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

描述：HIV-1相关的神经认知障碍(HAND)在抗逆转录病毒治疗后的时代仍然是一个重大的社会负担。在大脑中，基质金属蛋白酶(MMPs)通过其组织抑制物(TIMPs)调节仍然是星形胶质细胞重要的动态平衡功能。在过去的几年里，我们证明了TIMP家族的多功能诱导成员-星形胶质细胞-TIMP-1在神经炎症和手部神经发病机制中发挥着关键作用。我们确定了调控星形胶质细胞产生TIMP-1的关键信号通路，并描述了通过激活神经元促进生存机制来发挥新的TIMP-1神经保护作用。作为发育的组成部分，我们测试了多种纳米配方，并将A5P50标准化为脑递送研究的成功候选者。在这一竞争更新中，我们建议在新的方向上扩展我们的工作，以推进对具有临床相关性的机制的基本见解。具体地说，在这项提案中，我们将检验我们的主要假设，即星形胶质细胞-TIMP-1通过C/EBPMAPK/p38信号通路受上游启动子元件调控(AIM 1)，并通过TIMP-1/TIMP/整合素三部分复合体介导神经保护(AIM 2)。研究将包括启动子-报告基因和过度表达结构，以及关键信号元件的敲除或抑制，以破译这些机制。此外，我们之前优化了纳米配方，这些配方将用于成功地将GFAP限制性基因输送到大脑。在目标3中，我们建议测试A5P50纳米制剂对GFAP驱动的星形胶质细胞TIMP-1在手部环境中的表达。将使用GFAP/TAT转基因动物模型，并将通过尾静脉注射A5P50结合的GFAP Driven-TIMP-1。组织水平的变化将通过RNA、蛋白质和组织病理学的分析来评估，以逆转神经损伤和对细胞外基质的有害副作用。将进行行为研究，以验证临床结果的概念。将采用分子、细胞、体外、体内和翻译相结合的方法。综上所述，本次更新中提出的研究不仅将揭开星形胶质细胞在手部相关神经炎症期间做出反应的新机制，它们还将：提供对Tetraspanin/整合素神经元信号的创新洞察，对涉及基质金属蛋白酶/TIMP失衡的炎症性疾病具有更广泛的影响，并为未来与手部相关的治疗策略获得关键的概念验证数据。

DESCRIPTION: HIV-1-associated neurocognitive disorders (HAND) remain a significant societal burden in the era of post-antiretroviral therapy. In the brain, regulation of matrix metalloproteinases (MMPs), via their tissue inhibitors (TIMPs), remains an important homeostatic function of astrocytes. Over the past few years of funding we demonstrated that astrocyte-TIMP-1, the multifunctional inducible member of the TIMP family plays key roles in neuroinflammation and HAND neuropathogenesis. We identified critical signaling pathways in regulation of TIMP-1 production by astrocytes and described novel TIMP-1 neuroprotective effects through activation of neuronal pro-survival mechanisms. As a developmental component, we tested multiple nanoformulations and standardized A5P50 as a successful candidate for brain delivery studies. In this competing renewal, we propose to extend our works in novel directions for advancing basic insights into the mechanisms with clinical relevance. Specifically, in this proposal, we will test our overarching hypothesis that astrocyte-TIMP-1 is regulated by upstream promoter elements via C/EBPβ-MAPK/p38 signaling (Aim 1) and mediates neuroprotection via a TIMP-1/tetraspanin/integrin tripartite complex (Aim 2). Investigations will include promoter-reporter and overexpression constructs along with the knockdown or inhibition of key signaling elements to decipher these mechanisms. Furthermore, we previously optimized nanoformulations that will be used for successful GFAP-restrictive gene delivery to the brain. In Aim 3 we propose to test the A5P50 nanoformulations of GFAP-driven astrocyte TIMP-1 expression in the setting of HAND. The GFAP/Tat-transgenic animal model will be used and A5P50-conjugated GFAP driven-TIMP-1 will be administered via tail-vein injection. Changes will be evaluated at the tissue level by analysis of RNA, protein and histopathology for reversal of neural damage and deleterious side effects on extracellular matrix. Behavior studies will be performed for proof-of-concept of clinical outcomes. Combined molecular, cellular, in vitro, in vivo and translational approaches will be employed. Taken together, studies proposed in this renewal will not only unravel novel mechanisms underlying astrocyte responses during HAND-associated neuroinflammation, they will: provide innovative insight into the tetraspanin/integrin neuronal signaling, have broader implications for inflammatory diseases that involve MMP/TIMP imbalance and obtain critical proof-of-concept data for future therapeutic strategies relevant to HAND.