Mechanistic studies on stress, inflammation and neuroprotection

应激、炎症和神经保护的机制研究

• 批准号: 8158090
• 负责人: JUAN M SAAVEDRA
• 金额: $ 121.7万
• 依托单位: NATIONAL INSTITUTE OF MENTAL HEALTH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8158090
• 关键词: Inflammation; Stress; neuroprotection

Summary: Our first specific aim is to clarify the mechanisms involved in the beneficial effects of compounds which have not, until very recently, been considered of interest to the therapy of brain disorders. The second specific aim is to further establish the extent of therapeutic benefits of such compounds in diseases of the brain. We study a group of compounds collectively named sartans, or Angiotensin II AT1 receptor blockers (ARBs). Sartans are biphenyl derivatives with an excellent margin of safety, extensively used to treat high blood pressure because they antagonize Angiotensin II-induced vasoconstriction. We have previously discovered that sartan treatment reduces brain ischemia, stress, and anxiety, and increases lifespan in rodent models. During the current fiscal year, we studied the mechanisms of anti-inflammatory effects of sartans in the brain. There was a dual impact of sartan treatment in rats subjected to systemic administration of bacterial endotoxin, lipopolysaccharide (LPS). First, we observed that sartans decreased LPS-induced peripheral overproduction of pro-inflammatory cytokines (interleukin-1 beta, tumor necrosis factor alpha, interleukin-6, fractalkine). Enhanced cytokine production and release to the circulation is partially responsible for the central initial inflammatory response to LPS. Second, we found direct anti-inflammatory effects of sartans in brain parenchyma, evidenced by decreased LPS-induced activation of inflammatory cascades. Following sartan treatment in vivo, we found a decline in the LPS-induced activation of the transcription factors nuclear factor kappa-light-chain-enhancer of activated B cells (NFkappaBalpha) and activator protein-1 (AP-1). Sartans also reduced expression of inducible nitric oxide synthase, cyclooxygenase-2 and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase induced by LPS, enzymes involved in the production of excess nitric oxide, prostaglandin E2, and reactive oxygen species leading to brain inflammation and neuronal injury. In addition, treatment with sartans produced a widespread decrease in LPS-induced microglia activation throughout the brain. Using cell cultures of human circulating monocytes and primary neuronal cultures, we discovered direct anti-inflammatory effects of ARBs against LPS and interleukin-1beta. These effects involve decreased activation of the protein kinases and the transcription factor NFkappaBalpha. In addition, sartans neuroprotective and anti-inflammatory effects were clearly established by cell viability assays and determination of inflammatory factors in cultured neurons and cerebral microvascular endothelial cells. We hypothesized that the major anti-inflammatory and neuroprotective effects of sartans may not be the exclusive result of AT1 receptor inhibition. In