Mechanistic studies on stress, brain inflammation and neuroprotection

我们的第一个具体目标是阐明化合物的有益作用所涉及的机制，直到最近，这些化合物还没有被认为对治疗大脑疾病感兴趣。第二个具体目标是进一步确定这类化合物在治疗脑部疾病方面的益处程度。 我们研究了一组化合物，统称为沙坦，或血管紧张素II AT1受体阻滞剂(ARB)。沙坦是一种具有极高安全边际的联苯衍生物，广泛用于治疗心血管和代谢疾病，因为它们对抗血管紧张素II诱导的血管收缩和病理性细胞生长和纤维化，因为它们减少外周炎症，因为它们改善胰岛素敏感性。在我们最初发现沙坦可以减少高血压引起的脑血管炎症之后，我们后来发现沙坦治疗减少了啮齿动物模型中的脑缺血、压力和焦虑，并延长了寿命。最近，我们发现，沙坦的有益作用包括对大脑外周炎症的负面影响的重大改善。我们的结论是，ARB治疗的主要神经保护作用可能与几种机制有关，我们继续研究以进一步阐明这些机制。 在本财年，我们在澄清沙坦在大脑中的抗炎作用方面取得了进展。我们假设至少部分中枢抗炎和神经保护作用是ARB对脑细胞直接作用的结果。 沙坦的抗炎和神经保护作用(炎症诱导的转录因子核因子kappa轻链增强剂和激活蛋白-1的激活减弱，诱导型一氧化氮合酶、环氧合酶-2和烟酰胺腺嘌呤二核苷酸磷酸酶(NADPH)的表达，减少过量一氧化氮、前列腺素E_2的产生，以及导致脑炎症和神经元损伤的活性氧物种的产生)在脑实质中广泛存在。这表明，沙坦可能会影响多种脑细胞类型。利用小胶质细胞、原代皮质神经元、原代小脑颗粒细胞和脑微血管内皮细胞培养，我们发现ARB可以减轻所有研究类型的细胞的炎症。ARB对细菌内毒素脂多糖(LPS)、过量谷氨酸和促炎细胞因子IL-1β具有神经保护作用。涉及的机制包括几种蛋白激酶的活性降低和转录因子NFkappaBalpha的活性降低。 我们推测，沙坦的主要抗炎和神经保护作用可能不是AT1受体抑制的唯一结果。在人类循环单核细胞中，表达AT1受体的细胞很少，沙坦的抗炎作用部分依赖于过氧化体增殖物激活受体γ(PPARγ)的激活。我们发现一些沙坦可能具有双重作用机制：与抑制AT1受体有关的降压、抗生长和抗炎作用，以及部分由PPARγ直接激活所致的代谢和抗炎作用。我们现在证实，在THP-1细胞、缺乏显著AT1受体表达的大鼠皮质小胶质细胞的原代培养中，以及AT1A受体敲除小鼠的小脑颗粒细胞中，PPARγ激活是ARB效应的主要组成部分。这些结果表明，沙坦的部分有益作用是由于独立于AT1受体刺激的机制。 另一个新发现是，在啮齿动物模型中给予ARB显著保护大脑免受创伤性脑损伤。在该模型中，ARBS可缩小病变范围，减轻神经元损伤，保护神经功能。这是首次证明ARB在创伤性脑损伤中的神经保护作用。 我们的工作将不会继续，因为药理学部分已于2013年6月1日关闭。

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 cardiovascular and metabolic disorders because they antagonize Angiotensin II-induced vasoconstriction and pathological cellular growth and fibrosis, because they reduce peripheral inflammation and because they improve insulin sensitivity. Following our initial finding that sartans decrease hypertension-induced cerebrovascular inflammation, we later discovered that sartan treatment reduces brain ischemia, stress, and anxiety, and increases lifespan in rodent models. More recently, we established that the beneficial effects of sartans include a major amelioration of the negative effects of peripheral inflammation in the brain. Our conclusion was that several mechanisms may be responsible for the major neuroprotective effects of ARB treatment, and we continued studies to further clarify such mechanisms. During the current fiscal year, we advanced on the clarification of the anti-inflammatory effects of sartans in the brain. We hypothesized that at least part of the central anti-inflammatory and neuroprotective effects were the consequence of direct actions of ARBs on brain cells. The anti-inflammatory and neuroprotective effects of sartans (decline in inflammation-induced activation of the transcription factors nuclear factor kappa-light-chain-enhancer of activated B cells (NFkappaBalpha) and activator protein-1 (AP-1), expression of inducible nitric oxide synthase, cyclooxygenase-2 and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, reduction in the production of excess nitric oxide, prostaglandin E2, and reactive oxygen species leading to brain inflammation and neuronal injury) are widespread in the brain parenchyma. This suggested that sartans may influence multiple brain cell types. Using microglia, primary cortical neuron, primary cerebellar granule cell, and cerebral microvascular endothelial cell cultures, we discovered that ARBs ameliorate inflammation in all cell types studied. ARB neuroprotective effects were demonstrated against the bacterial endotoxin lipopolysaccharide (LPS), against excess glutamate and against the pro-inflammatory cytokine IL-1beta. Mechanisms involved include decreased activation of several protein kinases and reduced activation of the transcription factor NFkappaBalpha. 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. We now confirm that participation of PPARgamma activation as a major component of ARB effects in THP-1 cells, in primary cultures of rat cortical microglia devoid of significant AT1 receptor expression, and in cerebellar granule cells from AT1A receptor knock-out mice. These results suggest that part of the beneficial effects of sartans are due to mechanisms independent of AT1 receptor stimulation. An additional novel finding is that ARB administration in a rodent model significantly protects the brain from traumatic brain injury. ARBs decrease lesion size, reduce neuronal injury and protect neurological function in this model. This is the first demonstration of the neuroprotective effect of ARBs in traumatic brain injury. Our work will not continue because the Section on Pharmacology was closed June 1st, 2013.