Development of Brain Organoids to Study the Impact of HIV-1, Drugs of Abuse and Aging on Cognitive Impairment

项目摘要 HIV-1相关的神经炎症和神经毒性导致认知障碍(HIV-1相关 神经认知障碍或手)，即使在抑制性抗逆转录病毒治疗(ART)下也是如此。作为人类 与HIV-1年龄生活在一起，正在发生一种复合效应，与年龄相关的痴呆症增加， 导致神经认知缺陷的复杂网络。这将对医疗保健产生巨大的影响 系统不仅在美国，而且在发展中国家也是如此。手是如何发展的，以及它是如何发展起来的 修饰仍然是个谜，很大程度上是因为研究HIV-1感染和交出 人脑。在过去的几年里，干细胞技术的新发展使 从诱导的多能干细胞(IPSCs)中分化出“脑器官”，这些培养可以 在促进神经外胚层三维扩张的条件下体外生长，在脑器官中 或者是“微型大脑”的形式。大脑有机体是异质的，形成各种大脑区域，包括 前脑腹侧、大脑皮层、海马体、中、后脑边界。它们展示的神经元 有功能并能电刺激，并发育小胶质细胞。这些大脑器官也类似于 人类大脑皮层发育的基因表达水平，并允许深入分析神经网络、细胞 行为、药物筛选、疾病建模和大脑发育的变化。而大脑区域 不同IPSC品系的有机化合物组成不同，区域基因表达模式基本保持不变 可在不同个体之间复制。这些为研究艾滋病病毒感染创造了无与伦比的新机会 大脑。我们建议开发我们的ipsc衍生的器官模型，它将小胶质细胞结合到一个 更好地代表了成年成熟的大脑，可以支持强大的艾滋病毒-1感染。有了这个模型，我们可以测试 不同的病毒是否会导致不同的神经毒性，以及小胶质细胞以外的细胞是否会潜伏 感染了病毒。我们将测试病毒中的单个蛋白质，包括HIV-1Tat如何引起神经疾病 损害，以及ART或药物，如二氢皮质酮A(DCA)，它如何交叉中和TAT活性， 会影响进程。有趣的是，接触TAT还会加强可卡因介导的奖励机制， 这进一步促进了手，揭示了HIV-1感染和药物之间相互作用的复杂网络 滥用药物。我们将确定TAT和可卡因如何在神经损伤中协同作用，并确定是否 DCA可以逆转这一趋势。由于脑器官为HIV-1在大脑中的潜伏期提供了一个模型，我们可以测试 给药DCA，可以“阻断并锁定”任何残留病毒，最后是HIV-1特异性细胞毒T细胞 细胞(CTL)可以清除有机物中的病毒。总之，这些研究有望为我们提供新的见解 手部、TAT介导的神经毒性的发病机制、可卡因的影响以及它们之间的潜在联系 流程。最后，该模型承诺将增加关于HIV-1在大脑中潜伏的令人兴奋的发现，如果它可以实现的话 沉默或被淘汰。

PROJECT ABSTRACT HIV-1 associated neuroinflammation and neurotoxicity lead to cognitive impairments (HIV-1-associated neurocognitive disorders or HAND) even in those under suppressive antiretroviral therapy (ART). As people living with HIV-1 age, a compounding effect is occurring, with age associated dementia added to HAND, leading to a complex web of neurocognitive deficit. This will have tremendous implications for health care systems not only in the USA but also in the developing world. How HAND develops, and how it could be modified remain mysterious, largely because It has been very difficult to study HIV-1 infection and HAND in the human brain. Over the past few years new developments in stem cell technologies have permitted the differentiation of “cerebral organoids” from induced pluripotent stem cells (iPSCs), and these cultures can be grown in vitro in conditions that promote three-dimensional expansion of neuroectoderm, in cerebral organoid or “miniature brain” forms. Cerebral organoids are heterogeneous and form a variety of brain regions, including ventral forebrain, cerebral cortex, hippocampus, and mid- and hindbrain boundary. They exhibit neurons that are functional and capable of electrical excitation, and develop microglia. These brain organoids also resemble human cortical development at the gene expression level, and allow in depth analysis of neural networks, cell behavior, drug screening, disease modeling, and variations in brain development. While brain-region composition varies in organoids from different iPSC lines, regional gene-expression patterns remain largely reproducible across individuals. These create unparalleled new opportunities to study HIV-1 infection of the brain. We propose to develop our iPSC derived organoid model which incorporates microglia, into one that better represents an adult mature brain that can support robust HIV-1 infection. With this model, we can test whether different viruses lead to differential neurotoxicity and if cells other than microglia can be latently infected with virus. We will test how individual proteins from the virus, including HIV-1 Tat causes neurological damage, and how ART or drugs such as Didehydro-Cortistatin A (dCA), which cross-neutralizes Tat activity, affects the process. Interestingly, exposure to Tat also potentiates cocaine-mediated reward mechanisms, which further promotes HAND, revealing a complex web of interactions between HIV-1 infection and drugs of substance abuse. We will determine how Tat and cocaine collaborate in neurological damage and determine if dCA can reverse it. As cerebral organoids provide a model for HIV-1 latency in the brain we can test whether administration of dCA, can “block-and-lock” any residual virus, and finally whether HIV-1 specific cytotoxic T cells (CTL) can eliminate virus in the organoid. Together, these studies promise to provide novel insights into the pathogenesis of HAND, Tat mediated neurotoxicity, effects of cocaine, and the potential link between these processes. Finally, the model promises to add exciting findings on HIV-1 latency in the brain, and if it can be silenced or eliminated.