In situ and digital spatial profiling of the active HIV reservoir in autopsy-derived tissues

尸检组织中活性 HIV 储存库的原位和数字空间分析

• 批准号: 10614019
• 负责人: Timothy Jensen Henrich
• 金额: $ 49.71万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10614019
• 关键词: Acute myocardial infarction; Anatomy; Autopsy; B-Lymphocytes; Binding; Biological Assay; Biological Markers; Biopsy; Brain; CD4 Positive T Lymphocytes; Cardiac; Cell Count; Cell Lineage; Cells; Cessation of life; County; Cryopreserved Tissue; DNA; Data; Drug or chemical Tissue Distribution; Environment; Formalin; Freezing; Frequencies; Funding; Gene Expression; Gene Expression Profiling; Genetic Transcription; HIV; Heart; Hemorrhage; Histologic; Human; Immune; Immunohistochemistry; Immunologics; In Situ; In Situ Hybridization; Individual; Investigation; Length; Liver; Lung; Lymphoid Cell; Lymphoid Tissue; Macrophage; Maintenance; Measures; Medical Examiners; Metabolic Clearance Rate; Morbidity - disease rate; Myelogenous; Myeloid Cells; National Heart, Lung, and Blood Institute; Neuroglia; Neurologic; Nucleic Acids; Organ; Overdose; Paraffin Embedding; Participant; Persons; Pharmaceutical Preparations; Phenotype; Proteins; Proteomics; Proviruses; Pulmonary Embolism; RNA; Recrudescences; Residual state; Resolution; Resources; Role; Sampling; San Francisco; Source; Spleen; Time; Tissue Sample; Tissue-Specific Gene Expression; Tissues; Transcript; Viral; Virus; Visualization; Withdrawal; antiretroviral therapy; aspirate; cohort; comorbidity; digital; experience; immune activation; in vivo; innate immune pathways; innovation; interest; lymph nodes; mortality; nano-string; novel; prospective; protein expression; sudden cardiac death; synergism; targeted treatment; transcriptomics; viral RNA; viral rebound

Project Summary/Abstract (Project 2) The study of HIV rebound potential necessitates rigorous viral and immunological characterization directly within tissues in people with HIV (PWH) on ART with minimal comorbidities. As a result, this project leverages the longitudinal San Francisco POstmortemSystematic InvesTigation of Sudden Cardiac Death (POST SCD) Study, a postmortem study to bank samples and autopsy data on PWH and uninfected controls who were victims of sudden cardiac death (SCD). To date we have collected extensive tissue samples, including brain, multiple lymph node chains, liver, spleen, heart, pulmonary vasculature and other tissues of interest from47 HIV-infected and >500 uninfected individuals who experienced SCD. Importantly, ~80% of HIV+ SCD cases were on ART and died suddenly of non-HIV (i.e. cardiac) causes. As a result, the HIV POST SCD cohort is a one-of-kind resource for the study of tissue HIV persistence. This highly innovative project involves in situ hybridization and cutting-edge tissue-based transcriptomic/proteomic nanoString Digital Spatial Profiling (DSP) to clearly define how the reservoirs of intact and HIV-expressing proviruses and biomarker expression differ between tissues and determine how tissue-specific differences in the transcriptionally active HIV reservoir relate to in situ host cell gene and protein expression. Our central hypothesis is that infected CD4 T cells and various myeloid lineage cells within immune privileged histologic environments express full-length, intact HIV RNA transcripts and be capable of rapid viral recrudescence following ART withdrawal. We expect to observe lower expression of host antiviral factors and higher expression of pro-survival factors in cell clusters expressing HIV transcripts. Furthermore, we posit that transcriptional activity and immune states of reservoir cells that are identified as predictors of viral rebound in Project 3 will be visualized within specific lymphoid tissue regions of interest. Our aims are to: 1) measure the total burden of intact and defective proviruses and HIV transcripts across the full spectrum of different organs and tissues in vivo; 2) compare across tissues the cellular burden and phenotypes of cells that spontaneously transcribe HIV transcripts in SCD victims on and off ART at the time of death; and 3) determine the in situ impact of HIV burden and residual transcriptional activity on host cell factors (particularly antiviral restriction and innate immune pathways) in tissue-resident lymphoid and myeloid cells in SCD victims on ART compared to uninfected controls. As a result, this project will have the capacity to identify targets for therapeutic approaches to achieve HIV cure in synergy with Projects 1 & 3.

项目概要/摘要（项目2） HIV反弹潜力的研究需要直接在体内进行严格的病毒和免疫学表征。 组织中的艾滋病毒感染者（PWH）的ART与最低限度的合并症。因此，该项目利用了 旧金山弗朗西斯科心源性猝死（POST SCD）的纵向系统性研究， 对威尔斯亲王医院和未受感染的对照组的样本库和尸检数据进行了尸检研究， 心源性猝死（SCD）。到目前为止，我们已经收集了大量的组织样本，包括大脑，多个 淋巴结链、肝、脾、心脏、肺血管和其他组织，来自47例HIV感染者 和>500名未感染的患有SCD的个体。重要的是，约80%的艾滋病毒+ SCD病例接受了抗逆转录病毒治疗 突然死于非HIV（即心脏病）原因。因此，HIV SCD后队列是独一无二的 组织HIV持久性研究的资源。这个高度创新的项目涉及原位杂交， 尖端的基于组织的转录组学/蛋白质组学nanoString数字空间分析（DSP）， 完整的和表达HIV的前病毒的储存库和生物标志物的表达在组织之间如何不同， 确定转录活性HIV储库中的组织特异性差异如何与原位宿主细胞相关 基因和蛋白质表达。我们的中心假设是受感染的CD 4 T细胞和各种髓系细胞， 免疫豁免组织学环境中的细胞表达全长、完整的HIV RNA转录物， 在停止抗逆转录病毒治疗后病毒会迅速复发。我们希望观察到宿主的低表达 抗病毒因子和在表达HIV转录物的细胞簇中促存活因子的更高表达。 此外，我们证实，转录活性和免疫状态的水库细胞，被确定为 项目3中病毒反弹的预测因子将在感兴趣的特定淋巴组织区域内可视化。 我们的目标是：1）测量完整和有缺陷的前病毒和HIV转录本在整个组织中的总负担。 体内不同器官和组织的全谱; 2）比较组织间的细胞负荷， SCD患者接受和不接受ART时自发转录HIV转录物的细胞表型 死亡;和3）确定HIV负荷和残余转录活性对宿主细胞因子的原位影响 （特别是抗病毒限制和先天免疫途径）在组织驻留淋巴和骨髓细胞中的作用。 接受抗逆转录病毒治疗的SCD患者与未感染的对照组相比。因此，该项目将有能力确定 与项目1和项目3协同，为实现艾滋病毒治愈的治疗方法制定目标。