Extracellular Vesicle and Extracellular RNA Biomarkers of HIV-1 Central Nervous System Pathogenesis and Cigarette Use

摘要 HIV相关的神经认知障碍（HAND）和其他神经系统疾病预计会影响 大多数艾滋病毒感染者在生命的某个时候。这些疾病发生，尽管有效 通过抗逆转录病毒疗法（ART）进行病毒学抑制。然而，很少有人建立生物标记物。回顾 为艾滋病开发的血浆生物标志物（病毒载量和CD 4 + T细胞计数）在发展中至关重要。 艾滋病的治疗模式，加强研究重点神经系统疾病的生物标志物是必要的。 当物质使用障碍（SUD）与以下疾病共病时，改善生物标志物可能特别重要： 艾滋病。在艾滋病毒阳性个体中，吸烟的流行率比一般人群高出数倍。 并有助于炎症和衰老过程，加剧发病机制， 多器官系统有希望的生物标志物类别包括稳定的细胞外RNA（exRNA）， 其一部分被包装到细胞外囊泡（EV）中，所述细胞外囊泡（EV）可以离开组织如脑， 在容易接触到的生物体液中发现这些实体作为神经系统疾病的标志物受到越来越多的关注。 疾病，因为在神经变性中的作用和（对于EV），因为它们可以追溯到一个细胞或 组织起源。两类exRNA因其非凡的稳定性而特别有吸引力： microRNA（miRNAs）和circular RNAs（circRNAs）。已经在HIV疾病和SUD中研究了miRNA， 但不存在于脑组织的EV中，并且关于HIV-1发病机制或SUD中的circRNA几乎一无所知。在 在这个项目中，我们将开发一个HIV-1发病机制和吸烟的生物标志物目录，并确定如何 吸烟可能使生物标志物测试的解释复杂化。已充分表征的存档样品 将测试动物模型和人类群组以鉴定和验证候选生物标志物。我们的专业知识是 HIV-1疾病和动物模型;神经系统疾病的生物标志物; EV技术和生物学;以及 非编码RNA。我们将首先使用一种新的组织EV分离技术来寻找候选分子 脑组织中EV和exRNA的特征（目的1），然后研究脑组织中EV和exRNA的特征（目的1）。 匹配的纵向收集的脑脊液和血浆（目标2）。重要的是，目标3调查 人体样本平行。该项目将解决目前文献中的差距， 通过排除EV和exRNA分析的常见伪影，并通过最前沿的确认， 隔离EV的身份。总的来说，我们假设脑源性EV及其最稳定的exRNA 货物，miRNA和circRNA，作为HIV相关神经系统疾病的生物标志物。由于一些 标记物也将与吸烟相关，将调查烟草使用是否可能干扰 对神经系统疾病生物标志物测量的影响。

ABSTRACT The HIV-associated neurocognitive disorders (HAND) and other neurologic disorders are predicted to affect a majority of HIV-infected individuals at some time during the lifespan. These disorders occur despite effective virologic suppression by antiretroviral therapy (ART). Yet few established biological markers. Recalling that plasma biomarkers developed for AIDS (viral load and CD4+ T cell count) were essential in developing treatment paradigms for AIDS, an intensified research focus on biomarkers of neurologic disease is warranted. Improved biomarkers may be particularly important when substance use disorders (SUDs) are comorbid with HIV disease. Among HIV+ individuals, cigarette smoking is several times more prevalent than in the general population and contributes to inflammation and senescence processes that exacerbate pathogenesis in multiple organ systems. Promising classes of biomarkers include stable extracellular RNAs (exRNAs), a portion of which are packaged into extracellular vesicles (EVs) that can leave tissue such as brain to be found in easily accessed biological fluids. These entities receive increasing attention as markers of neurologic disease because of roles in neurodegeneration and (for EVs) because they can be traced back to a cell or tissue of origin. Two categories of exRNA are particularly attractive because of their extraordinary stability: microRNAs (miRNAs) and circular RNAs (circRNAs). miRNAs have been studied in HIV disease and SUDs, but not in EVs of brain tissue, and almost nothing is known about circRNAs in HIV-1 pathogenesis or SUDs. In this project, we will develop a catalog of biomarkers of HIV-1 pathogenesis and cigarette use and identify how cigarette use may complicate interpretation of biomarker tests. Archived samples from a well characterized animal model and human cohorts will be tested to identify and validate candidate biomarkers. Our expertise is in HIV-1 disease and animal models; biomarkers of neurologic disease; EV techniques and biology; and noncoding RNAs. We will first use a novel tissue EV isolation technique to seek candidate molecular signatures of EVs and exRNA in brain tissue (Aim 1), and then investigate EV and exRNA signatures in matched, longitudinally collected cerebrospinal fluid and blood plasma (Aim 2). Importantly, Aim 3 investigates human samples in parallel. the project will address current gaps in the literature in rigorous and reproducible fashion by ruling out common artifacts of EV and exRNA analysis and through cutting edge confirmation of the identity of isolated EVs. Overall, we hypothesize that brain-derived EVs and their most stable exRNA cargo, miRNAs and circRNAs, serve as biomarkers of HIV-associated neurologic disorders. Since some markers will also be associated with cigarette smoking, tobacco use will be investigated for possible interfering effects on measurement of neurologic disease biomarkers.