Protection of Brain Injury from Cyanide Poisoning by Carnosic Acid

描述（由申请人提供）：氰化物中毒是一种潜在的生物恐怖分子，危及生命，急性暴露会导致缺氧性脑损伤、心肺功能衰竭，并在数分钟至数小时内死亡。这种急性毒性可通过各种解毒剂治疗，但成功取决于快速给药和有效渗透各种组织。然而，即使经过治疗，人类急性或慢性氰化物中毒也会诱发迟发性神经系统综合征，包括肌张力障碍。通常情况下，这些患者在数周至数月后表现出帕金森病症状，伴有进行性僵硬和上肢屈曲和下肢伸展的主要特征。这些患者的CT和MRI检查一致显示基底节病变，包括苍白球和壳核。人类大脑的损伤已经通过尸检得到证实。在急性和慢性氰化物暴露后也报告了类似的观察结果。认为多种机制是氰化物诱导的神经元损伤的基础，包括响应于氰化物诱导的脂质过氧化而抑制细胞色素C氧化酶（CcOX）和产生活性氧（ROS）。对神经元特别有害的是谷氨酸转运功能障碍和离子泵衰竭，通过过度刺激NMDA型谷氨酸受体（NMDAR）导致兴奋性毒性神经元细胞死亡。此外，氰化物诱导NMDAR的氧化还原位点的化学还原，这最初是在我们的实验室中发现的，并导致NMDAR操作的离子通道的额外激活、Ca 2+内流和随后的神经元损伤;这些机制考虑因素说明了抗氧化剂和NMDAR拮抗剂可以预防氰化物诱导的神经元损伤的事实。由于氰化物通过抑制CcOX和增强NMDAR的毒性作用都集中在氧化应激上，因此开发氰化物中毒的神经保护药物的一种可能策略是寻找可以对抗氧化损伤的低分子量化合物。在这里，作为潜在的对策，对中枢神经系统的毒性氰化物，我们建议使用鼠尾草酸（CA），亲电子化合物从迷迭香提取物，它穿过血脑屏障发挥作用，通过上调内源性抗氧化酶通过Nrf 2转录途径。我们假设，这种亲电化合物具有优势，抗氧化剂分子抵消氰化物在大脑中的毒性作用，因为它们的行动是更持久和放大的转录介导的信号通路。我们将通过研究CA在暴露于氰化物的培养神经元和氰化物中毒小鼠模型中的神经保护活性来验证这一假设。本提案的具体目标如下：具体目标1。探讨CA对氰化物中毒离体模型的神经保护作用。具体目标2。在模拟氰化物摄入效应的体内小鼠模型中检查CA的神经保护作用。 公共卫生相关性：氰化物中毒是一种潜在的生物恐怖剂，威胁生命。以亚慢性方式施用的亚致死剂量的氰化物中毒导致脑损伤，引起帕金森病症状，并且是由引起神经元中氧化应激的会聚代谢途径引起的。在这里，我们建议使用鼠尾草酸，一种在草药迷迭香中发现的亲电子化合物，来预防氰化物亚慢性中毒引起的脑损伤。

DESCRIPTION (provided by applicant): Cyanide poisoning is a potential bioterrorist agent, is life threatening, and acute exposure results in hypoxic brain injury, cardiopulmonary failure, and death within minutes to hours. Such acute toxicity is treatable by various antidotes, but success depends on rapid administration and effective penetration of the various tissues. However, even with treatment, acute or chronic cyanide intoxication in humans can induce a delayed neurological syndrome, including dystonia. Typically, these patients show Parkinsonian symptoms after weeks to months, with progressive rigidity and predominant features of flexed upper limbs and extended lower limbs. CT and MRI examinations of these patients have consistently revealed lesions in the basal ganglia, including the globus pallidus and putamen. Damage in the human brain has been confirmed via autopsy. Similar observations have been reported following both acute and chronic cyanide exposure. Multiple mechanisms are thought to underlie cyanide-induced neuronal damage, including inhibition of Cytochrome C oxidase (CcOX) and generation of reactive oxygen species (ROS) in response to cyanide- induced lipid peroxidation. Particularly harmful to neurons is dysfunctional glutamate transport and ionic pump failure, contributing to excitotoxic neuronal cell death by overstimulation of NMDA-type glutamate receptors (NMDARs). Furthermore, cyanide induces chemical reduction of the redox site(s) of the NMDAR, which were originally discovered in our laboratory, and result in additional activation of NMDAR-operated ion channels, Ca2+ influx, and consequent neuronal damage; these mechanistic considerations account for the fact that anti- oxidants and NMDAR antagonists can prevent cyanide-induced neuronal damage. As both toxic effects of cyanide by inhibition of CcOX and potentiation of NMDARs converge on oxidative stress, one possible strategy for the development of neuroprotective drugs for cyanide poisoning is to search for low-molecular-weight compounds that can counter oxidative damage. Here, as potential countermeasures against CNS toxicity by cyanide, we propose to use carnosic acid (CA), an electrophilic compound from rosemary extract, which crosses the blood-brain-barrier to exert effects by up-regulating endogenous anti-oxidant enzymes via the Nrf2 transcriptional pathway. We hypothesize that such electrophilic compounds have an advantage over antioxidant molecules for counteract the toxic effect of cyanide in brain because their action is more sustained and amplified by transcription-mediated signaling pathways. We will test this hypothesis by investigating the neuroprotective activity of CA in culture neurons exposed to cyanide and in a mouse model of cyanide poisoning. Specific Aims of this proposal are as follows: Specific Aim 1. To investigate neuroprotective effects of CA in an in vitro model of cyanide poisoning. Specific Aim 2. To examine neuroprotective effects of CA in an in vivo mouse model mimicking the effects of cyanide ingestion. PUBLIC HEALTH RELEVANCE: Cyanide poisoning is a potential bioterroristic agent and life threatening. Sublethal doses of cyanide poisoning administered in a subchronic fashion result in brain injury causing Parkinsonian symptoms, and result from converging metabolic pathways causing oxidative stress in neurons. Here, we propose to use carnosic acid, a pro-electrophilic compound found in the herb rosemary, to prevent brain injury from cyanide subchronic poisoning.