Integrative Single Cell isoform and chromatin accessibility Mapping of Chronic Opioid Exposure in Cognitive Brain Areas in HIV

HIV认知脑区慢性阿片类药物暴露的综合单细胞亚型和染色质可及性图谱

• 批准号: 10494078
• 负责人: Teresa A Milner
• 金额: $ 71.56万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10494078
• 关键词: Affect; Animals; Area; Astrocytes; Autopsy; Behavior assessment; Biological Process; Brain; Brain region; Cells; Characteristics; Chromatin; Chronic; Cognitive; Computer Analysis; Computing Methodologies; Data; Data Set; Detection; Drug Exposure; Event; Exons; Exposure to; Functional disorder; Future; Gene Expression; Generations; Genes; Genetic Transcription; Genomics; Goals; HIV; HIV Infections; Health; Hippocampus (Brain); Human; Infection; Knowledge; Literature; Maps; Methods; Microglia; Modality; Modeling; Molecular; Monitor; Neuroglia; Neurons; Nomenclature; Opioid; Opioid Peptide; Opioid Receptor; Oxycodone; Peptide Receptor; Persons; Phenotype; Plant Roots; Poly(A)+ RNA; Prefrontal Cortex; Process; Protein Isoforms; Publishing; Regimen; Reporting; Resolution; Rodent; Signaling Molecule; Site; Slice; Slide; Stress; Tissues; Viral; Work; antiretroviral therapy; behavioral pharmacology; brain cell; brain health; cell type; classical conditioning; epigenome; epigenomics; experimental study; genome-wide; innovation; interest; molecular rearrangement; nonhuman primate; novel; opioid exposure; opioid use disorder; programs; response; sequencing platform; sex; simian human immunodeficiency virus; single cell sequencing; single-cell RNA sequencing; tool; transcriptome; transcriptome sequencing; transcriptomics

Opioid driven exacerbations of neuropathological events and alterations in HIV transcription contributing to HIV associated CNS dysfunction are well-reported. Despite years of continuous suppressive antiretroviral therapy (ART), latent HIV persists and finds sanctuary in many of the same brain regions involved in opioid use disorder (OUD) suggesting interactions between HIV and opioids in brain cells. However, there is a sizeable gap in our knowledge on how OUD impacts cellular responses and viral persistence in HIV-infected brain on ART in humans or relevant model organsims. This proposal seeks to generate topographical data sets and evidence at single cell resolution across the hippocampus and prefrontal cortex (PFC), two brain regions known for predilection for HIV persistence and OUD in non-human primate (NHP) and in post-mortem human brain. These data will provide an unprecedented cellular landscape of multiple modalities that can be harnessed to develop strategies to limit viral persistence and restore and retain optimal brain health in people living with HIV. In our published and preliminary work we have developed innovative single-cell approaches: (A) Single-cell isoform RNA sequencing (ScISOr-Seq), which enables single-cell long-read RNA sequencing of polyadenylated RNAs across thousands of single cells; (B) Slide-isoform sequencing (Sl-ISO-Seq) to spatially locate isoforms in brain slices and (C) a single-cell platform that identifies HIV sequences at single cell level (ScHIV-Seq). In concert these novel sequencing and computational methods, along with scATAC-Seq for chromatin accessibility, will permit the mapping of cellular gene expression, open chromatin regions, isoforms and the detection of HIV across single- cells of hippocampus and PFC. Recent literature supports the presence of HIV in the brain and more specifically in microglia and astrocytes present within the hippocampus and PFC. Importantly, these brain regions are also involved in associative learning processes for OUD. Moreover, our prior studies in rodent hippocampus have laid the groundwork for the proposed studies by establishing the regional and cell-specific distributions of opioid peptides and receptors as well as related signaling molecules, and how these distributions are impacted by sex, stress and opioid-associated learning. In further preliminary studies, we conduct opioid receptor mapping, brain spatial transcriptomics, NHP cognitive behavioral assessment and pharmacological profiling of current ART regimens in tissues. These approaches will provide a comprehensive regional landscape to support our single cell specific phenotypes. We propose an overarching hypothesis that: (i) our new integrated single-cell methods will map single-cell and cell-type specific human and NHP transcriptome and epigenome signatures in the hippocampus and PFC of S/HIV in NHPs and post-mortem human brain; (ii) chronic opioid exposure adds a distinguishable signature to S/HIV infection with long-term ART and defines cell subtypes in which these signatures are rooted; and (iii) these signatures are different from chronic opioid exposure on uninfected brain. These studies further an understanding of molecular mechanisms in HIV and OUD in brain.

阿片类药物导致神经病理学事件的恶化和 HIV 转录的改变，从而导致 HIV 相关的中枢神经系统功能障碍已有充分报道。尽管多年来持续进行抑制性抗逆转录病毒治疗 （ART），潜伏的艾滋病毒持续存在，并在涉及阿片类药物使用障碍的许多相同大脑区域中找到庇护所 （OUD）表明艾滋病毒和阿片类药物在脑细胞中存在相互作用。但我们的差距还很大 关于 OUD 如何影响人类 ART 中感染 HIV 的大脑中的细胞反应和病毒持久性的知识 或相关模型有机体。该提案旨在一次性生成地形数据集和证据 海马体和前额皮质 (PFC) 的细胞分辨率，这两个大脑区域以偏爱 非人类灵长类动物 (NHP) 和死后人类大脑中的 HIV 持久性和 OUD。这些数据将提供 前所未有的多种模式的细胞景观，可以用来制定限制策略 病毒持久性以及恢复和保持艾滋病毒感染者的最佳大脑健康。在我们发布的和 我们开发了创新的单细胞方法的前期工作：（A）单细胞亚型RNA测序 (ScISOr-Seq)，可对数千个多腺苷酸化 RNA 进行单细胞长读长 RNA 测序 单细胞； (B) 载玻片异构体测序 (Sl-ISO-Seq)，用于在脑切片中空间定位异构体；(C) 在单细胞水平上识别 HIV 序列的单细胞平台 (ScHIV-Seq)。配合这些小说 测序和计算方法，以及用于染色质可及性的 scATAC-Seq，将允许 细胞基因表达图谱、开放染色质区域、亚型以及跨单细胞的 HIV 检测 海马和 PFC 细胞。最近的文献支持艾滋病毒在大脑中的存在，更具体地说 存在于海马和 PFC 内的小胶质细胞和星形胶质细胞中。重要的是，这些大脑区域也 参与 OUD 的联想学习过程。此外，我们之前对啮齿动物海马体的研究表明 通过建立阿片类药物的区域和细胞特异性分布，为拟议的研究奠定了基础 肽和受体以及相关信号分子，以及这些分布如何受到性别的影响， 压力和阿片类药物相关的学习。在进一步的初步研究中，我们进行了阿片受体图谱、大脑 当前 ART 的空间转录组学、NHP 认知行为评估和药理学分析 组织中的治疗方案。这些方法将提供全面的区域景观来支持我们的单一 细胞特异性表型。我们提出一个总体假设：（i）我们新的集成单细胞 方法将绘制单细胞和细胞类型特定的人类和 NHP 转录组和表观基因组特征 NHP 和死后人脑中 S/HIV 的海马体和 PFC； (ii) 长期接触阿片类药物会增加 长期 ART 的 S/HIV 感染的可区分特征，并定义了这些细胞亚型 签名已扎根； (iii) 这些特征与未感染大脑长期接触阿片类药物不同。 这些研究进一步加深了对大脑中 HIV 和 OUD 分子机制的理解。