A new brain organoid model for NeuroHIV and the impact of opioids

摘要 HIV-1感染中枢神经系统（CNS）的驻留细胞，导致神经发病。艾滋病毒- 神经发病机制可能由直接和间接的病毒和宿主因素引起。然而， 机制尚不清楚。尽管抗逆转录病毒疗法（ART）在抑制艾滋病毒复制方面取得了成功， 近一半的艾滋病毒感染者（PLWH）仍然患有不同程度的艾滋病毒相关神经系统疾病 （手）。也有证据表明，中枢神经系统作为一个艾滋病毒水库和避难所网站，可能允许低 水平的病毒血症，导致持续的神经炎症。HIV潜在的进化分子事件 神经发病机制难以描述，部分原因是缺乏现实的HIV动物模型， 人的脑组织很少能用于研究，直到病人死亡，通常是由于晚期疾病。 人脑皮质类器官（BCO）是研究神经病理学的新兴尖端技术 由于它们的人类起源，它们更好地匹配发育中细胞的基因组和结构特征。 人类大脑与动物模型的比较该模型由自组装动态三维结构组成， 提供了不同细胞类型的相互作用，这在传统的单层培养中是有限的。我们优化 协议，以产生长期可行的和功能性的BCO。我们的BCO模型具有前所未有的细胞类型 多样性，通过从祖细胞到神经元细胞的动态发育， 随着时间的推移，星形胶质细胞处于静止状态;这是一种难以离体获得的表型。与细胞成分， 在适当的功能神经网络的生成，我们的BCO模型显示出强大的细胞外电活动 在早期阶段，并逐步发展成为一个有组织的振荡网络。此外，我们还有 先前建立的用于将iPSC衍生的小胶质细胞整合到BCO中形成类胶质细胞的方法， 这对这项研究至关重要。含有相关免疫细胞类型的BCO类胶质细胞模型将使 艾滋病毒的易感性和研究不同类型的细胞对神经系统的影响， 感染使用这种强大的和功能性的BCO类胶质细胞，我们建议开发一种新的人类模型来研究 HIV神经发病机制的细胞和分子机制，以及潜在的相互作用， 抗逆转录病毒治疗（ART）和阿片类药物暴露的叠加或协同效应。这种注入小胶质细胞的BCO 与内源性星形胶质细胞将允许艾滋病毒感染及其相关的病理生理事件，并有助于 解开相关免疫细胞对神经发病机制的贡献和相互作用。

Abstract HIV-1 infects resident cells of the central nervous system (CNS) leading to neuropathogenesis. HIV- neuropathogenesis is likely caused by direct and indirect viral and host factors. However, the exact underlying mechanisms remain unclear. Despite the success of antiretroviral therapy (ART) in suppressing HIV replication, near half of people living with HIV (PLWH) still have varying degrees of HIV-associated neurological disorders (HAND). There is also evidence that the CNS serves as an HIV reservoir and sanctuary site that may allow low level viremia, contributing to persistent neuroinflammation. The evolving molecular events underlying HIV neuropathogenesis are difficult to delineate, partially due to the lack of realistic HIV animal models and because human brain tissues rarely become available for studies until patients die, often due to advanced diseases. Human brain cortical organoids (BCO) are an emerging, cutting-edge technology for studying neuropathological disorders; because of their human origin, they better match the genomic and structural features of the developing human brain compared to animal models. This model consists of a self-assembled dynamic 3-D structure that provides an interplay of different cell types, which is limited in traditional monolayer cultures. We optimized protocols to generate long-term viable and functional BCO. Our BCO model has an unprecedented cell type diversity, via a dynamic development from progenitor cells to neuronal cells, that become interspersed with quiescent astrocytes over time; a difficult phenotype to obtain ex vivo. With the cellular components for generation of a functional neural network in place, our BCO model shows a robust extracellular electrical activity at early stages and progressively develops into an organized oscillatory network. Additionally, we have previously established methods for integration of iPSC-derived microglia into the BCO forming an assembloid, which is crucial tor this study. A BCO assembloid model containing relevant immune cell types will enable susceptibility to HIV and the study of the contributions of different cell types to the neurological consequences of infection. Using this robust and functional BCO assembloid, we propose to develop a new human model to study the cellular and molecular mechanisms underlying HIV neuropathogenesis, and the potential interactive, additive, or synergistic effects of antiretroviral treatment (ART) and opioid exposure. This microglia-infused BCO with endogenous astrocytes will allow HIV infection and its related pathophysiological events and help to disentangle the contribution and interplay of relevant immune cells to neuropathogenesis.