Neuronal Survival, HIV-1 and Astrocyte-TIMP-1

描述（由申请人提供）：星形胶质细胞产生金属蛋白酶组织抑制剂（TIMP）-1在中枢神经系统稳态和炎症性疾病（如HIV-1相关痴呆（HAD））中起重要作用。TIMP-1最初是作为基质金属蛋白酶（MMP）的抑制剂被发现的，最近，TIMP-1已经成为促进细胞存活的多功能分子。MMP/TIMP失衡与几种神经炎性疾病有关。我们以前的研究独特地证明了HAD患者脑中TIMP-1水平降低。星形胶质细胞-TIMP-1表达在急性和慢性炎症性疾病中受到不同的调节，急性炎症性疾病中的水平升高，慢性炎症性疾病中的水平降低。这些观察结果表明，TIMP-1补充HAD可能具有治疗价值。我们认为星形胶质细胞TIMP-1通过神经营养活性和/或通过激活抗凋亡蛋白来调节HAD中神经元的存活。TIMP-1神经保护可能涉及或可能不涉及MMP。为了理解HAD中TIMP-1调节和神经保护的机制，提出了以下问题： 1. TIMP-1在免疫激活的星形胶质细胞中是如何在分子水平上调节的，TIMP-1表达和慢性胶质细胞激活之间的时间关系是什么？ 2. TIMP-1介导神经元存活的机制是什么？TIMP-1是否通过与MMP相互作用介导神经保护作用？ 3. TIMP-1能否作为治疗剂减轻HIV-1介导的神经退行性变？为了解决这些问题，将利用HAD的细胞和动物模型来模拟急性和慢性炎症性疾病过程。将使用由原代人星形胶质细胞、人神经元、神经毒素、病毒和病毒蛋白组成的实验室体外系统。分子操作如TIMP-1启动子-荧光素酶报告基因构建体、腺病毒感染和siRNA分子的基因沉默将用于检查TIMP-1启动子元件和参与astrocvte-TIMP-1调节和神经保护作用的信号转导途径。将评价TIMP-1的神经保护机制，包括诱导抗凋亡途径和神经营养活性。将使用具有不同MMP结合能力的TIMP-1突变体研究这些现象的MMP依赖性。为了检查TIMP-1是否可以减弱神经变性，将使用颅内或脑室内注射将腺病毒表达的TIMP-1递送至HIV脑炎SCID小鼠模型中的损伤区域。这些研究将揭示HAD中的神经胶质炎症反应，揭示神经胶质-神经元相互作用的新机制，并可能揭示未来神经退行性疾病的潜在治疗策略。

DESCRIPTION (provided by applicant): Astrocyte production of tissue inhibitor of metalloproteinase (TIMP)-1 plays an important role in central nervous system homeostasis and inflammatory diseases such as HIV-1-associated dementia (HAD). While originally discovered as an inhibitor of matrix metalloproteinases (MMPs), recently, TIMP-1 has emerged as a multifunctional molecule promoting cell survival. MMP/TIMP imbalance is implicated in several neuro-inflammatory diseases. Our previous studies uniquely demonstrate that HAD patients have reduced levels of TIMP-1 in the brain. Astrocyte-TIMP-1 expression is differentially regulated in acute and chronic inflammatory conditions with elevated levels in acute and diminished levels in chronic. These observations indicate that TIMP-1 replenishment in HAD may have therapeutic value. We propose that astrocyte-TIMP-1 regulates neuronal survival in HAD, either through neurotrophic activities and/or via activation of anti-apoptotic proteins. TIMP-1 neuroprotection may or may not involve MMPs. To understand the mechanisms of TIMP-1 regulation and neuroprotection in HAD, the following questions are proposed: 1. How is TIMP-1 regulated in immune-activated astrocytes at the molecular level and what is the temporal relationship between TIMP-1 expression and chronic glial activation? 2. What is the mechanism of TIMP-1-mediated neuronal survival? Is TIMP-1 neuroprotection mediated through interactions with MMP? 3. Can TIMP-1 be utilized as a therapeutic agent to attenuate HIV-1-mediated neurodegeneration? To address these questions, cellular and animal models for HAD will be utilized to mimic acute and chronic inflammatory disease processes. Laboratory in vitro systems consisting of primary human astrocytes, human neurons, neurotoxins, virus and viral proteins will be used. Molecular manipulations such as TIMP-1 promoter-luciferase reporter constructs, adenoviral infections and gene silencing with siRNA molecules will be used to examine TIMP-1 promoter elements and signal transduction pathways involved in astrocvte-TIMP-1 regulation and neuroprotective effects. Neuroprotective mechanisms of TIMP-1 including induction of anti-apoptotic pathways and neurotrophic activities will be evaluated. MMP-dependence of these phenomena will be investigated using TIMP-1 mutants with differential MMP binding abilities. To examine if TIMP-1 can attenuate neurodegeneration, adenoviral-expressed TIMP-1 will be delivered to areas of injury in an HIV encephalitis SCID mouse model using intra-cranial or intra-ventricular injections. These studies will shed light on glial inflammatory responses in HAD, unravel novel mechanisms of glial-neuronal interactions, and may uncover potential future therapeutic strategies for neurodegenerative diseases.