Research Support Core B: Primary Cell, Biomimetic, and iPSC-derived Cell Models

项目摘要 - 原代细胞、仿生学和 iPSC 衍生细胞模型核心 B 了解可卡因、甲基苯丙胺、大麻和阿片类药物的作用机制 放大 HIV 潜伏期的损害是 CWRU 卓越中心的核心目标 药物使用对艾滋病毒的影响。我们对艾滋病毒潜伏期和持久性的理解是 由于循环中发现的少量潜伏感染细胞使情况变得复杂，困难 从患者那里获取全面的组织样本，缺乏已知的表型 可以区分潜伏感染细胞和未感染细胞的标记，信息有限 关于体内组织储库的行为。 Core B 将提供两种离体原代细胞模型 （目标 1）和高度灵敏的基于核酸的测定法来测量物质使用的影响 on the HIV reservoir (Aim 2) in clinical samples.一个尚未解决的关键问题是每个人如何 该药物可调节多种细胞类型和组织中的潜在 HIV 储存库。原代细胞， 核心 B 团队将为中心提供和开发仿生和 iPSC 衍生细胞模型 研究人员针对三个主要领域建立了创新且信息丰富的细胞和仿生模型 吸毒和艾滋病毒的目标器官——大脑、肠道和免疫系统。类器官衍生 来自组织外植体和诱导多能干细胞 (iPSC) 将用于模拟组织 微环境并扩展检测范围，超出使用单细胞所能实现的范围 离体模型。 Core B 提供的模型将在整个卓越中心使用 评估药物使用对 HIV 潜伏期的影响，也将通过样本进行离体实验 源自接受抗逆转录病毒治疗的药物滥用障碍和艾滋病毒患者。这些 每个模型都将反映控制药物反应的分子机制的多样性 外周 CD4 T 细胞、肠道粘膜 T 细胞的使用和 HIV 潜伏期，以及 来自中枢神经系统的小胶质细胞、神经元和微血管内皮细胞，还将提供 对比体内物质使用与直接细胞生物学影响的独特机会 离体分析。重要的是，Core B 团队将继续开发新的检测方法和方法 研究物质使用对艾滋病毒持续性和潜伏期机制的影响。在 特别是，这项工作包括 (a) 开发单细胞 RNA 诱导测定，(b) 建模 TFH 室，以及 (c) 推进 iPSC 衍生的大脑类器官来研究药物使用和 HIV 中枢神经系统的潜伏期。因此，核心 B 与核心 C 中的分子服务合作，将 提供评估细胞、蛋白质组、微生物、 与物质使用相关的代谢组学和转录组学变化及其对艾滋病毒的影响 Core D 中提供的患者样本中的储存库以及原代细胞培养物和工具 操纵我们的物质使用研究人员无法获得的细胞 机构和全国范围内。

Project Summary - Primary Cell, Biomimetics, and iPSC-derived Cellular Models Core B Understanding the mechanism(s) by which cocaine, methamphetamine, cannabis, and opioids amplify the damage of HIV latency is a central goal of the CWRU Center for Excellence on the Impact of Substance Use on HIV. Our understanding of HIV latency and persistence has been complicated by the small numbers of latently infected cells found in the circulation, the difficulty of obtaining comprehensive sets of tissue samples from patients, the lack of known phenotypic markers that can distinguish latently infected cells from uninfected ones, and limited information about the behavior of tissue reservoirs in vivo. Core B will provide both ex vivo primary cell models (Aim 1) and highly sensitive nucleic acid-based assays to measure the impact of substance use on the HIV reservoir (Aim 2) in clinical samples. A key unresolved issue is how each individual drug modulates the latent HIV reservoir in multiple cell types and tissues. The Primary Cell, Biomimetic, and iPSC-derived Cell Models Core B team will provide and develop for Center investigators innovative and highly informative cellular and biomimetic models for the three main organ targets of drug use and HIV – the brain, gut, and the immune system. Organoids derived from tissue explants and induced pluripotent stem cells (iPSC) will be used to mimic the tissue microenvironment and extend the range of assays beyond what can be achieved using single cell ex vivo models. The models offered by Core B will be used throughout the Center of Excellence to evaluate the impact of drug use on HIV latency and will also be established ex vivo with samples derived from persons with Substance Use Disorder and HIV on anti-retroviral therapy. These models will each reflect the diversity of molecular mechanisms governing the response to drug use and HIV latency in CD4 T cells from the periphery, mucosal T cells from the gut, and microglial, neuronal, and microvascular endothelial cells from the CNS and will also provide unique opportunities to contrast the effects of substance use in vivo with direct cell biological analyses ex vivo. Importantly, the Core B team will continue to develop new assays and methods to study the impact of substance use on the mechanisms of HIV persistence and latency. In particular, this effort includes (a) developing single cell RNA induction assays, (b) modeling the TFH compartment, and (c) advancing iPSC-derived cerebral organoids to study drug use and HIV latency in the CNS. Thus, Core B, in collaboration with the molecular services in Core C, will provide access to novel technologies for evaluating the cellular, proteomic, microbiological, metabolomic, and transcriptomic changes associated with substance use and their impact on HIV reservoirs in patient samples available in Core D and to primary cell cultures and tools to manipulate cells that would not otherwise be available to Substance Use researchers at our institutions and nationwide.