Modeling HIV and methamphetamine-induced neuroinflammation in cerebral organoids

联合抗逆转录病毒疗法(CART)显著延长了艾滋病毒感染者的寿命 个人。然而，尽管CART有效，但与艾滋病毒相关的疾病造成了重大损失。艾滋病毒-- 相关的神经认知障碍(手)在高达50%的慢性感染者中发生，尽管 手推车。HAND的发病机制仍在调查中。神经炎症是手部的一个特征， 通过临床研究、尸检研究和动物模型确定。持续或间歇性复制 中枢神经系统中的艾滋病毒可能通过直接作用于受感染的细胞或 通过释放病毒蛋白和炎症介质。使用神经刺激剂，包括 甲基苯丙胺可以加剧手部出现的神经认知能力下降，但其机制 这种合并症的潜在原因尚不清楚。小胶质细胞是大脑中主要的髓样细胞， 在急性感染艾滋病毒或SIV后的早期感染，可作为中枢神经系统的病毒库，以及 都被认为在手的发育过程中起着核心作用。小胶质细胞的致病途径 HIV感染后的激活和功能障碍仍未完全确定。小胶质细胞来源于 诱导多能干细胞(IPSCs)为研究其分子机制提供了独特的机会。 潜在的小胶质细胞激活。IPSC来源的小胶质细胞将被引入脑器官，提供 确定小胶质细胞激活对周围星形胶质细胞、神经元、 和其他细胞。Tetherin是一个宿主限制因素，在组装过程中捕获HIV 并在感染细胞内产生促炎信号级联反应。AIM中的实验 本项目的1将以公正的方式和通过 直接评估Tetherin介导的信号作为小胶质细胞炎症的触发因素的作用。 将使用RNAseq、细胞因子产生和免疫荧光显微镜来定义小胶质细胞 艾滋病毒感染后的激活。在目标2中，我们将把感染艾滋病毒的小胶质细胞引入脑器官。 明确HIV引起的神经炎症和神经元功能障碍的分子基础。单细胞 RNAseq和神经元健康和电生理学的评估将在以下模型中进行 急性感染和ART抑制后的慢性脑部感染。甲基苯丙胺对 在HIV感染的小胶质细胞/器质模型中促进神经炎症和神经元损伤 然后对其进行界定，并确定相关路径。总之，这些研究将为我们提供对 手的发病机制和甲基苯丙胺对神经认知功能下降的潜在贡献。

Combination antiretroviral therapy (cART) has led to dramatic increases in lifespan among HIV-infected individuals. Despite effective cART, however, HIV-associated morbidities exert a significant toll. HIV- associated neurocognitive disorders (HAND) occur in up to 50% of chronically infected individuals despite cART. The pathogenesis of HAND remains under investigation. Neuroinflammation is a hallmark of HAND, as established by clinical studies, autopsy studies, and animal models. Ongoing or intermittent replication of HIV in the CNS is likely to contribute to neuroinflammation through direct effects on the infected cells or through release of viral proteins and inflammatory mediators. Use of neural stimulants including methamphetamine can exacerbate the neurocognitive decline seen in HAND, but the mechanisms underlying this comorbidity are not understood. Microglia are the primary resident myeloid cells of the brain, are infected at early times following acute infection with HIV or SIV, can act as a CNS viral reservoir, and are thought to play a central role in the development of HAND. The pathways responsible for microglial activation and dysfunction following HIV infection remain incompletely defined. Microglia derived from induced pluripotent stem cells (iPSCs) provide a unique opportunity to examine the molecular mechanisms underlying microglial activation. iPSC-derived microglia will be introduced into cerebral organoids, providing the additional opportunity to define the effects of microglial activation on surrounding astrocytes, neurons, and other cells. Tetherin is a host restriction factor that captures HIV during the assembly process in infected cells and generates a proinflammatory signaling cascade within infected cells. Experiments in Aim 1 of this project will evaluate HIV-induced neuroinflammation both in an unbiased way and through a directed evaluation of the role of tetherin-mediated signaling as a trigger of microglial inflammation. RNAseq, cytokine production, and immunofluorescence microscopy will be employed to define microglial activation following HIV infection. In Aim 2, we will introduce HIV-infected microglia into cerebral organoids to define the molecular basis of HIV-induced neuroinflammation and neuronal dysfunction. Single-cell RNAseq and evaluation of neuronal health and electrophysiology will be performed in models representing acute infection and in ART-suppressed, chronic infection of the brain. The potential of methamphetamine to contribute to neuroinflammation and neuronal damage in the HIV-infected microglia/organoid model will then be defined, and the relevant pathways identified. Together, these studies will provide insights into the pathogenesis of HAND and the potential contribution of methamphetamine to neurocognitive decline.