Viral dynamics of rebound and reservoir HIV species in IPSC-derived myeloid cells

IPSC 衍生骨髓细胞中反弹和储存 HIV 物种的病毒动态

• 批准号: 10548292
• 负责人: Cagla Akay Espinoza
• 金额: $ 22.81万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10548292
• 关键词: Address; Adherence; Adult; Anatomy; Blood; Cell Lineage; Cells; Cerebrospinal Fluid; Characteristics; Conceptions; Data; Development; Exhibits; Fibroblasts; HIV; Human; Infection; Interferons; Interruption; Knowledge; Lead; Microglia; Modeling; Molecular Cloning; Mus; Myelogenous; Myeloid Cells; Neuraxis; Patients; Phenotype; Resistance; Role; Signal Transduction; Site; Source; Surface; System; T-Lymphocyte; Technology; Testing; Tissues; Toxic effect; Tropism; Up-Regulation; Variant; Viral; Viral reservoir; Viremia; Virus; Virus Replication; antiretroviral therapy; chronic infection; density; follow-up; in vitro Model; in vivo; induced pluripotent stem cell; lymph nodes; macrophage; monocyte; novel strategies; pressure; trafficking; viral DNA; viral rebound

Eradicative HIV cure requires elimination of virus from all tissues, including the central nervous system (CNS), and a thorough understanding of HIV reservoirs that harbor intact and/or replication-competent virus and give rise or support rebound viremia upon treatment interruption is paramount. As long-lived CNS myeloid cells, namely microglia and resident macrophages are an HIV reservoir. While HIV enters the CNS during primary viremia and the predominant HIV variants isolated soon after infection exhibit T-cell tropism, HIV variants that infect cells with lower surface CD4 density, such as microglia, are found later during chronic infection, supported by HIV viral DNA isolated from perivascular and CNS myeloid cells and sustained HIV replication in macrophages independently of T cells in humanized myeloid-only mice. HIV from a T-cell source has also been detected in the cerebrospinal fluid (CSF) of PLWH on suppressive ART. HIV variants detected in the CSF are likely a result of a combination of independent replication of HIV compartmentalized in the CNS in CD4+ T and/or myeloid­ lineage cells and HIV trafficking via T cells or monocytes or as free virus from the periphery. Technical and anatomical constraints limit the comprehensive examination of (i) cellular and viral factors involved in HIV replication in the CNS and (ii) selection pressures that give rise to rebound viruses in the correct cellular context, especially in microglia. Among the reasons are limited availability of freshly isolated primary human microglia, which nonetheless rapidly lose key in vivo features, and the paucity of in vitro models that successfully recapitulate key microglial characteristics. We will use human induced pluripotent stem cell-derived microglia (iMg) differentiated from adult human fibroblasts as a tractable system to examine host and viral determinants governing rebound virus replication in CNS myeloid cells with detailed analyses that are not possible with currently available models. Two recent developments regarding rebound viruses has led to the conception of this proposal. First, in a recent study examining key viral phenotypes in patients undergoing analytical treatment interruption (ATI), our collaborator Dr. Bar found that viruses that rebounded after ATI exhibited higher IFN-1 resistance than any other viruses, including transmitted founder (TF) viruses and reservoir viruses, among viruses isolated from the blood throughout the longitudinal follow-up of patients. The origins of these uniquely IFN-1-resistant rebound viruses remain unclear. Persistent upregulation of IFN-1 signaling in the CNS of PLWH despite suppressive ART raises the possibility that selection pressures might lead to the establishment of a productive reservoir of IFN-1-resistant virus within CNS myeloid-lineage cells. Second, our preliminary data suggest that infectious molecular clones (IMCs) of some rebound viruses identified by Dr. Bar may replicate better than the IMCs of TF viruses in iMg. In this proposal, we will use iMg with a panel of well-characterized IMCs of TF, reservoir, and rebound viruses to test our hypothesis that CNS myeloid cells may be a site for replication of IFN-1-resistant rebound viruses.

根除艾滋病毒治疗需要消除包括中枢神经系统（CNS）在内的所有组织中的病毒，并且彻底了解含有完整和/或具有复制能力的病毒并在治疗中断时引起或支持病毒血症反弹的艾滋病毒储存库至关重要。由于长寿命的中枢神经系统髓样细胞，即小胶质细胞和常驻巨噬细胞是艾滋病毒储存库，虽然艾滋病毒在原发性病毒血症期间进入中枢神经系统，并且感染后不久分离出的主要艾滋病毒变体表现出T细胞趋向性，但艾滋病毒变体表面 CD4 密度较低的感染细胞，如小胶质细胞，在慢性感染过程中后期被发现，得到从血管周围和中枢神经系统骨髓细胞中分离出的 HIV 病毒 DNA 的支持，并且在人源化纯骨髓 HIV 小鼠中，独立于 T 细胞在巨噬细胞中持续进行 HIV 复制。在接受抑制性 ART 的感染者的脑脊液 (CSF) 中也检测到了 T 细胞来源，在 CSF 中检测到的 HIV 变异可能是 HIV 独立复制组合的结果。 CD4+ T 和/或骨髓谱系细胞中的 CNS 以及通过 T 细胞或单核细胞或作为外周游离病毒进行的 HIV 运输限制了对 (i) CNS 中 HIV 复制所涉及的细胞和病毒因素的全面检查。 （ii）选择压力导致病毒在正确的细胞环境中反弹，尤其是在小胶质细胞中，原因之一是新鲜分离的原代人类小胶质细胞的可用性有限，但它们很快就失去了体内的关键特征，并且缺乏体内的病毒。我们将使用从成人成纤维细胞分化而来的人诱导多能干细胞衍生的小胶质细胞（iMg）作为易于处理的系统来检查控制中枢神经系统骨髓细胞中反弹病毒复制的宿主和病毒决定因素。目前可用的模型不可能实现这一点。最近关于反弹病毒的两项发展导致了这一提议的构想。 Bar博士发现，在对患者进行的纵向随访中，从血液中分离出的病毒中，ATI后反弹的病毒比任何其他病毒（包括传播的创始人（TF）病毒和储存病毒）表现出更高的IFN-1抵抗力。尽管抑制性抗逆转录病毒治疗，这些独特的抗干扰素1病毒反弹病毒在感染者中枢神经系统中的持续上调仍不清楚，这增加了选择压力可能导致抗干扰素病毒在体内建立的可能性。其次，我们的初步数据表明，Bar 博士发现的一些反弹病毒的感染性分子克隆 (IMC) 可能比 TF 病毒的 IMC 在 iMg 中复制得更好。一组经过充分表征的 TF、储存病毒和反弹病毒的 IMC，以检验我们的假设，即中枢神经系统骨髓细胞可能是 IFN-1 抗性反弹病毒的复制位点。