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: 10598469
• 负责人: Timothy W Schacker
• 金额: $ 74.15万
• 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/10598469
• 关键词: Address; Age; Anatomy; Anti-Retroviral Agents; Antigens; B-Lymphocytes; Biological Assay; C-reactive protein; CD4 Positive T Lymphocytes; Cells; Circulation; Coagulation Process; Collection; Cytomegalovirus; Data; Detection; Development; Fibrin fragment D; Foundations; Frequencies; Future; Gene Expression Profile; Genes; Genetic Transcription; HIV; HIV Infections; HIV Seronegativity; HIV envelope protein; Health; Helper-Inducer T-Lymphocyte; Herpesviridae; Herpesviridae Infections; Human Herpesvirus 4; Image Analysis; Immune; Immune response; Immunologic Stimulation; 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; Proliferating; Pulmonary Hypertension; RNA; Research; Rest; Serum Markers; Simplexvirus; Site; Source; Strategic Planning; T-Lymphocyte; T-Lymphocyte Subsets; TNF gene; Technology; Testing; Tissues; United States National Institutes of Health; Viral; Viral Antigens; Viral Load result; Viral reservoir; Virion; Virus; Virus Replication; Work; antibody test; antiretroviral therapy; cardiovascular disorder risk; cell envelope; comorbidity; detection limit; endothelial dysfunction; experience; genetic signature; high throughput screening; immune activation; immune reconstitution; improved; microbial; mortality; neutralizing antibody; quantitative imaging; transcriptomics; translational approach; viral detection

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.

抗逆转录病毒治疗 (ART) 充分抑制 HIV 复制，将血浆病毒载量 (pVL) 降低至低于 检测限，但免疫激活 (IA) 未标准化，并且 IA 标记物 - IL-6、TNF、 TGFβ、C 反应蛋白 (CRP) 和 D-二聚体保持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、非艾滋病相关恶性肿瘤的发展以及中枢神经系统异常。多重机制 有人提出解释 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，我们在这里提出，艾滋病毒本身就是造成这种情况的一个主要原因。 持续性 IA 是由于接受 ART 期间淋巴组织 (LT) 中持续产生病毒所致。在本次修订中 提案中，我们有一个具体目标来测试两个假设：1）ART 期间持续的 IA 是由以下因素驱动的： 在重新激活的潜伏感染细胞中产生病毒颗粒和/或表达病毒抗原，有或没有 病毒持续低水平复制； 2) ART 期间持续低水平的病毒复制是由于 LT 中抗逆转录病毒 (ARV) 药物的细胞内浓度 (IC) 不能完全抑制复制和 病毒的产生。对于我们的第一个假设，我们将寻求持续性 IA 和 IA 之间相关性的直接证据。 通过在单细胞水平上鉴定以下内容，在 LT 中产生病毒/抗原表达：1) 产生病毒的 CD4 T 我们之前已经证明，缺乏激活和增殖标记的细胞可以维持低水平的 病毒的产生； 2) B 细胞滤泡中的滤泡辅助 T 细胞 (Tfh) 最近被证明是一种 病毒持久性的独立储存库； 3) 重新激活潜伏感染的 T 细胞，产生病毒或 p24。 我们建议通过经过验证的高灵敏度原位杂交 (ISH) 对 IA 的这些驱动因素进行研究 在 LT 中检测、表型和定位病毒 (v) RNA+ 和病毒产生细胞的方法；由经过高度验证的 灵敏的高通量“基于诱导转录的测序的包膜检测”（EDITS）测定；和 一种广泛中和抗体 (bNab) 方法，用于富集 HIV 包膜 (ENV)+ 细胞。对于假设的 病毒驱动因素与 IA 的相关性，我们将使用基于标准流的抗体测定法和 对 LT 单核细胞进行单细胞转录组分析，以确定与 IA。我们的第二个假设将通过 LT 和 LT 中 ARV 细胞内浓度 (IC) 的量化来检验 确定 ARV-IC 之间的关系以及检测三种假定病毒驱动因素的频率 IA。确定艾滋病毒本身是 IA 的一个原因将为未来的发展指明全面抑制病毒的方向 LT 水库的生产不仅有利于减少 IA 和相关病理，而且是一种重要的 HIV 治疗策略的组成部分。