Cellular and Molecular Mechanisms of HIV Neuropathology

描述（由申请人提供）：这些研究旨在评估细胞周期蛋白在HIV神经发病机制中的作用。我们将专注于激活关键的CDK/RWE 2F-1途径在培养的神经元，以确定潜在参与HIV诱导的神经变性的靶基因。然后将在来自HIV痴呆和非痴呆患者的脑组织样品中分析这些凋亡靶点的表达，以确定它们在体内的相关性。实验已经被设计成收集与趋化因子受体与神经元细胞周期蛋白偶联相关的基本信息，并确定HIV包膜蛋白对Rb/E2 F通路的影响。目的1将剖析神经元培养物中趋化因子受体CXCR 4对Rb和E2 F-1的调节机制，并评估非神经元细胞对SDF-1对神经元作用的贡献。目的2将关注不同性质/结构的gp 120对Rb的影响以及控制p53和E2 F-1活性的途径。目标3将集中在HIV感染患者组织样本中E2 F特异性靶点的表达，并确定细胞周期蛋白改变在神经元变性中的作用。为了解决这些问题，一个定义明确的原代培养系统，其中纯的大鼠中枢神经元的群体可以在存在和不存在的非神经元细胞的情况下进行检查，将被用来辨别直接影响的神经胶质细胞和其他非神经元细胞介导的神经元的趋化因子。然后，将使用人类神经元和神经胶质细胞，以确认人类体外模型中最相关的观察结果。最后，检查脑组织样本的艾滋病毒感染患者和没有HAD，将包括确定我们的体外研究结果的人类病理学的相关性。趋化因子，即SDF-1，和HIV包膜蛋白的表达和参与细胞凋亡和分化的细胞周期组分的活性的影响将通过多学科的方法，包括蛋白质组学，药理学，分子生物学，和新的成像/荧光方法进行研究。拟议的实验将提供新的信息，在艾滋病的神经系统并发症的背景下，趋化因子和神经胶质细胞的相互作用的作用。蛋白质组学方法将帮助我们识别与HIV神经发病机制相关的差异蛋白质谱，这可能导致识别新的疾病生物标志物。

DESCRIPTION (provided by applicant): These studies aim to assess the role of cell cycle proteins in HIV neuropathogenesis. We will focus on the activation of the key CDK/RWE2F-1 pathway in cultured neurons to identify target genes potentially involved in HIV-induced neurodegeneration. The expression of such apoptotic targets will be then analyzed in brain tissue samples from HIV demented and non-demented patients to establish their relevance in vivo. Experiments have been designed to gather fundamental information related to the coupling of chemokine receptors to neuronal cell cycle proteins, and to determine the effect of HIV envelope proteins on the Rb/E2F pathway. Aim 1 will dissect the mechanisms implicated in the regulation of Rb and E2F-1 by the chemokine receptor CXCR4 in neuronal cultures and will evaluate the contribution of non-neuronal cells to the effect of SDF-1 on neurons. Aim 2 will focus on the effect of gp120s of different nature/structure on Rb and the pathways that controls the activity of p53 and E2F-1. Aim 3 will concentrate on the expression of E2F-specific targets in tissue samples from HIV-infected patients and will determine the role of cell cycle protein alterations in neuronal degeneration. To address these issues, a well-defined primary culture system, in which pure populations of rat central neurons can be examined both in the presence and in the absence of non-neuronal cells, will be used to discern direct effects of chemokines on neurons from indirect effects mediated by the glia and other non-neuronal cells. Then, human neurons and glia will be used, to confirm the most relevant observations in a human in vitro model. Finally, examination of brain tissue samples from HIV-infected patients with and without HAD, will be included to determine the relevance of our in vitro findings to the human pathology. The effects of chemokines, namely SDF-1, and HIV envelope proteins on the expression and activity of cell cycle components involved in apoptosis and differentiation will be investigated by a multidisciplinary approach including proteomics, pharmacology, molecular biology, and novel imaging/fluorescence methods. The proposed experiments will provide novel information on the roles of chemokines and neuronal-glia interactions in the context of the neurological complications of AIDS. The proteomic approach will help us recognize differential protein profiles associated with HIV neuropathogenesis, which may lead to the identification of novel disease biomarkers.