Synergistic Mechanisms of chronic Innate Immune Activation in Microglia by Opiates and HIV Infection

摘要 持续的免疫激活是体内HIV-1感染的决定性特征，也是进展的驱动力- 爱滋病分期。艾滋病毒携带者的系统免疫激活被假设为占更高的 慢性炎症性疾病的发病率，包括与艾滋病毒相关的神经认知障碍(手)。 重要的是，许多研究表明，患有阿片使用障碍(OUD)的人群 HIV并存感染的发生率较高，这表明潜在的社会经济和神经认知 影响力。这种OUD和HIV的共病也被证明会增加神经炎症并加剧 HIV感染和手部疾病的进展。虽然一些研究已经调查了OUD的交互效应 和少数分离的细胞类型，连接这两种情况的分子机制在很大程度上仍然是 未知，尤其是在原代人类小胶质细胞、神经元和星形胶质细胞中。因此，我们认为这一提议 通过建立原代人类神经元细胞培养和公正、系统的 在阿片类药物和HIV-1感染共同暴露的背景下对这些培养物进行单细胞转录研究。 在这项提议中，我们将建立由诱导多能干细胞(IPSC)来源的3D球体培养 人小胶质细胞、星形胶质细胞和神经元，来自北京理工大学建立的10个独立供体系， 将概括大脑中细胞间相互作用的空间复杂性，并询问艾滋病毒的影响 感染和吗啡暴露对持续天然免疫激活的影响。我们将利用尖端的单细胞 转录组学分析确定小胶质细胞促进促炎性反应的内在途径 HIV和阿片类药物联合暴露的反应及神经退行性变通路对神经元的影响 细胞损伤。最后，来自这些转录研究的信息将被用来向CRISPR/CAS提供信息 选择性地删除ipsc-小胶质细胞中的关键通路以减轻神经炎症的敲除策略。我们 相信这些研究的结果将为未来预防艾滋病毒和阿片类药物的治疗开发提供信息- 在HIV物质使用人群中诱发神经炎症和预防手部发育。

ABSTRACT Persistent immune activation is the defining feature of HIV-1 infection in vivo and a driver of progression to end- stage AIDS. Systemic immune activation in people living with HIV has been hypothesized to account for higher incidence of chronic inflammatory diseases, including HIV-associated neurocognitive disorders (HAND). Importantly, numerous studies have demonstrated that populations living with opioid-use disorder (OUD) have a higher occurrence of comorbid HIV infection, suggesting both potential socio-economic and neurocognitive influence. This comorbidity of OUD and HIV has also been shown to increase neuroinflammation and exacerbate the progression of HIV infection and HAND. While some studies have investigated the interactive effects of OUD and HAND in few cell types in isolation, the molecular mechanisms linking the two conditions is still largely unknown, especially in primary human microglia, neurons and astrocytes. Hence, we believe that this proposal fulfills a critical unmet need by establishing primary human neuronal cell cultures and an unbiased, systematic single cell transcriptomic survey of these cultures in the context of co-exposure of opiates and HIV-1 infection. In this proposal, we will establish 3D spheroid cultures consisting of induced pluripotent stem cell (iPSC)-derived human microglia, astrocytes and neurons, derived from 10 independent donor lines established at BUMC, that will recapitulate spatial complexity of cell-to-cell interactions in the brain, and interrogate the effect of HIV infection and morphine exposure on persistent innate immune activation. We will utilize cutting-edge single cell transcriptomics analysis to identify the microglia-intrinsic pathway that promotes induction of pro-inflammatory responses upon co-exposure to HIV and opiates and the neurodegenerative pathways contributing to neuronal cell injury. Finally, information from these transcriptomic studies will be leveraged to inform CRISPR/Cas-based knock-out strategies to selectively delete key pathways in iPSC-microglia to alleviate neuroinflammation. We believe that results from these studies will inform future therapeutic development to prevent HIV and opiate- induced neuroinflammation and prevent development of HAND in HIV+ substance use population.