Validation of Adenylosuccinate as a Novel Endogenous Pro-Angiogenic Factor in the Brain

成人的脑血管生成需要适应低能量条件，如缺氧和缺血，以及 脑损伤后的康复。脑血管生成减少可能导致脑缺血、神经变性和 衰老过程中的损伤，而血管生成增加与肿瘤的发生有关，其中包括胶质母细胞瘤 其他病症。因此，对成人脑血管生成调控机制的理解既有 基础神经科学和临床/翻译对治疗这些疾病的血管调节的重要性。 目前的基础神经科学项目旨在研究一种以前未被认识到的 通过一种新的内源性促血管生成因子--腺苷琥珀酸酯(AdSucc)调节脑血管生成。 我们的初步研究表明，AdSucc在大脑中具有重要的细胞信号作用 而不仅仅是作为代谢中间代谢物发挥作用。我们第一次展示了AdSucc 在体外和体内的检测中激活血管生成，AdSucc诱导磷脂酶C 胞浆内钙离子依赖性增加，提示有受体依赖性作用。这两种影响 在大脑低能量条件下发现的内源性AdSucc浓度下被激活， 这些情况需要血管生成来进行长期补偿。根据我们的初步数据，我们 中心假说是AdSucc是一种大脑内源性促血管生成因子。我们的长期目标是 更好地了解脑血管生成的代谢调节，并确定AdSucc是否有效 治疗涉及异常血管生成的疾病的靶点。为了检验我们的假设，我们有 一种新的AdSucc合酶条件性基因敲除(KO)小鼠模型的建立与鉴定 这允许对AdSucc衰减进行空间和时间控制。我们的实验室还建立了一个 使用双光子显微镜对活体动物脑血管生成进行量化的微创分析。 我们的中心假设将通过以下具体目标得到验证：1.确定AdSucc在大脑中的作用 慢性低氧条件下的血管生成。我们将使用活体脑血管成像来确定血管生成 野生型和ADSS条件性KO小鼠在常氧和慢性低氧条件下，并与 组织化学和功能分析。2.确定AdSucc在常氧条件下脑血管生成中的作用。 在这个目标中，我们将确定AdSucc是否独立于氧气供应而诱导脑血管生成。 这些目标的成功完成将测试一种调节脑血管生成的新机制。

Adult brain angiogenesis is required for adaptation to low energy conditions e.g. hypoxia and ischemia, and for recovery after brain injury. Decreased brain angiogenesis may cause brain ischemia, neurodegeneration, and damage during aging while increased angiogenesis is linked to tumorigenesis including glioblastoma among other pathologies. Thus, an understanding of the mechanisms regulating adult cerebral angiogenesis has both basic neuroscience and clinical/translational importance to therapeutic vessel modulation for these diseases. The present basic neuroscience project is designed to investigate a previously unrecognized mechanism for brain angiogenesis regulation through a novel endogenous pro-angiogenic factor, adenylosuccinate (AdSucc). Our preliminary studies have revealed that AdSucc has an important cellular signaling role in the brain rather than functioning just as a metabolic intermediate metabolite. We demonstrated, for the first time, that AdSucc activates angiogenesis in assays both in vitro and in vivo, and that AdSucc induces phospholipase C dependent increase in cytoplasmic calcium, suggesting a receptor dependent action. Both of these effects were activated under AdSucc endogenous concentrations found under low energy conditions in the brain, which are conditions that require angiogenesis for long term compensation. Based on our preliminary data, our central hypothesis is that AdSucc is a brain endogenous pro-angiogenic factor. Our long term objective is to better understand metabolic regulation for brain angiogenesis, and determine if AdSucc is a valid therapeutic target for treating conditions where aberrant angiogenesis is involved. To test our hypothesis, we have established and characterized a novel AdSucc synthase (ADSS) conditional knockdown (KO) mouse model that allows for spatial and temporal control of AdSucc attenuation. We have also established in our laboratory a minimally invasive assay to quantify brain angiogenesis in a living animal in vivo using two photon microscopy. Our central hypothesis will be tested via the following Specific Aims: 1. Determine the role of AdSucc in brain angiogenesis under chronic hypoxia. We will use brain vascular imaging in vivo to determine angiogenesis in wild type and ADSS conditional KO mice under normoxia and chronic hypoxia, and confirm data with histochemical and functional assays. 2. Determine the role of AdSucc in brain angiogenesis under normoxia. In this aim, we will determine if AdSucc induces brain angiogenesis independently of oxygen availability. Successful completion of these aims will test a novel mechanism for regulation of brain angiogenesis.