Investigation of persistent HIV immune stimulation in lymphoid tissues during therapy as a cause of sustained immune activation

研究治疗期间淋巴组织中持续的 HIV 免疫刺激作为持续免疫激活的原因

• 批准号: 10376189
• 负责人: Timothy W Schacker
• 金额: $ 74.8万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-03 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10376189
• 关键词: Address; Age; Anatomy; Anti-Retroviral Agents; Antigens; B-Lymphocytes; Biological Assay; Blood Circulation; C-reactive protein; CD4 Positive T Lymphocytes; Cells; Coagulation Process; Collection; Cytomegalovirus; Data; Detection; Development; Endothelium; Fibrin fragment D; Foundations; Frequencies; Future; Gene Expression Profile; Genes; Genetic Transcription; HIV; HIV Infections; HIV Seronegativity; Health; Helper-Inducer T-Lymphocyte; Herpesviridae; Human Herpesvirus 4; Image Analysis; Immune; Immune response; Immunization; In Situ Hybridization; Individual; Infection; Interleukin-6; Interruption; Interstitial Nephritis; Investigation; Link; Lymphoid Tissue; Measurement; Measures; Metabolic; Methods; Mononuclear; Morbidity - disease rate; Outcome; Pathology; Patients; Pharmaceutical Preparations; Phenotype; Plasma; Production; Pulmonary Hypertension; RNA Viruses; Research; Rest; Serum Markers; Simplexvirus; Site; Source; Strategic Planning; T-Lymphocyte; T-Lymphocyte Subsets; TNF gene; Technology; Testing; Tissues; Transforming Growth Factor beta; United States National Institutes of Health; Viral; Viral Antigens; Viral Load result; Viral reservoir; Virion; Virus; Virus Diseases; Virus Replication; Work; antibody test; antiretroviral therapy; base; cardiovascular disorder risk; cell envelope; comorbidity; detection limit; experience; genetic signature; high throughput screening; immune activation; immune reconstitution; improved; microbial; mortality; neutralizing antibody; quantitative imaging; transcriptomics; translational approach

Antiretroviral therapy (ART) sufficiently suppresses HIV replication to reduce plasma viral load (pVL) below the limit of detection, but immune activation (IA) is not normalized and the elevated levels of IA markers- IL-6, TNF, TGFß, C reactive protein (CRP), and D-Dimer remain elevated are associated with increased risk for cardiovascular disease, endothelial disfunction and clotting abnormalities, pulmonary hypertension, interstitial nephritis, development of non-AIDS associated malignancies, and CNS abnormalities. Multiple mechanisms have been proposed to explain the persistence of IA under ART including microbial translocation and herpes virus infections (e.g., HSV, CMV, EBV). However, here we propose that HIV itself is a major cause of persistent IA because of ongoing virus production in lymphoid tissues (LT) while on ART. In this revised proposal, we have one specific aim that tests two hypotheses: 1) Sustained IA during ART is driven by production of virions and/or expression of viral antigens in reactivated latently infected cells with or without persistent low-level virus replication; and 2) that persistent low-level virus replication during ART is the result of intracellular concentrations (IC) of antiretroviral (ARV) drugs in LT that do not completely inhibit replication and virus production. For our first hypothesis we will seek direct evidence of correlations between persistent IA and virus production/antigen expression in LT by identification, at the single cell level, of: 1) virus-producing CD4 T cells lacking markers of activation and proliferation that we have previously shown can sustain low levels of virus production; 2) T follicular helper cells (Tfh) in B cell follicles that have recently been shown to be an independent reservoir for viral persistence; and 3) reactivated latently infected T cells producing virus or p24. We propose investigations of these drivers of IA by our validated and highly sensitive in situ hybridization (ISH) methods to detect, phenotype and locate virus (v) RNA+ and virus-producing cells in LT; by a validated highly sensitive high throughput “envelope detection by induced transcription-based sequencing” (EDITS) assay; and a broadly neutralizing antibody (bNab) method to enrich for HIV-envelope (ENV)+ cells. For the hypothesized correlations of the virus drivers with IA, we will measure IA with standard flow-based antibody assays and with single cell transcriptomic analysis of LT mononuclear cells to identify unique gene signatures associated with IA. Our second hypothesis will be tested by quantification of ARV-intracellular concentrations (ICs) in LT and determine the relationships among ARV-ICs and the frequency of detecting the three putative virus drivers of IA. Establishing that HIV itself is a cause of IA would point future developments to fully suppress virus production in the LT reservoir with benefits both in reducing IA and associated pathologies and as an essential component of HIV Cure Strategies.

抗逆转录病毒疗法（ART）充分抑制HIV复制，以将血浆病毒载量（pVL）降低至低于 检测极限，但免疫活化（IA）未正常化，IA标志物- IL-6，TNF， TGF β、C反应蛋白（CRP）和D-二聚体持续升高与以下风险增加相关： 心血管疾病、内皮功能障碍和凝血异常、肺动脉高压、间质性 肾炎、非AIDS相关恶性肿瘤的发展和CNS异常。多种机制 已被提出来解释在ART下IA的持续存在，包括微生物易位和疱疹 病毒感染（例如，HSV、CMV、EBV）。然而，在这里我们提出，艾滋病毒本身是一个主要原因， 持续性IA，因为在ART治疗期间淋巴组织（LT）中持续产生病毒。 根据这项建议，我们有一个具体的目标，测试两个假设：1）ART期间持续IA由以下因素驱动： 在再活化的潜伏感染细胞中产生病毒体和/或表达病毒抗原， 持续的低水平病毒复制;和2）ART期间持续的低水平病毒复制是以下因素的结果 LT中不完全抑制复制的抗逆转录病毒（ARV）药物的细胞内浓度（IC）， 病毒生产。对于我们的第一个假设，我们将寻找持续性IA和 通过在单细胞水平鉴定LT中的病毒产生/抗原表达：1）产生病毒的CD 4 T 缺乏活化和增殖标志物的细胞，我们以前已经证明，可以维持低水平的 病毒产生; 2）B细胞滤泡中的T滤泡辅助细胞（Tfh）最近被证明是一种 病毒持久性的独立储库;和3）再活化产生病毒或p24的潜伏感染的T细胞。 我们建议调查这些驱动程序的IA我们验证和高度敏感的原位杂交（ISH） 方法检测，表型和定位病毒（v）RNA+和病毒生产细胞在LT;通过一个验证的高度 灵敏的高通量“通过基于诱导转录的测序的包膜检测”（EDITS）测定;以及 一种用于富集HIV包膜（ENV）+细胞的广泛中和抗体（bNab）方法。对于假设的 为了确定病毒驱动因子与IA的相关性，我们将用标准的基于流动的抗体测定和用 LT单核细胞的单细胞转录组学分析，以鉴定与 内务部我们的第二个假设将通过定量LT和LT中的ARV细胞内浓度（IC）来检验。 确定ARV-IC之间的关系和检测三种推定病毒驱动程序的频率， 内务部确定艾滋病毒本身是IA的原因将指出未来的发展，以充分抑制病毒 在LT储层中的生产具有减少IA和相关病理的益处， 艾滋病毒治疗战略的组成部分。