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艾滋病毒神经发病中的作用。我们将重点研究培养神经元中关键的CDK/RWE2F-1通路的激活，以确定可能与HIV诱导的神经变性有关的靶基因。然后将分析HIV痴呆症患者和非痴呆症患者的脑组织样本中这种凋亡靶点的表达，以确定它们在体内的相关性。实验的目的是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/E2F途径的影响。目的1分析趋化因子受体CXCR4在神经元培养中调节Rb和E2F-1的机制，并评价非神经细胞在SDF-1对神经元影响中的作用。目的2将重点研究不同性质/结构的gp120对Rb的影响以及控制P53和E2F-1活性的途径。目的3将集中在HIV感染患者的组织样本中E2F特异性靶标的表达，并将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，一个定义良好的原代培养系统将被用来区分趋化因子对神经元的直接影响和由胶质细胞和其他非神经细胞介导的间接影响。在该系统中，可以在存在和不存在非神经细胞的情况下检查大鼠中枢神经元的纯群体。然后，将使用人类神经元和神经胶质细胞，以证实在人类体外模型中最相关的观察结果。最后，对患有和不患有HAD的HIV感染患者的脑组织样本进行检查，以确定我们的体外研究结果与人类病理学的相关性。趋化因子SDF-1和HIV包膜蛋白对参与细胞凋亡和分化的细胞周期成分的表达和活性的影响将通过蛋白质组学、药理学、分子生物学和新的成像/荧光方法等多学科方法来研究。拟议的实验将提供有关趋化因子和神经元-神经胶质细胞相互作用在艾滋病神经系统并发症中的作用的新信息。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组，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.