PARS ACTIVATION AFTER TBI

聚(ADP-核糖)合成酶，又称聚(ADP-核糖)聚合酶或PARP，是一种普遍存在的核蛋白，在消耗NAD和ATP的过程中促进DNA链断裂的修复。几项针对非神经细胞的体外研究支持“PARs激活的自杀理论”，即活跃的PARs会导致能量衰竭和细胞坏死。我们的初步研究提供了创伤性脑损伤(TBI)后激活PARS的证据。PARS缺陷的小鼠表现出明显的功能缺陷，这通常是由脑损伤造成的，这表明在脑损伤后，PARS的激活是有害的。我们的假设是，过氧亚硝酸盐引起的DNA氧化损伤激活了PARS，扰乱了细胞能量学，损伤了线粒体，并导致了坏死细胞的死亡。解决这一假说的具体目的包括使用药理PARS抑制剂测试过氧亚硝酸盐诱导的PARS激活对体外培养的神经元DNA氧化损伤、细胞能量学、线粒体功能以及细胞凋亡性坏死和总细胞死亡的影响。通过以下步骤确定脑损伤后PARS的作用：首先确定小鼠、大鼠和人类脑损伤后PARS激活的特征；然后使用药物抑制剂或基因敲除小鼠(PARS基因敲除小鼠)，测试PARS激活对NAD储备、线粒体损伤、脑水肿和脑血流、细胞凋亡和总细胞死亡以及神经病理结果的影响。通过抑制PARs来保存细胞能量储存可能是治疗TBI的关键策略。重要的是，抑制PARS可能针对坏死性神经元死亡，这在以前一直被认为是无法操纵的。然而，PAR也可能通过参与维持基因组的完整性而在脑损伤后发挥有益的作用。在实施针对急性脑损伤后PARS的临床治疗策略之前，需要仔细研究脑损伤后PARS的潜在双重作用。脑损伤在没有警告的情况下发作，是成人和儿童发病和死亡的主要原因。细胞能量衰竭和坏死性神经元死亡会导致发病率和死亡率，目前只有少数非特异性治疗方法可用。拟议的实验解决了关键问题，过氧亚硝酸盐产生的氧化应激是否在体外和体内脑损伤后触发PARS介导的线粒体损伤、能量衰竭和神经元死亡？针对脑外伤后能量衰竭和坏死的新的和临床相关的药理学策略的发展可能被证明是治疗脑损伤的有效药物。

Poly(ADP-ribose) synthetase (PARS), also termed poly (ADP- ribose)polymerase or PARP, is a ubiquitous nuclear protein that facilitates repair of DNA strand breaks in a process that consumes NAD and ATP. Several in vitro studies in non-neuronal cells support a "suicide theory of PARS activation" where active PARS leads to energy failure and cell death by necrosis. Our preliminary studies provide evidence that PARS is activated after traumatic brain injury (TBI). Mice deficient in PARS showed dramatic from functional deficits typically produced by TBI, suggesting that in toto PARS activation is detrimental after TBI. Our hypothesis is that oxidative DNA damage produced by peroxynitrite activates PARS, disturbs cellular energetics, injures mitochondria, and contributes to necrotic cell death. Specific aims to address this hypothesis include testing the effects of peroxynitrite-induced PARS activation on oxidative DNA damage, cellular energetics, mitochondrial function, and apoptotic necrotic and total cell death in neurons in vitro using pharmacologic PARS inhibitors. The role of PARS after TBI will be established by first characterizing PARS activation in mice, rats, and human samples after TBI; then testing the effects of PARS activation on NAD stores, mitochondrial injury, brain edema and cerebral blood flow, apoptotic necrotic, and total cell death, and neuropathologic outcome after TBI using pharmacologic inhibitors or genetic disruption (PARS knockout mice). Preserving cellular energy stores by inhibiting PARS may represent a key strategy for the treatment of TBI. Importantly, PARS inhibition may target necrotic neuronal death which has been previously felt to be unmanipulable. However, PARS may also play a beneficial role after TBI by participating in the maintenance of genomic integrity. The potential dual-role of PARS after TBI needs to be carefully addressed prior to implementation of clinical treatment strategies targeting PARS after acute brain injury. TBI strikes without warning and is a major cause of morbidity and mortality in adults and children. Cellular energy failure and necrotic neuronal death contribute to morbidity and mortality and currently only few non-specific therapies are available. The proposed experiments address the key question, does oxidative stress produced by peroxynitrite trigger PARS-mediated mitochondrial damage, energy failure, and neuronal death in vitro and after TBI in vivo? The development of novel and clinically-relevant pharmacologic strategies that target energy failure and necrosis after TBI may prove to be powerful and efficacious agents for the treatment of TBI.