Neuroprotective Engineering Based on Innate Responses to Stroke

Pi：Liu，Shu Q.提案编号：1403036机构：西北大学题目：基于中风先天反应的神经保护工程中风是一种常见的疾病，通常由动脉斑块阻塞流向大脑，导致脑损伤、抑郁、智力低下和/或瘫痪。受伤的大脑通常与骨样结构形成有关，即所谓的脑钙化，这是一个扰乱大脑结构并加剧脑损伤的过程。到目前为止，人们对中风是如何导致脑钙化的了解仍然很少，也没有什么有效的方法来预防脑钙化。在这项应用中，研究人员打算阐明一种与细胞膜相关的钙携带分子家族--膜联蛋白在中风小鼠模型脑钙化诱导中的作用。当脑细胞受到损伤时，这些分子可能会从细胞膜移动到细胞内的收缩细丝，导致钙沉积或钙化，因为这些分子携带钙离子。研究人员发现了一种由肝脏产生的分子，称为三叶因子3，它可以通过阻止膜联蛋白的沉积来潜在地保护受损的大脑免受钙化的影响。这一发现的意义在于，三叶因子3可能被潜在地用作预防中风患者脑钙化和损伤的药物。在这个项目中，研究人员将开发一种工程策略，通过传递三叶因子3基因或蛋白质来促进三叶因子3在中风小鼠模型中的产生，并测试该工程方法对防止依赖膜联蛋白的钙化的脑保护的有效性。如果成功，可以通过生物技术方法生产三叶因子3，并将其应用于人类中风患者，以防止脑钙化，从而减少脑损伤和功能缺陷。这一努力可能会降低由中风引起的人类发病率和死亡率。除了这些科学方面的内容外，研究人员还将致力于建立一种将自主研究融入讲座主题的本科教育新模式。这种教育形式将让学生从实践中理解科学概念，更多地参与前沿研究，提高学生的创造力和解决现实世界问题的能力。脑缺血或缺血性中风是一种常见的疾病，通常由脑动脉血栓和/或动脉粥样硬化引起，导致脑损伤和神经功能障碍，包括抑郁、智力低下和/或瘫痪。缺血性中风通常与脑钙化或羟基磷灰石沉积有关，这是一个扰乱神经元结构并加剧脑损伤的过程。到目前为止，脑钙化的机制仍然难以捉摸，几乎没有建立起保护大脑免受钙化的方法。研究人员发现，在缺血神经元中，与细胞膜相关的钙携带分子家族--膜联蛋白可能会从细胞膜转移到细胞骨架微丝，以促进羟基磷灰石的形成。此外，肝脏产生的内分泌分子三叶因子3(TFF3)在中风时上调，通过阻止膜联蛋白沉积来潜在地保护缺血大脑免受钙化。在拟议的研究中，研究人员打算实现三个目标：(1)评估Annexins A2、A3和A5在缺血性脑钙化和损伤中的作用；(2)评估TFF3在保护缺血大脑免受Annexin依赖的钙化和损伤中的作用；以及(3)建立基于TFF3作用机制的神经保护工程策略，以最大限度地保护中风患者的脑钙化。在缺血性中风小鼠模型中，将通过使用siRNA介导的膜联蛋白丢失和基于重组膜联蛋白的获得膜联蛋白的方法来评估膜联蛋白的作用；将使用TFF3-/-小鼠模型来测试TFF3的抗钙化作用，无论是否使用重组TFF3；以及在存在或不存在TFF3的情况下，通过分子结合分析来评估TFF3在干扰膜联蛋白与神经元微丝结合中的作用。基于TFF3基因转染和受控的TFF3蛋白传递技术，将建立保护性的工程方法，以促进缺血性卒中后TFF3的表达。这些研究将为了解缺血性脑钙化的机制和建立预防缺血性脑钙化和损伤的工程技术奠定基础，从而潜在地降低由中风引起的人类发病率和死亡率。

PI: Liu, Shu Q. Proposal Number: 1403036 Institution: Northwestern UniversityTitle: Neuroprotective Engineering Based on Innate Responses to StrokeStroke is a prevalent disorder commonly caused by arterial plaques that block blood flow to the brain, resulting in brain injury, depression, mental retardation, and/or paralysis. The injured brain is often associated with bone-like structure formation, known as brain calcification, a process disrupting the brain structure and intensifying brain injury. To date, it remains poorly understood how stroke causes brain calcification and there are few approaches effective for prevention of brain calcification. In this application, the investigators intend to elucidate the role of a cell membrane-associated family of calcium-carrying molecules known as annexins in the induction of brain calcification in a mouse model of stroke. These molecules may move from the cell membrane to the intracellular contractile filaments when brain cells are injured to cause calcium deposition or calcification, as these molecules carry calcium ions. The investigators have discovered a liver-produced molecule known as trefoil factor 3 that potentially protects the injured brain from calcification by blocking annexin deposition. The significance of this discovery is that trefoil factor 3 may be potentially used as a drug to prevent brain calcification and injury in patients with stroke. In this project, the investigators will develop an engineering strategy for boosting trefoil factor 3 production in a mouse model of stroke by delivery of the trefoil factor 3 gene or protein and test the efficacy of the engineering approach for brain protection against annexin-dependent calcification. If