human circulating monocytes, cells expressing very few AT1 receptors, the anti-inflammatory effects of sartans were partially dependent on peroxisome proliferator-activated receptor gamma (PPARgamma) activation. We have found that some sartans may have dual mechanisms of action: anti-hypertensive, anti-growth and anti-inflammatory effects related to their inhibition of AT1 receptors, and metabolic and anti-inflammatory effects, partially the consequence of direct PPARgamma activation. Studies on rodent models of systemic inflammation established that anti-inflammatory effects of sartans in the brain are widespread, and not only related to classical brain targets for circulating inflammatory factors such as the hypothalamic paraventricular nucleus and areas outside the blood brain barrier. Reduced inflammation was also noted in multiple cortical areas including the prefrontal cortex, a key regulatory structure controlling the emotional response to stress. Our demonstration of protection from endotoxin in cerebrovascular endothelial cells indicated that the widespread anti-inflammatory effect of sartans involves AT1 receptor blockade in brain endothelial cells. These receptors, first characterized in our laboratory, are fundamental elements regulating the brain response to inflammation and the reaction to stress. In addition, sartans limit the exaggerated hormonal and sympathetic response to stress, effects dependent on the type and intensity of the challenge. For instance, while sartans completely prevent the hormonal and sympathetic response to isolation, these compounds are able to preserve the anti-inflammatory corticosterone response in inflammatory stress models. Moreover, sartans can prevent formation and release of aldosterone, a pro-inflammatory hormone stimulating brain mineralocorticoid receptors in the hippocampus. We also found, in rats submitted to acute restraint, that the anti-stress and anti-anxiety effects of sartan administration included prevention of the cortical gamma-aminobutyric acid (GABAA) receptor alterations characteristic of stress. Prevention of decreased activation of a major inhibitory system such as GABAA, explains the consistent anti-anxiety and anti-stress effects of sartan administration. These discoveries highlight the multiple central roles of AT1 receptors in the control of stress, inflammation, and mood. The anti-stress effects of ARBs have behavioral correlates. Sartan administration prevented LPS-induced sickness behavior, characterized by anorexia and weight loss, which occurs with systemic inflammation. In addition, sartans eliminated the anxiety responses produced by systemic LPS administration. In collaborative studies, we are analyzing the anti-depressant effects of sartan treatment, and the degree of involvement of the brain Angiotensin II system in the mechanisms of susceptibility to chronic variable stress. We have continued to clarify the mechanisms of neuroprotection and prolongation of lifespan produced by life-long sartan administration. We have found profound anti-inflammatory effects in the