MRS OF NEUROPROTECTION IN AN SIV MODEL OF NEUROAIDS

描述（由申请人提供）：在艾滋病流行早期，HIV引起的神经系统疾病被认为是该疾病的重要临床表现。尽管HAART取得了成功，但艾滋病感染的神经认知并发症仍然是一个重要的问题，而且可能会日益严重。这项研究的长期目标是证明二甲胺四环素是一种耐受性良好、价格低廉的抗生素，可以在神经艾滋病的加速动物模型中预防神经元损伤并促进神经元恢复。二甲胺四环素已经在包括神经艾滋病在内的多种神经疾病中进行了测试。然而，其作用机制尚不清楚。我们的研究旨在提供初步的数据来了解米诺环素的作用机制。我们建议对神经健康、单核/巨噬细胞活化和血液、脑脊液和大脑中的病毒产生进行连续评估，以揭示米诺环素的神经保护、抗炎和/或抗病毒特性。为此，7只siv感染的CD8+细胞耗尽的猕猴将在接种后28天开始接受每日剂量的米诺环素，并使用磁共振波谱（MRS）、免疫学、病毒学和病理学收集数据。他们的结果将与一个未感染的队列、两个SIV感染CD8耗尽的未治疗队列和一个接受抗逆转录病毒联合治疗（CART）的队列进行比较。这四个队列已经在单独的R01中获得资助。神经元标志物将在体内通过MRS和死后通过免疫组织化学（IHC）和离体MRS进行量化，以收集初步数据，阐明米诺环素治疗在SIV感染期间的神经保护作用（特异性目的1）。当神经元标记物NAA显著降低确定神经元损伤时，将开始治疗，以确定二甲胺四环素是否仅能防止进一步损伤，还是实际上能够促进神经元恢复。采用突触素（突触前完整性的标志物）和微管相关蛋白2 （MAP2，树突完整性的标志物）的死后免疫结构将用于确认体内研究结果。由于预计神经元恢复是二甲胺四环素抗炎作用的结果，NAA的变化将在假定的神经胶质代谢物标记物，胆碱（Cho）和肌醇（MI）的减少之前，通过体内MRS测量（特异性目的2）。这些动物的巨噬细胞/小胶质细胞和星形胶质细胞的相对活化程度将在死后进行评估。巨噬细胞的IHC标记物包括CD68、CD163和主要组织相容性复合体（MHC） II类，而胶质纤维酸性蛋白（GFAP）将被用作星形胶质细胞形成的标记物。最后，将比较接受二甲胺四环素治疗和接受抗逆转录病毒治疗的动物之间SIV感染的病毒学标志物（特定目的3）。采用定量SIV RNA PCR、原位杂交和免疫组化检测CNS感染SIV及相关细胞类型。NAA与血液、脑脊液和脑内病毒水平的线性回归分析将进一步揭示米诺环素的抗病毒特性。项目描述：相当数量的hiv感染患者出现神经系统症状，从轻微的认知障碍到严重的痴呆（神经艾滋病），这被认为是大脑神经元损伤的结果。二甲胺四环素是一种耐受性良好、价格低廉的抗生素，已被用于治疗多种神经疾病；然而，其工作机制尚不清楚。本研究的重点是利用神经艾滋病加速动物模型，通过连续访问体内神经元健康，以及单核细胞/巨噬细胞激活和病毒载量，揭示二甲胺四环素的潜在功能。

DESCRIPTION (provided by applicant): Early in the AIDS epidemic, neurological disorders produced by HIV were recognized as an important clinical manifestation of the disease. Despite the success of HAART, the neurocognitive complication of AIDS infection continues to be an important and possibly increasing problem. The long-term goal of this study is to show that minocycline, a well-tolerated, inexpensive antibiotic, can prevent neuronal injury and promote neuronal recovery in an accelerated animal model of neuroAIDS. Minocycline has been tested in a variety of neuronal diseases including neuroAIDS. However, its mechanism of action is still unknown. Our study seeks to produce preliminary data to understand the mechanism of minocycline. We propose to serially assess neuronal health, monocyte/macrophage activation and viral production in the blood, CSF and brain to reveal minocycline's neuroprotective, anti-inflammatory and/or antiviral properties. To this end, seven SIV-infected CD8+ cell depleted macaques will receive daily doses of minocycline beginning 28 days post inoculation, and data will be collected using magnetic resonance spectroscopy (MRS), immunology, virology and pathology. Their results will be compared to an uninfected cohort, two SIV infected CD8 depleted untreated cohorts and a cohort treated with combination antiretroviral therapy (CART). These four cohorts are already funded in a separate R01. Neuronal markers will be quantified in vivo by MRS and post mortem by immunohistochemistry (IHC) and ex vivo MRS to collect preliminary data elucidating the neuroprotective effect of minocycline treatment during SIV infection (specific aim 1). Treatment will be initiated when neuronal damage has been identified by a significant decrease in the neuronal marker NAA, in order to determine if minocycline merely prevents further injury or if in fact it is able to promote neuronal recovery. Post mortem IHC using synaptophysin (a marker of pre-synaptic integrity) and microtubule associated protein 2 (MAP2, a marker of dendritic integrity) will be used to confirm the in vivo findings. Since it is expected that neuronal recovery is a result of minocycline's anti- inflammatory effect, changes in NAA will be preceded by a reduction in the putative glial metabolite markers, choline (Cho) and myo-Inositol (MI), measured by in vivo MRS (specific aim 2). The relative degree of macrophage/microglia and astrocyte activation in these animals will be assessed post mortem. IHC for macrophage markers include CD68, CD163, and major histocompatibility complex (MHC) class II, while glial fibrillary acidic protein (GFAP) will be used as an astrogliosis marker. Finally, virologic markers for SIV infection will be compared between animals that receive minocycline therapy and those that undergo antiretroviral therapy (specific aim 3). SIV infection of the CNS and relative cell types infected will be assessed using quantitative SIV RNA PCR, in situ hybridization and IHC. Linear regression analysis between NAA and the viral levels in the blood, CSF and brain will be performed to further reveal the antiviral properties of minocycline. PROJECT NARRATIVE: A significant number of HIV-infected patients develop neurological symptoms ranging from minor cognitive impairment to severe dementia (neuroAIDS), which are thought to be a result of injury to neurons in the brain. Minocycline, a well-tolerated, inexpensive antibiotic has been tested in a variety of neuronal diseases; however the mechanism of how this works is unknown. The focus of this study is to unravel the underlying function of minocycline by serially accessing neuronal health in vivo, as well as monocyte/macrophage activation and viral loads, utilizing an accelerated animal model of neuroAIDS.