successful, trefoil factor 3 can be produced by a biotechnology approach and applied to human patients with stroke to prevent brain calcification, thereby reducing brain injury and functional deficits. This effort may potentially lead to a reduction in stroke-induced human morbidity and mortality. In addition to these scientific aspects, the investigators will devote efforts to establish a new undergraduate education model integrating independent research into lecture topics. This form of education will allow students to understand scientific concepts from hands-on experience and to be more engaged in cutting-edge research, enhancing students creativity and capability of solving real-world problems. Cerebral ischemia or ischemic stroke is a prevalent disorder commonly caused by cerebral artery thrombosis and/or atherosclerosis, resulting in cerebral injury and neurological deficits including depression, mental retardation, and/or paralysis. Ischemic stroke is often associated with cerebral calcification or hydroxyapatite deposition, a process disrupting neuronal structure and intensifying cerebral injury. To date, the mechanisms of cerebral calcification remain elusive and few approaches have been established for protecting the cerebrum from calcification. The investigators have found that a cell membrane-associated family of calcium-carrying molecules known as annexins may translocate from the cell membrane to the cytoskeletal microfilaments in ischemic neurons to facilitate hydroxyapatite formation. Furthermore, a liver-produced endocrine molecule known as trefoil factor 3 (TFF3) is upregulated in response to stroke, potentially protecting the ischemic cerebrum from calcification by blocking annexin deposition. In the proposed research, the investigators intend to achieve three aims: (1) evaluate the role of annexins A2, A3, and A5 in ischemic cerebral calcification and injury; (2) assess the role of TFF3 in protection of the ischemic cerebrum from annexin-dependent calcification and injury; and (3) establish neuroprotective engineering strategies based on the mechanisms of TFF3 action for maximizing protection against cerebral calcification in stroke. In a mouse model of ischemic stroke, the role of annexins will be evaluated by using siRNA-mediated loss-of-annexin and recombinant annexin-based gain-of-annexin approaches; the anti-calcification role of TFF3 will be tested by using a TFF3-/- mouse model with or without recombinant TFF3 administration; and the role of TFF3 in interference with annexin binding to neuronal micro-filaments will be assessed by molecular binding assays in the presence or absence of TFF3. Protective engineering approaches will be established based on TFF3 gene transfection and controlled TFF3 protein delivery technologies to boost TFF3 expression following ischemic stroke. These investigations will provide a foundation for understanding the mechanisms of ischemic cerebral calcification and establishing engineering technologies for protection against ischemic cerebral calcification and injury, thus potentially reducing stroke-induced human morbidity and mortality.