brain, including prevention of NADPH activation, which is responsible for age-dependent generation of neurotoxic reactive oxygen species, and a reversal of age-related cerebrovascular remodeling. We also found that the anti-anxiety effects of sartans persist throughout life. Our findings may, at least in part, explain the major beneficial effects of sartan therapy in Alzheimers disease. Our work continues with more advanced mechanistic studies including the use of gene knockout mouse models, microarray assays, and selective gene-silencing and phenotype-rescue experiments in cultured cells, and studies on animal models to determine the precise therapeutic range of sartans and related molecules. We are also interested in determining the extent of AT1 receptor blockade-dependent and independent effects of sartans. This information is very valuable for further discovery of sartan molecules of even better therapeutic value. Our mechanistic and translational studies will hopefully lead to novel treatment for disorders of the brain.

概括： 我们的第一个具体目的是阐明化合物的有益作用所涉及的机制，直到最近才被认为是脑部疾病治疗感兴趣的。第二个具体目的是进一步确定大脑疾病中这种化合物的治疗益处的程度。 我们研究一组统称为sartans或血管紧张素II AT1受体阻滞剂（ARB）的化合物。 sartans是具有极大安全缘的双苯基衍生物，广泛用于治疗高血压，因为它们会拮抗血管紧张素II诱导的血管收缩。 我们以前已经发现，Sartan治疗可减少脑缺血，压力和焦虑，并在啮齿动物模型中增加寿命。 在当前财政年度，我们研究了大脑中萨坦的抗炎作用的机制。在全身服用细菌内毒素，脂多糖（LPS）的大鼠中，Sartan治疗对大鼠产生了双重影响。首先，我们观察到Sartans降低了LPS诱导的促炎性细胞因子（白介素-1β，肿瘤坏死因子α，白介素-6，Fractalkine）的周围周围过量产生。 增强的细胞因子产生和循环释放是部分原因是中央初始炎症反应对LPS的中心炎症反应。其次，我们发现了sartans在脑实质中的直接抗炎作用，这是通过LPS诱导的炎症级联反应降低证明的。在体内治疗后，我们发现LPS诱导的转录因子的激活下降了活化B细胞（NFKappabalpha）和激活蛋白-1（AP-1）的转录因子Kappa-Light-light链增强剂。 Sartans还降低了诱导型一氧化氮合酶，环氧合酶-2和烟酰胺腺苷二核苷酸磷酸腺苷（NADPH）氧化酶，LPS诱导的LPS，酶，参与过量氧化氮，前列腺素E2的产生，前列腺素E2的产生，并导致脑炎症和Neuronaronal损伤。 此外，用sartans的治疗导致LPS诱导的小胶质细胞激活在整个大脑中的广泛降低。 利用人循环单核细胞和原发性神经元培养的细胞培养物，我们发现了ARB对LPS和白介素1Beta的直接抗炎作用。这些作用涉及蛋白激酶的激活减少和转录因子NFKappaBalpha。此外，通过细胞活力测定法和确定培养的神经元和脑微血管内皮细胞中炎症因子的测定清楚地确定了sartans神经保护作用和抗炎作用。 我们假设萨坦的主要抗炎和神经保护作用可能不是AT1受体抑制作用的独特结果。 在人循环的单核细胞中，表达很少的AT1受体的细胞，sartans的抗炎作用部分取决于过氧化物酶体增殖物激活的受体伽马（Ppargamma）激活。 我们发现，某些sartans可能具有双重的作用机制：与抑制AT1受体有关的抗高血压，抗发育和抗炎作用，以及代谢和抗炎作用，部分原因是直接ppargamma激活的结果。 对啮齿动物炎症模型的研究表明，大脑中的萨尔特人的抗炎作用是广泛的，不仅与经典的大脑靶标有关循环炎症因子（例如下丘脑脑室核和血液脑屏障外的区域）的抗炎作用。 在包括前额叶皮层在内的多个皮质区域中还注意到了减少的炎症，这是控制压力情绪反应的关键调节结构。我们对脑血管内皮细胞中内毒素免受内毒素的保护的证明表明，SARTAN的广泛抗炎作用涉及脑内皮细胞中的AT1受体阻断。这些受体首先在我们的实验室中表征，是调节大脑对炎症和压力反应的基本元素。 此外，萨特人限制了对压力的夸张的激素和交感反应，其影响取决于挑战的类型和强度。例如，虽然sartans完全防止了对隔离的激素和同情反应，但这些化合物能够在炎症应激模型中保留抗炎皮质酮反应。此外，sartans可以防止醛固酮的形成和释放，醛固酮是一种促炎激素，刺激海马中刺激脑矿物皮质激素受体。 我们还发现，在服从急性约束的大鼠中，萨坦给药的抗压力和抗焦虑作用包括预防压力的特征的皮质γ-氨基丁酸（GABAA）受体的改变。 预防诸如GABAA之类的主要抑制系统的激活减少，解释了萨坦给药的一致性抗焦虑和抗压力。 这些发现突出了AT1受体在控制压力，炎症和情绪中的多重核心作用。 ARB的抗压力效应具有行为相关性。 萨坦给药可阻止LPS引起的疾病行为，其特征是厌食和体重减轻，这是随着全身性炎症而发生的。此外，Sartans消除了全身LPS给药产生的焦虑反应。 在协作研究中，我们正在分析Sartan治疗的抗抑郁作用，以及脑血管紧张素II系统的参与程度，在慢性可变应力的易感性机制中。 我们继续阐明神经保护的机制以及终生萨坦给药产生的寿命的延长。 我们发现大脑中的深刻抗炎作用，包括预防NADPH激活，这是导致年龄依赖性神经毒性活性氧的产生，以及与年龄相关的脑血管重塑的逆转。 我们还发现，萨特人的抗焦虑作用始终存在。 我们的发现至少可以部分解释萨坦疗法对阿尔茨海默氏病的主要有益作用。 我们的工作继续进行了更高级的机械研究，包括使用基因基因敲除小鼠模型，微阵列测定以及在培养细胞中的选择性基因分解和表型 - 响应实验，以及对动物模型的研究来确定sartans和相关分子的精确治疗范围。 我们还有兴趣确定萨坦的AT1受体阻滞依赖性和独立效应的程度。 这些信息对于进一步发现具有更好治疗价值的sartan分子非常有价值。 我们的机械和转化研究将有望导致对大脑疾病的新型治疗。