Protection of Brain Injury from Cyanide Poisoning by Carnosic Acid

描述(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、心肺衰竭，并在几分钟至几小时内死亡。这种急性毒性可以用各种解毒剂治疗，但成功与否取决于快速给药和各种组织的有效渗透。然而，即使接受治疗，人类的急性或慢性氰化物中毒也会导致迟发性神经综合征，包括肌张力障碍。通常，这些患者在几周到几个月后就会出现帕金森症状，并伴有渐进性僵硬，主要特征是上肢弯曲和下肢伸展。这些患者的CT和MRI检查始终显示基底节病变，包括苍白球和壳核。人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。多种机制被认为是氰化物诱导的神经元损伤的基础，包括抑制细胞色素C氧化酶(CcOX)和在氰化物诱导的脂质过氧化反应中产生活性氧(ROS)。对神经元尤其有害的是谷氨酸转运功能障碍和离子泵衰竭，过度刺激NMDARs导致兴奋性毒性神经细胞死亡。此外，氰化物导致NMDAR氧化还原位点(S)的化学还原，导致NMDAR操作的离子通道的额外激活，钙内流，从而导致神经元损伤；这些机制考虑解释了抗氧化剂和NMDAR拮抗剂可以防止氰化物引起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个可能的策略是寻找能够对抗氧化损伤的低分子化合物。在这里，我们建议使用迷迭香提取物中的亲电化合物鼠尾草酸(CA)，作为对抗氰化物对中枢神经系统毒性的潜在对策，它通过Nrf2转录途径上调内源性抗氧化酶，从而跨越血脑屏障发挥作用。我们推测，与抗氧化剂分子相比，这种亲电化合物在对抗氰化物对大脑的毒性作用方面具有优势，因为它们的作用更持久，并通过转录介导的信号通路得到放大。我们将通过研究CA在氰化物暴露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。本研究的具体目的如下：1.在体外氰化物中毒模型上观察CA的神经保护作用。具体目的2.在模拟氰化物摄入的小鼠体内模型中，检测CA的神经保护作用。

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.