Are HIV-1-Infected Alveolar Macrophages Productive Sites of Viral Persistence?

HIV-1 感染的肺泡巨噬细胞是病毒持续存在的产生场所吗？

• 批准号: 10224994
• 负责人: DAVID G RUSSELL
• 金额: $ 42.76万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10224994
• 关键词: Alveolar Macrophages; Apoptosis; Award; Behavior; Belief; Biology; Blood; Bronchoalveolar Lavage; Cell Line; Cell Lineage; Cell Survival; Cells; Cellular Tropism; Coculture Techniques; Collaborations; Data; Data Set; Development; Embryonic Development; Etiology; Failure; Fluorescent in Situ Hybridization; Frequencies; Gene Expression Profiling; Genetic Transcription; Genome; Goals; HIV; HIV-1; Hospitals; Human; Individual; Infection; Longevity; Lung; Lung Transplantation; Lymphocyte; Maintenance; Malawi; Maps; Messenger RNA; Methods; Mus; Operative Surgical Procedures; Pathway interactions; Penetrance; Peripheral; Peripheral Blood Mononuclear Cell; Phagocytes; Phase; Phenotype; Physiological; Pilot Projects; Play; Population; Protocols documentation; Reporter; Reporting; Resolution; Role; Route; Site; Small Interfering RNA; Sorting - Cell Movement; Source; Testing; Time; Transcript; Transplant Recipients; Tropism; Viral; Viral reservoir; Viremia; Virus; Virus Diseases; Yolk Sac; base; cohort; gain of function; loss of function; macrophage; macrophage product; memory CD4 T lymphocyte; peripheral blood; single cell sequencing; stem cells; transcriptome sequencing; viral detection; viral rebound; volunteer

Project Summary / Abstract Despite several reports of HIV-1-infected alveolar macrophages (AM) in the lungs of HIV-1-infected individuals, the roles played by these cells in the maintenance or persistence of infection remain unresolved. Recent studies in Malawi, conducted on AM from HIV-1-infected individuals that are effectively virally- suppressed by long-term ART, reproducibly detected the presence of HIV-1 mRNA by fluorescent in situ hybridization (FISH). In these studies, we also detected HIV-1 transcripts through single cell sequencing protocols, and have isolated infectious HIV-1 virus from cells recovered by bronchoalveolar lavage from ART- naïve, HIV-1-infected volunteers. The hypothesis we propose to test is that the presence of HIV-1 transcripts in the alveolar macrophages of HIV- infected individuals is indicative of a productive viral infection that has significance for persistence of the virus during ART. R21 Phase: Aim 1. Are HIV-1-infected AM productively infected? We propose co-culture approaches to detect and capture infectious HIV-1 from the AM and peripheral blood of HIV-1-infected volunteers in Malawi. Using permissive, HIV-1-susceptible reporter cells we have already shown that we can acquire infectious virus from ART-naïve individuals in a pilot study on 12 volunteers. We propose expanding this analysis to a larger cohort, including ART-treated, virally-suppressed individuals. R33 Phase: Aim 2. Transcriptional Profiling of Viral and Host Transcripts in HIV-1-Infected Cells. Using methods already established in Malawi, we will generate transcriptional profiles of HIV-infected and uninfected AMs by single-cell Seq-Well and Flow-FISH RNASeq methods to obtain datasets reflecting both single-cell resolution and depth of coverage. We will use these datasets to probe the impact of HIV-1 in cell longevity and to study the cellular tropism of the envs from AM-derived HIV-1 virus. R33 Phase: Aim 3. Perturbation of HIV-1-infected AM Function with Synthetic mRNA and SiRNA. We will use gain-of-function (synthetic mRNA) and loss-of-function (siRNA) approaches to manipulate the phenotype and behavior of HIV-1-infected HMDMs and AMs. The goal is to identify pathways that will drive programmed cell death specifically in those AMs that are HIV-1-infected as a route for selective eradication of this potential viral reservoir. The proposal is based on the contention that AM in virally-suppressed individuals will prove to be productively- infected. The verification of this contention is the major milestone of the R21 period of this phased award.

项目摘要 /摘要 尽管有几个报道了HIV-1感染的HIV-1感染的HIV-1感染的肺泡巨噬细胞（AM） 这些细胞在维持或持续性感染中所扮演的角色尚未解决。 马拉维的最新研究，对AM进行了来自HIV-1感染的个体的AM进行，这些人实际上是实际上是 被长期艺术抑制，可重复检测到荧光原位存在HIV-1 mRNA 杂交（鱼）。在这些研究中，我们还通过单细胞测序检测了HIV-1转录本 方案，并具有从ART-从支气管肺泡灌洗中恢复的细胞中分离出的感染性HIV-1病毒 幼稚，HIV-1感染的志愿者。 我们建议检验的假设是，HIV-肺泡巨噬细胞中HIV-1转录本的存在 感染的个体表明了生产性病毒感染，该病毒感染对病毒的持久性具有重要意义 在艺术期间。 R21阶段：AIM 1。HIV-1感染的AM是否有效感染？ 我们提出了共同培养方法，以检测和捕获AM和外周血感染性HIV-1 在马拉维的HIV-1感染志愿者。使用允许的HIV-1启发性记者单元格，我们已经显示 在一项针对12名志愿者的试点研究中，我们可以从没有艺术的人那里获得传染病。我们建议 将此分析扩展到更大的队列，包括经过艺术治疗的病毒抑制的个体。 R33期：AIM 2。在HIV-1感染细胞中病毒和宿主转录物的转录分析。 使用马拉维已经建立的方法，我们将生成HIV感染的转录概况，并 通过单细胞SEQ-WELL和FLOW-FISH RNASEQ方法的未感染AM，以获得反映两者的数据集 单细胞分辨率和覆盖深度。我们将使用这些数据集探测HIV-1在细胞中的影响 寿命并研究来自AM衍生的HIV-1病毒的ENV的细胞向潮流。 R33期：目标3。与合成mRNA和siRNA的HIV-1感染AM功能的扰动。 我们将使用功能获得（合成mRNA）和功能丧失（siRNA）方法来操纵 HIV-1感染的HMDMS和AMS的表型和行为。目标是确定会驱动的途径 在那些被HIV-1感染的AMS中的程序性细胞死亡中，作为选择性放疗的途径 这种潜在的病毒储层。 该提议是基于以下论点：在几乎被抑制的个体中被证明是有效的 - 已感染。该论点的验证是该分阶段奖励奖的R21时期的主要里程碑。