Defining the impact of cannabinoids on the HIV reservoir in humanized mice

摘要 大麻（CB）的使用在艾滋病毒感染者（PWH）中很普遍，但它对艾滋病毒感染和潜伏感染的影响 病毒储存库还没有被充分研究。特别是，CB对实体组织中HIV储库的影响 例如大脑是完全未知的。因此，可以仔细控制给药的动物模型 和组织样本进行分析将是重要的，以充分了解如何CB影响艾滋病毒水库 组织中的特征。我们的中心假设是CB诱导免疫抑制细胞状态， 影响病毒转录和组织中HIV储存库的维持。本提案的总体目标是 开发一种在HIV感染期间暴露于CB的人源化小鼠模型， CB的使用会影响组织中HIV宿主的特性。在R61期，HIV感染者的人源化 表达hIL-34以促进小胶质细胞CNS重建的小鼠将暴露于一系列THC剂量 在感染期间，在用抗逆转录病毒疗法（ART）抑制病毒之前和之后。THC的影响 将检查暴露对外周血中病毒血症、病毒抑制、免疫标志物和细胞因子的影响。 在R33阶段，我们将使用该模型来解决三个主要问题。首先，病毒DNA（vDNA）和病毒 已经用THC给药的受感染小鼠的大脑和脾脏中的RNA（vRNA）水平将被降低。 定量以确定THC对这些病毒库的大小和转录活性的影响。 组织中其次，我们将研究THC暴露是否影响体内HIV潜伏期逆转。我们假设 由强效潜伏期逆转剂（LRA）激活的炎症信号通路之间的相互作用 如非经典的NF-kB激动剂AZD 5582和CB使用激活的抗炎通路， 显著改变体内LRA给药的结果。为了验证这一假设，THC或对照暴露于HIV 感染的小鼠将被给予AZD 5582，THC对组织中病毒血症和病毒再活化的影响 将被审查。最后，我们将使用该模型对受感染细胞进行深层单细胞表征 在存在和不存在THC暴露的ART抑制的人源化小鼠的脑和脾中。 将使用组合的单细胞ATACseq/RNAseq方法对脾和脑组织进行分析，以鉴定和分析脾和脑组织。 表征HIV vDNA+或vRNA+细胞，并比较这些细胞的丰度和表型， CB给药小鼠和对照。如果成功的话，这一提议将揭示CB如何影响艾滋病毒的新见解 感染和组织驻留HIV储库的特征。这反过来将指导新的艾滋病毒治疗 为CB使用PWH优化的策略。

Abstract Cannabis (CB) use is prevalent amongst people with HIV (PWH) but its impact on HIV infection and the latent viral reservoir has not been fully examined. In particular, the impact of CB on the HIV reservoir in solid tissues such as the brain is completely unknown. As such, animal models in which dosing can be carefully controlled and tissue samples can be analyzed will be important to fully understand how CB impacts HIV reservoir characteristics in tissues. Our central hypothesis is that CB induces an immunosuppressive cellular state that impacts viral transcription and maintenance of the HIV reservoir in tissues. The overall goal of this proposal is to develop a humanized mouse model of CB exposure during HIV infection and to use this model to learn how CB use affects characteristics of the tissue resident HIV reservoir. In the R61 phase, HIV infected humanized mice expressing hIL-34 to promote microglial CNS reconstitution will be exposed to a range of THC doses during infection, both before and after viral suppression with antiretroviral therapy (ART). The impact of THC exposure on viremia, viral suppression, immune markers and cytokines in peripheral blood will be examined. In the R33 phase, we will use this model to address three major questions. First, viral DNA (vDNA) and viral RNA (vRNA) levels in the brains and spleens of infected mice that have been dosed with THC will be quantified to determine the impact of THC on the size and transcriptional activity of the viral reservoir in these tissues. Second, we will examine whether THC exposure affects HIV latency reversal in vivo. We hypothesize that cross-talk between inflammatory signaling pathways activated by potent latency reversing agents (LRAs) such as the non-canonical NF-kB agonist AZD5582 and anti-inflammatory pathways activated by CB use could significantly alter the outcome of LRA dosing in vivo. To test this hypothesis, THC or control exposed HIV infected mice will be dosed with AZD5582, and the impact of THC on viremia and viral reactivation in tissues will be examined. Finally, we will use this model to perform a deep single cell characterization of infected cells in the brains and spleens of ART-suppressed humanized mice in the presence and absence of THC exposure. Spleen and brain tissues will be profiled using a combined single cell ATACseq/RNAseq method to identify and characterize HIV vDNA+ or vRNA+ cells and to compare the abundance and phenotype of these cells between CB dosed mice and controls. If successful, this proposal will reveal new insights into how CB affects HIV infection and the characteristics of the tissue-resident HIV reservoir. This in turn will guide novel HIV cure strategies that are optimized for CB-using PWH.