Characterize replication competent myeloid reservoirs in the central nervous system

表征中枢神经系统中具有复制能力的骨髓库

基本信息

批准号：
10327112
负责人：
Guochun Jiang
金额：
$ 24.76万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-01 至 2023-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10327112
关键词：
Address Animal Model Animals Autopsy Biological Assay Blood Brain Brain region CCAAT-Enhancer-Binding Proteins CD4 Positive T Lymphocytes Cells Cessation of life DNA Data Disease remission Enrollment Environment Epigenetic Process Exposure to Frequencies Generations Genes Gifts Goals HIV HIV Infections HIV-1 Histone Deacetylase Inhibitor Histone Deacetylation Human Immune Immune system In Vitro Infection Interruption Knowledge Macaca Macaca mulatta Measurable Measures Microglia Modification Molecular Myelogenous Myeloid Cells Neuraxis Pathway interactions Penetration Peripheral Play Population Primates Proteins Protocols documentation RNA Regulation Resources Rest Role SIV Sampling Source Study models T-Lymphocyte TLR3 gene Techniques Technology Testing Tissues Universities Viral Viral reservoir Viremia Virus Virus Latency antiretroviral therapy brain cell brain tissue cohort epigenetic regulation histone methylation histone methyltransferase in vivo in vivo imaging inhibitor/antagonist latent HIV reservoir latent infection macrophage monocyte nonhuman primate novel strategies peripheral blood viral rebound virology

项目摘要

SUMMARY The eradication of HIV-1(HIV) infection must address all tissues where infection persists. Clearance of infection in the central nervous system (CNS) is not achieved by antiretroviral therapy (ART) alone, just as in the periphery. Latent HIV reservoirs in the CNS may allow viral rebound upon discontinuation of ART, as HIV can egress from the brain into the peripheral blood. However, studies are needed to elucidate the contributions of subsets of latently infected CNS cells to ongoing HIV persistence in the CNS. Circulating T cells have been well characterized as the major HIV reservoirs in the peripheral blood, and may circulate into the CNS, contributing to HIV persistence in the brain. Nevertheless, it is still not known whether T cells are the only major viral reservoir in the CNS. Myeloid cells are a major cellular compartment of the immune system infected by HIV in the brain. In vivo imaging and immunostaining studies have revealed that brain myeloid cells (BMCs) harbor HIV DNA and produce HIV RNA and proteins. However, it is not clear whether BMCs, and especially long-lived microglia, are latently infected, and if they are true HIV reservoirs encoding replication-competent HIV despite durable, successful ART. Our team at the UNC HIV Cure Center has recently established a protocol to isolate highly pure myeloid cells from the brain of SIV-infected macaques, in whom ART was interrupted. These SIV containing BMCs can be cultured for many generations ex vivo. Further application of this technology to other fully ART-suppressed animals found that BMCs contain proviral SIV DNA, and that SIV RNA can be effectively induced by latency reversal agents. This platform to study myeloid cells ex vivo allows rigorous characterization of brain myeloid cells to address their role as true HIV reservoirs that can produce replication- competent viruses. Our preliminary data highlights that epigenetic regulation, such as histone deacetylation and histone methylation, may play an essential role in the modulation of HIV latency in the CNS. We hypothesize that BMCs are latently infected by HIV and harbor replication competent HIV. In this proposal, we will isolate highly pure brain myeloid cells from different regions of the brain in both people living with HIV (PLWH) enrolled in the “Last Gift” cohort and fully ART-suppressed SIV-infected rhesus macaques. We will examine whether BMCs harbor replication-competent HIV or SIV using the intact proviral DNA assay (IPDA) and quantitative viral outgrowth assay (QVOA) (Aim1). We will examine whether latent HIV in myeloid cells responds to the currently available latency reversal agents related to epigenetic regulation of HIV latency (Aim 2), which may provide clues to the mechanisms that allow HIV latency in the brain. Our study will address critical gaps in knowledge to better understand whether BMCs serve as true HIV reservoirs and how HIV persists in these CNS cells.

概括 HIV-1（HIV）感染的根除必须解决感染持续存在的所有组织。清除单独的抗逆转录病毒疗法（ART）并不能像在中枢神经系统（CNS）中感染（CNS）一样外围。中枢神经系统中的潜在艾滋病毒水库可以在停用艺术时允许病毒反弹，如艾滋病毒可以从大脑流入外周血。但是，需要研究以阐明贡献 CNS中持续的HIV持续性的潜在感染的中枢神经系统细胞的子集。循环T细胞已经被很好地描述为外围血液中主要的HIV储量，并且可能圆满CNS 导致大脑中的艾滋病毒持久性。然而，仍然不知道T细胞是唯一的中枢神经系统中的主要病毒库。髓样细胞是由大脑中HIV感染的免疫系统的主要细胞室。在体内成像和免疫染色研究表明，脑髓样细胞（BMC）携带HIV DNA和产生HIV RNA和蛋白质。但是，尚不清楚BMC，尤其是长寿的小胶质细胞是否是受到潜在感染，如果它们是真正的艾滋病毒水库，编码具有复制能力的HIV目标耐用性，成功的艺术。我们在UNC HIV Cure Center的团队最近建立了一项协议，以高度隔离来自SIV感染猕猴的大脑的纯髓样细胞中断。这些SIV 可以在许多世代培养含有BMC。将该技术进一步应用于其他完全抑制艺术的动物发现BMC含有前病毒SIV DNA，并且可以有效地有效地进行SIV RNA 由潜伏期逆转剂引起。这个研究髓样细胞的平台可以实现严格脑髓样细胞的表征以解决它们作为真正的HIV储存库的作用，可以产生复制 - 合格的病毒。我们的初步数据强调了表观遗传调节，例如组蛋白脱乙酰化 Hisstone甲基化可能在中枢神经系统中的艾滋病毒潜伏期的调节中起着至关重要的作用。我们假设BMC被艾滋病毒和港口复制能力的艾滋病毒感染。在此提案中，我们将把高度纯净的大脑髓样细胞与大脑的不同区域分离出来患有艾滋病毒（PLWH）的人参加了“最后的礼物”队列，并完全抑制了SIV感染的恒河神猕猴。我们将使用完整的前病毒术检查BMCS是否携带复制能力的艾滋病毒或SIV DNA分析（IPDA）和定量病毒出血测定（QVOA）（AIM1）。我们将检查潜在的艾滋病毒在髓样细胞中，对当前可用潜伏期的反向剂反应与表观遗传调节有关艾滋病毒潜伏期（AIM 2），这可能为允许大脑艾滋病毒潜伏期的机制提供线索。我们的研究将解决知识中的关键差距，以更好地了解BMC是否充当真正的艾滋病毒水库和这些CNS细胞中HIV如何持续。