CNS Viral Persistence and Neuropsychiatric Perturbations in HIV: Single cell and Molecular Interrogation

项目摘要 尽管抗逆转录病毒疗法（ART）长期抑制艾滋病毒，但根除或持续缓解艾滋病毒/艾滋病的可能性仍然存在。 感染尚未实现。在外周血单核细胞中仍然可以检测到低水平的HIV DNA。 来自接受ART的HIV感染者（PWH）的PBMC，以及含有可反弹病毒的细胞， 存在于庇护组织部位，包括淋巴结、肠道、生殖道和中枢神经系统 （CNS）。在艾滋病临床试验小组进行的一项研究中，我们最近使用了高度敏感的 在活体供者中进行病毒学检测，以检测脑脊液（CSF）细胞中高达50%的HIV DNA， 条重要的是，那些脑脊液中可检测到HIV DNA的人的整体认知功能比那些 CSF HIV DNA未检出。我们随后发现，在CD 4 + T细胞中， 与同期PBMC相比，来自CSF的细胞，以及包括骨髓细胞在内的非典型细胞系， 可能在CSF中感染。最后，我们已经成功地使用单细胞转录组学来鉴定独特的， PWH患者CSF中的罕见细胞类型与HIV疾病状态相关，并进一步证明了 以鉴定PWH与健康对照中富集的细胞转录物。了解CNS HIV的关键差距 持久性包括CSF中感染的免疫细胞的特征和功能， 与血液中的前病毒相比，CSF中的前病毒是完整的，并且在遗传上是区室化的， 这些特征与长期HIV的神经精神功能有关。由于CSF中存在的细胞数量是 抗逆转录病毒疗法抑制的PWH低，免疫学和病毒学的全面、同时表征 CNS的景观尚未实现。使用优化的腰椎穿刺程序和新的 分子技术，我们已经克服了这些障碍，既严格检查表型， CSF细胞，并彻底表征的大小，稳定性，完整性，和序列多样性的持久性艾滋病毒， 脑脊液与血液相比。我们将使用单细胞技术来测量转录和细胞因子谱 结合完整HIV DNA的新定量和单基因组测序， CNS储层内的新相关性。最重要的是，我们建议严格审查认知 艾滋病毒感染者的功能和心理健康，使用新的NIMH的复杂实施 研究领域标准（RDoC）框架，以及神经精神病学结果的差异如何与 在一个不同的参与者队列中，CNS的特异性免疫学和病毒学特征 神经精神共病的范围。

Project Abstract Despite prolonged suppression of HIV on antiretroviral therapy (ART), eradication or sustained remission of the infection has not been achieved. Low levels of HIV DNA are still detectable in peripheral blood mononuclear cells (PBMC) from people living with HIV (PWH) taking ART, and cells containing rebound-competent virus can reside in sanctuary tissue sites, including lymph nodes, gut, genital tract and the central nervous system (CNS). In a study conducted within the AIDS Clinical Trials Group, we have recently used highly sensitive virologic assays in living donors to detect HIV DNA in cerebrospinal fluid (CSF) cells in up to 50% on long-term ART. Importantly, those with detectable CSF HIV DNA had poorer global cognitive function that those in whom CSF HIV DNA was not detected. We have subsequently shown higher concentrations of HIV DNA in CD4+ T cells from CSF compared to contemporaneous PBMC, and that atypical cell lineages including myeloid cells may be infected in CSF. Finally, we have successfully used single cell transcriptomics to identify unique and rare cell types in the CSF in PWH associated with HIV disease status and have further demonstrated the ability to identify cellular transcripts enriched in PWH versus healthy controls. Critical gaps in understanding CNS HIV persistence include what the characteristics and function of infected immune cells are in CSF, whether HIV proviruses in CSF are intact and genetically compartmentalized compared to proviruses in blood, and how these features relate to neuropsychiatric function of long-term HIV. Since the number of cells present in CSF is low in PWH suppressed on ART, thorough, simultaneous characterization of the immunologic and virologic landscape of the CNS has not been achieved. Using optimized lumbar puncture procedures and novel molecular techniques, we have overcome these obstacles to both rigorously examine the phenotype of CSF cells and thoroughly characterize the size, stability, intactness, and sequence diversity of persistent HIV in CSF compared to blood. We will use single cell technology to measure the transcriptional and cytokine profile of CNS cells combined with novel quantification of intact HIV DNA and single genome sequencing to discover new correlations within the CNS reservoir. Most importantly, we propose to rigorously examine the cognitive function and mental health of people living with HIV using sophisticated implementation of the new NIMH Research Domain Criteria (RDoC) framework, and how differences in neuropsychiatric outcomes relate to specific immunological and virological characteristics of the CNS in a diverse cohort of participants with a range of neuropsychiatric comorbidity.