Regulation of HIV latency by microglial-neuronal interactions

小胶质细胞-神经元相互作用对 HIV 潜伏期的调节

• 批准号: 10674037
• 负责人: JONATHAN KARN
• 金额: $ 78.49万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10674037
• 关键词: 3-Dimensional; Biological Models; Brain; CRISPR/Cas technology; CX3CL1 gene; Cells; Cerebrum; Coculture Techniques; Collaborations; Communication; Defect; Development; Drug abuse; Drug usage; Effector Cell; Fractalkine; GABA Receptor; Genetic; Genetic Transcription; Glutamate Receptor; HIV; HIV Infections; HIV Seropositivity; Hand; Human; Imaging technology; Individual; Infection; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Response; Injury; Investigation; Laboratories; Lead; Libraries; Mediating; Memory; Methamphetamine; Microglia; Modeling; Molecular Biology; Morbidity - disease rate; Mutation; Natural Products; Nerve Degeneration; Neurobiology; Neurocognitive; Neuroglia; Neurons; Organoids; Pathway interactions; Patients; Pattern recognition receptor; Pharmaceutical Preparations; Production; Productivity; Purinoceptor; Rattus; Receptor Gene; Receptor Signaling; Recovery; Regulation; Rest; Risk; Role; Signal Pathway; Signal Transduction; Stress; Symptoms; System; T-Lymphocyte; Testing; Therapeutic; United States; Viral; Viral Proteins; Virus; antiretroviral therapy; cell transformation; design; dopaminergic neuron; drug of abuse; experimental study; gamma-Aminobutyric Acid; genome editing; glial activation; induced pluripotent stem cell; inhibitor; knock-down; methamphetamine effect; methamphetamine exposure; migration; mortality; multidisciplinary; neurocognitive disorder; neurotoxic; neurotransmission; novel; pharmacologic; protective effect; receptor; reconstitution; response; small hairpin RNA; substance use; transcriptome sequencing

Summary Over 40% of HIV-positive individuals in the United States engage in substance use. This not only represents a major cause of enhanced morbidity and mortality, but also is associated with increased risks of neurocognitive disorders, such as HAND. Neurons possess refined systems for maintaining constant communication with glia through propagation of “Off” and “On” signals controlling microglial activation states. Using HIV latency models in immortalized human microglial cells (hµglia/HIV), we have shown previously that cellular activation and inflammatory responses induce HIV production. Remarkably, co-culture of productively infected microglia with an excess of healthy neurons leads to viral silencing. We have also shown that hµglia/HIV cells can migrate into brain organoids where they become silenced. However, damaging neurons with a variety of agents, including methamphetamine (METH), a frequently-used abuse substance among HIV-infected individuals, produce reactivation signals for HIV, and this initiates a cycle of microglial activation and further neuronal damage. This cycle of shutdown and reactivation seems to parallel the M1 to M2 transition model of microglial cells, much as HIV latency in T cells is a product of the natural transition of effector cells to resting memory cells. In this proposal, we seek to define the key signals mediating the cycle of viral silencing and reactivation in microglial cells by neurons in the context of iPSC-derived cerebral organoids. This multidisciplinary investigation is designed as a close collaboration between the laboratories of Dr. Jonathan Karn (CWRU, HIV molecular biology), Dr. Anthony Wynshaw-Boris (CWRU, iPSC cells, brain organoids), Dr. Kurt Hauser (VCU, neurobiology and drug abuse), and Dr. Pamela Knapp (VCU, brain organoids). To avoid the limitations of working with transformed cells, we have recently initiated experiments using co-cultures between iPSC-derived cerebral organoids and microglia. Using co-cultures between iPSC-derived cerebral organoids and microglia, we will thoroughly test the hypothesis that the exaggerated responses of HIV-infected microglia to neuronal damage leads to enhanced neurodegeneration. We will also test the hypothesis that exposure to METH, given this background of faulty microglia-neuron crosstalk, enhances HIV replication. Using genome editing approaches, we will identify the specific contribution of “On” and “Off” receptor systems in controlling HIV latency in microglia, and study how METH impacts neuronal-microglial signaling to augment HIV production. In parallel with our genetic investigations, we will also evaluate a number of pharmacological agents against microglial receptors, HIV transcription inhibitors, and mediators of inflammation in order to define therapeutic approaches that might be expected to slow the development of HAND, especially in patients who abuse drugs.

概括 在美国，超过 40% 的艾滋病毒阳性者有药物滥用行为。这 不仅是发病率和死亡率增加的主要原因，而且还与 神经认知障碍（例如手部疾病）的风险增加。神经元拥有精致的 通过传播“关”和“开”来与神经胶质细胞保持持续通信的系统 控制小胶质细胞激活状态的信号。在永生人类中使用 HIV 潜伏期模型 小胶质细胞（hμglia/HIV），我们之前已经证明细胞激活和炎症 反应会诱导艾滋病毒的产生。值得注意的是，有效感染的小胶质细胞与 过多的健康神经元会导致病毒沉默。我们还表明 hμglia/HIV 细胞 可以迁移到大脑类器官中，在那里它们变得沉默。然而，损伤神经元 各种药物，包括甲基苯丙胺 (METH)，一种经常使用的滥用物质 在艾滋病毒感染者中，产生艾滋病毒重新激活信号，这启动了一个循环 小胶质细胞激活和进一步的神经元损伤。这种关闭和重新激活的循环似乎 与小胶质细胞的 M1 到 M2 转变模型相似，就像 T 细胞中的 HIV 潜伏期一样 效应细胞自然转变为静息记忆细胞的产物。在本提案中，我们寻求 定义介导小胶质细胞病毒沉默和重新激活周期的关键信号 由 iPSC 衍生的大脑类器官中的神经元产生。这项多学科调查 是 Jonathan Karn 博士（CWRU、HIV 分子生物学）、Anthony Wynshaw-Boris 博士（CWRU、iPSC 细胞、脑类器官）、Kurt 博士 Hauser（VCU，神经生物学和药物滥用）和 Pamela Knapp 博士（VCU，脑类器官）。到 为了避免使用转化细胞的局限性，我们最近开始了实验 使用 iPSC 衍生的脑类器官和小胶质细胞之间的共培养。使用共培养 在 iPSC 衍生的脑类器官和小胶质细胞之间，我们将彻底检验这一假设 HIV感染的小胶质细胞对神经元损伤的过度反应导致增强 神经变性。鉴于此，我们还将检验接触冰毒的假设 有缺陷的小胶质细胞-神经元串扰的背景，增强了艾滋病毒的复制。使用基因组编辑 方法，我们将确定“开”和“关”受体系统的具体贡献 控制小胶质细胞中的 HIV 潜伏期，并研究 METH 如何影响神经元-小胶质细胞信号传导 增加艾滋病毒的产生。在进行基因研究的同时，我们还将评估 针对小胶质细胞受体、HIV 转录抑制剂的药物数量 炎症介质以确定可能预期的治疗方法 减缓手部疾病的发展，尤其是滥用药物的患者。