Effect of Flavanol-Rich Cocoa Extract in Acute Neurodegenerative Conditions

描述（由申请人提供）：据推测，急性和慢性神经退行性疾病中神经元细胞死亡的决定因素是由自由基损伤介导的。表儿茶素和可可（可可），已被报道是神经保护和潜在的预防药物，但潜在的细胞机制仍不清楚。我们的初步结果促使我们把注意力集中在富含（-）-表儿茶素和黄烷醇的可可提取物上，并测试血红素加氧酶（HO）活性可能参与可可提取物神经保护功能的假设。HO切割血红素（促氧化剂）以形成胆绿素/胆红素（抗氧化剂）、一氧化碳（血管扩张剂）和铁（通过增加铁蛋白将具有保护作用），已显示在氧化应激、缺血、炎症和高血压中起保护作用。虽然HO 2是组成型表达的，但HO 1是诱导型的。因此，通过增加HO 1水平来增加这种内源性抗氧化系统的活性可能是在细胞和器官水平上实现神经保护的一种方式。在我们使用原代神经元培养物的初步实验中测试的化合物中，表儿茶素是最有效的HO 1诱导剂之一。我们使用中风模型的研究结果也表明，口服表儿茶素可显著预防缺血再灌注脑损伤。总之，我们的研究结果表明，预处理与单一化合物表儿茶素，这是最丰富的可可提取物中的黄烷醇-并可能可可提取物本身-足以提供内源性神经保护，这表明在氧化应激过程中的共同治疗是不必要的。这些初步结果表明，HO 1的特异性诱导可能是可可提取物发挥其神经保护作用的一种机制，并促使我们提出，归因于可可提取物的一些神经保护作用可以通过导致内源性抗氧化剂途径刺激的途径来介导。在目标1中，我们将确定用表儿茶素和/或富含黄烷醇的可可提取物预处理（急性或慢性）的野生型（WT）小鼠全脑缺血后的神经元细胞死亡和行为结果，并测试这些效应是否在敲除小鼠中减弱。在目标2中，我们将确定由表儿茶素和/或富含黄烷醇的可可提取物诱导的HO 1表达的变化是否导致来自WT和基因敲除小鼠的神经元培养物中细胞存活的变化。总之，这些结果将帮助我们确定食用标准化可可提取物是否有益，以及可可提取物可以为大脑提供抵抗急性衰弱性神经退行性疾病的途径。 公共卫生相关性：几个世纪以来，可可（Theobroma cacao）一直被报道为加强神经系统的预防性药物，但其潜在的细胞机制仍不清楚。我们的初步结果促使我们把注意力集中在表儿茶素和可可提取物上，并检验导致血红素加氧酶（HO）诱导的途径可能参与可可的神经保护功能的假设。HO切割血红素（促氧化剂）以形成胆绿素/胆红素（抗氧化剂），已显示在氧化应激、缺血、炎症和高血压中起保护作用。使用临床前实验室小鼠模型，我们将确定是否预防性消费的标准化的富含黄烷醇的可可提取物可以防止神经功能下降和神经元细胞死亡后，全球缺血，从而提供新的途径，可可可以提供对急性神经系统疾病的脑抵抗力。

DESCRIPTION (provided by applicant): It has been postulated that the determinants of neuronal cell death in acute and chronic neurodegenerative conditions are mediated by free radical damage. Epicatechin and cocoa (Theobroma cacao), has been reported to be neuroprotective and a potential preventive medicine, but the underlying cellular mechanisms are still unclear. Our preliminary results prompted us to focus our attention on (-)-epicatechin and flavanol-rich cocoa extract and test the hypothesis that heme oxygenase (HO) activity could participate in cocoa extract neuroprotective function. HO, which cleaves heme (a prooxidant) to form biliverdin/bilirubin (antioxidants), carbon monoxide (a vasodilator), and iron (which by increasing ferritin would be protective) has been shown to play a protective role in oxidative stress, ischemia, inflammation, and hypertension. Although HO2 is constitutively expressed, HO1 is inducible. Consequently, an increase in the activity of this endogenous antioxidant system via an increase in HO1 levels, could be a way to achieve neuroprotection both at a cellular and organ levels. Of the compounds tested in our preliminary experiments using primary neuronal cultures, epicatechin was one of the most potent HO1 inducers. Our results using stroke models also suggest that epicatechin given orally significantly prevented ischemic-reperfusion brain damage. Together, our results indicate that pretreatment with the single compound epicatechin which is the flavanol most enriched in cocoa extract - and likely to cocoa extract itself - is sufficient to provide endogenous neuroprotection, suggesting that co-treatment during oxidative stress is not necessary. These preliminary results implied that specific induction of HO1 could be a mechanism by which cocoa extract exerts its neuroprotective actions and motivated us to propose that some of the neuroprotective effects attributed to Cocoa extract could be mediated through a pathway leading to stimulation of an endogenous antioxidant pathway. In Aim 1, we will determine neuronal cell death and behavioral outcomes following global ischemia in wildtype (WT) mice pre-treated (acutely or chronically) with epicatechin and/or flavanol-rich cocoa extract and test whether these effects are attenuated in knockout mice. In Aim 2, we will determine whether changes in HO1 expression induced by epicatechin and/or flavanol-rich cocoa extract result in changes in cell survival in neuronal cultures derived from WT and knockout mice. Together, these results will help us determine whether consumption of a standardized cocoa extract could be beneficial and the pathways by which cocoa extract could provide the brain with resistance to acute debilitating neurodegenerative conditions. PUBLIC HEALTH RELEVANCE: For centuries, cocoa (Theobroma cacao) has been reported as preventive medicine to strengthen the nervous system, but the underlying cellular mechanisms are still unclear. Our preliminary results prompted us to focus our attention on epicatechin and cocoa extract and test the hypothesis that the pathway leading to heme oxygenase (HO) enzyme induction could participate in cocoa's neuroprotective function. HO, which cleaves heme (a pro-oxidant) to form biliverdin/bilirubin (anti-oxidants) has been shown to play a protective role in oxidative stress, ischemia, inflammation, and hypertension. Using pre-clinical laboratory mouse models, we will determine whether prophylactic consumption of a standardized flavanol-rich cocoa extract can prevent neurological decline and neuronal cell death following global ischemia; thus providing new pathways by which cocoa could provide brain resistance against acute neurological disorders.