HIV Infection and Latency In Astrocytes

摘要 联合抗逆转录病毒疗法(CART)显著延长了艾滋病毒感染者的寿命和 改变了艾滋病毒/艾滋病的面貌。然而，细胞/组织中潜伏的艾滋病毒阻碍了 功能性治愈或完全根除。有限的获得抗逆转录病毒疗法和艾滋病毒建立潜伏感染的能力 使中枢神经系统(CNS)成为独一无二的艾滋病毒蓄水池。小胶质细胞/巨噬细胞是HIV的主要靶细胞 中枢神经系统的感染和ALL可以是生产性的、潜伏的或持续的感染。相比之下，星形胶质细胞被感染，但 以有限的方式；我们对这些细胞作为HIV潜伏库的理解及其在HIV潜伏期中的作用 艾滋病毒/神经艾滋病相当有限。我们最近发现，细胞之间的接触导致成功的艾滋病毒感染 星形胶质细胞和在这些细胞中建立艾滋病毒潜伏期，正在进行的艾滋病毒复制水平极低。在……里面 此外，我们还发现，在星形胶质细胞中表达HIV早期基因TAT可诱导miR-132并下调甲基 CpG结合蛋白2(MeCP2)是一种染色质重塑表观遗传因子，通过miRNA-2引起神经毒性。 含有外切体的。此外，我们已经证明可卡因激活了潜伏感染艾滋病毒的星形胶质细胞中的艾滋病毒复制。 通过miR-132的表达，Tat的表达与星形胶质细胞中HIV潜伏期的建立有关。最后，我们 已有初步数据表明，HIV感染者脑脊液中miR-132表达升高 有轻微认知和运动障碍的受试者。作为我们已发表和初步研究的合乎逻辑的延伸，我们 建议表征星形胶质细胞中HIV感染和潜伏期及其对星形胶质细胞功能和功能的贡献 购物车时代的艾滋病毒/神经艾滋病。这一提议的基本假设是，感染艾滋病毒的星形胶质细胞构成 艾滋病毒潜伏在中枢神经系统中，并直接导致艾滋病毒/神经艾滋病。为了检验这一假设，我们建议 解决以下四个相互关联的具体目标：(1)确定细胞-细胞接触介导的艾滋病毒感染的特征 (2)阐明星形胶质细胞中HIV潜伏的分子机制；(3)确定潜伏的HIV的作用 对星形胶质细胞和神经元的感染；以及(4)确定脑脊液中艾滋病毒在大脑中潜伏的生物标志物。我们将使用 分子、细胞、生化和遗传学相结合的方法，包括使用原代小鼠皮质星形胶质细胞 培养和神经元培养，多西环素诱导的脑特异性HIV TAT转基因小鼠(ITAT)，原代人胎儿 脑/星形胶质细胞培养，脑组织和脑脊液样本HIV-1在我们的研究中。我们预计这项提议 将使我们能够确定星形胶质细胞中HIV潜伏期的调节机制以及这些细胞作为 艾滋病毒储藏者在CART的耳朵和可卡因滥用的背景下。这些发现很可能会通知控制组和 根除中南地区艾滋病毒携带者的战略。关于艾滋病毒感染的大量可用信息 以及中枢神经系统/星形胶质细胞的发病机制和外周的HIV潜伏期以及我们初步的研究结果 学习使这些目标的实现变得切实可行。

ABSTRACT Combination antiretroviral therapy (cART) has considerably prolonged the lifespan of HIV-infected individuals and changed the landscape of the HIV/AIDS disease. However, latent HIV in the cells/tissues has prevented from achieving a functional cure or complete eradication. Limited access to cART and the ability of HIV to establish latent infection have made the central nervous system (CNS) unique HIV reservoirs. Microglia/macrophages are the main target cells for HIV infection in the CNS and all can be productively, latently or persistently infected. In contrast, astrocytes are infected but in a restricted manner; our understanding of these cells as HIV latent reservoirs and their roles of HIV latency in HIV/neuroAIDS is quite limited. We have recently found that cell-cell contact leads to successful HIV infection of astrocytes and establishment of HIV latency in these cells with an extremely low level of ongoing HIV replication. In addition, we have found that expression of HIV early gene Tat in astrocytes induces miR-132 and down-modulates methyl CpG-binding protein 2 (MeCP2), a chromatin-remodeling epigenetic factor and causes neurotoxicity through miRNA- containing exosomes. Moreover, we have shown that cocaine activates HIV replication in HIV latently infected astrocytes through miR-132 expression and that Tat expression is linked to establishment of HIV latency in astrocytes. Lastly, we have obtained the preliminary data that miR-132 expression in the cerebrospinal fluid (CSF) is elevated in HIV-infected subjects with minor cognitive and motor disorder. As a logical extension of our published and preliminary studies, we propose to characterize HIV infection and latency in astrocytes and their contribution to astrocyte function and HIV/neuroAIDS in the era of cART. The underlying hypothesis of this proposal is that HIV-infected astrocytes constitute latent HIV reservoirs in the CNS and directly contribute to HIV/neuroAIDS. To test this hypothesis, we propose to address the following four interrelated specific aims: (1) To characterize cell-cell contact-mediated HIV infection of astrocytes; (2) To elucidate molecular mechanisms of HIV latency in astrocytes; (3) To determine effects of latent HIV infection on astrocytes and neurons; and (4) To identify CSF biomarkers for HIV latency in the brain. We will use a combined molecular, cellular, biochemical, and genetic approach including use of primary mouse cortical astrocyte cultures and neuron cultures, doxycycline-inducible brain-specific HIV Tat transgenic mice (iTat), primary human fetal brain/astrocyte cultures, brain tissues and CSF samples of HIV-1 cohorts in our studies. We anticipate that this proposal will allow us to determine the regulatory mechanisms of HIV latency in astrocytes and the significance of these cells as HIV reservoirs in the ear of cART and in the context of cocaine abuse. The findings will likely inform control and eradication strategies for the HIV reservoirs in the CNS. The enormous amount of information available on HIV infection and pathogenesis in the CNS/astrocytes and HIV latency in the periphery and the results obtained from our preliminary studies make accomplishment of these aims practical.