Modeling CNS dynamics in HIV infection and cannabinoids with forebrain organoids

项目总结 艾滋病毒相关神经认知障碍(HAND)在高达50%的艾滋病毒阳性人群中持续存在，通常在 尽管由于有效的抗逆转录病毒疗法(ART)，病毒载量很低或检测不到。小胶质细胞是常住细胞 中枢神经系统(CNS)中的巨噬细胞，在ART中被认为是HIV的持久储存库 治疗过的病人。小胶质细胞分泌一系列影响神经元功能的因子，这些因子 在HIV感染小胶质细胞的过程中，因子发生了变化。除了由以下因素介导的HIV病毒与宿主的相互作用 小胶质细胞，许多外源性因素可以改变这些相互作用或直接影响中枢神经系统本身，使其 难以剖析导致手部神经病理的根本原因。在生理上建立一种- 相关平台在抗逆转录病毒治疗和其他辅助因素的背景下调查艾滋病毒感染的动态，我们 将使用人类诱导的多能干细胞产生人类小胶质细胞，我们可以与我们的 发育中的大脑皮层的特性化3D模型。这个集成的3D大脑器官模型将 概括了许多被认为对手部相关病理有贡献的关键细胞群，包括 谷氨酸能神经元、星形胶质细胞和小胶质细胞。我们将评估脑有机化合物的疗效 通过测量艾滋病毒复制动力学和抑制以及艾滋病毒蛋白表达来模拟艾滋病毒感染 不同的细胞类型(目标1)。然后，我们将测量艾滋病毒对神经元发育和功能的影响 急性暴露后的几个时间点(目标2)。我们将进行形态和电生理检查 在单细胞和群体水平上进行分析和RNA测序，以确定特定细胞类型 对艾滋病毒暴露的转录反应。最后，我们将评估外源性药物对这些因素的影响。 衡量病毒复制和神经功能(目标3)。作为概念验证，我们将测试首选的 ART方案，以及大麻素，在有和没有艾滋病毒暴露的情况下开始剖析其影响 这些共同的细胞属性辅助因素可能有助于HAND的病理基础。成功 这些实验的完成将产生一个经过验证的平台来模拟神经炎症的动力学 使用脑区特异性脑器官与人类小胶质细胞整合的疾病。

PROJECT SUMMARY HIV-Associated Neurocognitive Disorder (HAND) persists in up to 50% of the HIV-positive population, often in spite of low or undetectable viral loads due to effective antiretroviral therapy (ART). Microglia are the resident macrophages in the central nervous system (CNS) and are thought to be a persistent reservoir of HIV in ART treated patients. Microglia secrete a panoply of factors that impact neuronal function and these secreted factors are altered during HIV infection of microglia. In addition to HIV virus-host interactions mediated by microglia, many exogenous factors can modify these interactions or directly impact the CNS itself, making it difficult to dissect the underlying causes of neural pathology leading to HAND. To establish a physiologically- relevant platform to investigate the dynamics of HIV infection in the context of ART and other co-factors, we will use human induced pluripotent stem cells to generate human microglia that we can co-culture with our well-characterized 3D model of the developing cerebral cortex. This integrated 3D cerebral organoid model will recapitulate many of the critical cell populations thought to contribute to HAND-related pathology including glutamatergic neurons, astrocytes, and microglia. We will evaluate the efficacy of the cerebral organoids to model HIV infections by measuring HIV replication kinetics and suppression, as well as HIV protein expression in different cell types (Aim 1). We will then measure the impact of HIV on neuronal development and function at several time points following acute exposure (Aim 2). We will perform morphological and electrophysiological analyses and RNA sequencing at both the single cell and population level to determine the cell type-specific transcriptional responses to HIV exposure. Finally, we will evaluate the impact of exogenous drugs on these measures of viral replication and neuronal function (Aim 3). As a proof-of-concept, we will test a preferred regimen of ART, as well as cannabinoids, both with and without HIV exposure to begin to dissect the impact of these common co-factors on cellular properties that may contribute to pathology underlying HAND. Successful completion of these experiments will result in a validated platform to model the dynamics of neuroinflammatory disorders using brain region-specific cerebral organoids integrated with human microglia.