miR-132, a new player in HIV/neuroAIDS

 描述(由申请人提供)：HIV-1感染中枢神经系统(CNS)发生在大多数艾滋病患者中，并导致各种神经功能障碍和神经病理，通常称为神经艾滋病。小胶质细胞/巨噬细胞和星形胶质细胞是中枢神经系统中HIV-1的主要靶细胞，而神经元很少被HIV-1感染，但在HIV/NeuroAIDS中受影响最多。因此，对HIV/NeuroAIDS的发病机制提出了几种间接机制。其中就有HIV-1Tat蛋白。我们已经证明，在没有HIV-1感染的情况下，TAT的表达足以引起神经行为异常和病理，类似于艾滋病毒/神经艾滋病中提到的大多数。此外，我们还发现TAT激活了星形胶质细胞中胶质纤维酸性蛋白的表达，并促进了星形胶质细胞功能障碍和随后的神经元死亡。在初步研究中，我们发现TAT在星形胶质细胞中的表达诱导miR-132以外切体的形式表达和分泌，并改变轴突生长和神经元存活。重要的是，我们还获得了将脑脊液(CSF)miR-132水平与HIV/NeuroAIDS发病机制联系起来的初步证据。作为我们研究的逻辑延伸，我们建议进一步剖析miR-132在星形胶质细胞介导的TAT神经毒性和HIV/神经艾滋病发病机制中的作用。此外，我们还将确定将脑脊液miR-132水平用作新的HIV/NeuroAIDS生物标志物的可行性。因此，当前提议的基本假设是TAT通过调节miR-132及其靶基因对星形胶质细胞和神经元的功能以及神经元的存活产生不利影响。换句话说，miR-132不仅是一种介质，而且是TAT神经毒性和HIV/NeuroAIDS发病机制的指标。为了验证这一假说，我们建议解决以下相互关联的特定目标：(1)表征TAT诱导星形胶质细胞miR-132表达的机制；(2)确定TAT诱导的miR-132表达对星形胶质细胞的影响；(3)阐明miR-132介导的TAT神经毒性的机制；以及(4)探讨将脑脊液miR-132用作HIV/神经艾滋病生物标志物的可能性。在我们的研究中，我们将使用分子、细胞、生化和遗传学相结合的方法，包括使用原代小鼠星形胶质细胞培养和神经元培养、TAT转基因和miR-132基因敲除小鼠、原代人胎脑培养、脑组织和HIV-1队列的脑脊液样本。所寻求的答案对于理解HIV-1Tat这一关键和普遍存在的蛋白及其在HIV/NeuroAIDS发病机制中的作用具有重要意义。此外，这些答案还将有助于确定艾滋病毒/神经艾滋病生物标记物和制定抗艾滋病毒/神经艾滋病治疗策略。关于HIV-1 TAT、星形胶质细胞的HIV感染、miR-132/靶基因在神经退行性疾病中的作用的大量信息、我们的初步研究结果以及CNS HIV抗逆转录病毒治疗效果研究(CHECT)资源使这些目标的实现成为可能。

 DESCRIPTION (provided by applicant): HIV-1 infection of the central nervous system (CNS) occurs in a majority of AIDS patients and causes a variety of neurologic dysfunction and neuropathologies generally termed neuroAIDS. Microglia/macrophages and astrocytes to a less extent are the main HIV-1 target cells in the CNS, whereas neurons are rarely infected by HIV-1 but mostly affected in HIV/neuroAIDS. Therefore, several indirect mechanisms have been proposed for HIV/neuroAIDS pathogenesis. Among them is HIV-1 Tat protein. We have shown that Tat expression in the absence of HIV-1 infection is sufficient to cause neurobehavioral abnormalities and pathologies similar to most of those noted in HIV/neuroAIDS. Moreover, we have shown that Tat activates glial fibrillary acidic protein expression in astrocytes and contributes astrocyte dysfunction and subsequent neuron death. In the preliminary studies, we have found that Tat expression in astrocytes induces miR-132 expression and secretion in the form of exosomes and alters neurite growth and neuron survival. Importantly, we have also obtained preliminary evidence to link the cerebrospinal fluid (CSF) miR-132 levels to HIV/neuroAIDS pathogenesis. As a logical extension of our studies, we propose to further dissect the miR-132 function in astrocyte-mediated Tat neurotoxicity and HIV/neuroAIDS pathogenesis. Besides, we will determine the feasibility of using the CSF miR-132 level as a novel HIV/neuroAIDS biomarker. Thus, the underlying hypothesis for the current proposal is that Tat adversely affects astrocyte and neuron function and neuronal survival through regulation of miR-132 and its target genes. In other words, miR-132 is not only a mediator but also an indicator of Tat neurotoxicity and HIV/neuroAIDS pathogenesis. To test this hypothesis, we propose to address the following interrelated specific aims: (1) To characterize the mechanisms of Tat-induced miR-132 expression in astrocytes; (2) To determine effects of Tat-induced miR-132 expression on astrocytes; (3) To elucidate the mechanisms of miR-132-mediated Tat neurotoxicity; and (4) To investigate the potential of using CSF miR-132 as an HIV/neuroAIDS biomarker. We will use a combined molecular, cellular, biochemical, and genetic approach including use of primary mouse astrocyte cultures and neuron cultures, Tat transgenic and miR-132 knockout mice, primary human fetal brain cultures, brain tissues and CSF samples of HIV-1 cohorts in our studies. The answers sought have fundamental significance for understanding of this critical and pervasive protein HIV-1 Tat, and its role in HIV/neuroAIDS pathogenesis. In addition, these answers shall also aid in identification of HIV/neuroAIDS biomarkers and development of anti-HIV/neuroAIDS therapeutic strategies. The enormous amount of information available on HIV-1 Tat, HIV infection of astrocytes, and roles of miR-132/target genes in neurodegenerative diseases, the results obtained from our preliminary studies, and the CNS HIV Antiretroviral Therapy Effects Research (CHARTER) resources make accomplishment of these aims practical.