NADPH oxidases in ischemic stroke: cellular sources and pharmacological translation

尽管进行了大量的研究工作，重组组织型纤溶酶原激活剂（rt- PA）仍然是唯一被批准用于治疗急性缺血性中风的化合物，而急性缺血性中风是全球第二大死亡原因。活性氧（ROS）和氧化应激在脑卒中的病理生理中起着至关重要的作用，但它们的确切作用尚不清楚。氧化应激的一个有吸引力的候选来源是NADPH氧化酶，这是唯一已知的以ROS为唯一酶产物的酶家族。NADPH氧化酶是寡聚蛋白复合物，NOX亚基催化整个反应。在啮齿类动物中存在4个NOX基因，在鼠脑中NOX主要在神经元和脉管系统中表达，其中NOX4是最丰富的异构体。该项目的目的是阐明NOX4在缺血性卒中病理生理中的作用，使用基因定义的动物模型。暂时性和永久性脑缺血将应用于构成型和组织特异性NOX4敲除小鼠。梗死动态将通过常规1.5 T磁共振成像（MRI）和组织学来确定。通过多模态超高场MRI (17.6 T)，包括弥散（DWI）和灌注（PWI）加权序列，对半影进行定量评估。此外，通过与德国卡尔斯鲁厄Bruker BioSpin GmbH和澳大利亚墨尔本莫纳什大学Harald Schmidt教授合作，我们可以使用传统技术（双氢乙啶[DHE]和硝基酪氨酸染色）以及电子顺磁共振成像（EPRI）对体内和体外自由基形成进行成像。为了将我们的病理生理发现转化为治疗应用，唯一的特异性NOX抑制剂（VAS2870）是可用的。

Despite tremendous research efforts, recombinant tissue plasminogen activator (rt- PA) is still the only approved compound for the treatment of acute ischemic stroke which represents the second leading cause of death worldwide. Reactive oxygen species (ROS) and oxidative stress are critically involved in the pathophysiology of stroke but their precise role is still unknown. One attractive candidate source for oxidative stress are NADPH oxidases, the only known enzyme family that has ROS as their sole enzymatic product. NADPH oxidases are oligomeric protein complexes, with the NOX subunit catalysing the entire reaction. In rodents 4 NOX genes exist, and in the rodent brain NOX are mainly expressed in neurons and the vasculature with NOX4 being the most abundant isoform. The aim of this project is to elucidate the role of NOX4 in the pathophysiology of ischemic stroke using genetically defined animal models. Transient and permanent cerebral ischemia will be applied to constitutive and tissue specific NOX4 knockout mice. The infarct dynamics will be determined by routine 1.5 T magnetic resonance imaging (MRI) and histology. The penumbra will be quantitatively assessed by using multimodal ultra-high field MRI (17.6 T) including diffusion- (DWI) and perfusion (PWI)-weighted sequences. Furthermore, we can image free radical formation ex vivo and in vivo using conventional techniques (dihydroethidium [DHE] and nitrotyrosine staining) as well as electron paramagnetic resonance imaging (EPRI) through cooperation with Bruker BioSpin GmbH, Karlsruhe, Germany and Prof. Dr. Harald Schmidt, Monash University, Melbourne, Australia. For translating our pathophysiological findings into therapeutic applications, the only specific NOX inhibitor (VAS2870) is available.