对NMDA的抗性，没有神经毒性，氧气葡萄糖剥夺诱导了神经元损伤。与这些体外发现一致，PARP敲除突变小鼠在短暂性局灶性缺血后的梗塞体积大大减少。 PARP基因敲除培养物和小鼠的保护超出了NNOS敲除小鼠观察到的保护，或者在用NMDA受体拮抗剂治疗后，MK801表明其他细胞死亡信号可能会在PARP上汇聚以引起缺血再灌注损伤的毒性。 PARP是一种复杂的蛋白质。 PARP的生物化学和细胞生物学在有丝分裂后神经元中尚不清楚。假定有丝分裂神经元中PARP激活的机理被认为是DNA迹象，但这尚不清楚。 PARP介导的神经毒性的机制尚不清楚。提出了以下具体目的来研究PARP神经生物学并发现导致神经元细胞死亡的PARP的作用：1。PARP在兴奋性毒性与凋亡神经元死亡中的作用是什么？ 2。活性氧和DNA损伤在PARP激活中的作用是什么？ 3。胱天蛋白酶激活与PARP介导的神经毒性之间的相互作用是什么？ 4。能量障碍在PARP介导的神经元死亡中的作用是什么？ 5。蛋白质核糖基化在PARP介导的神经元死亡中的作用是什么？