Involvement of HIV-1 Vpr in neuronal degeneration.

HIV-1 Vpr 参与神经元变性。

• 批准号: 8512828
• 负责人: BASSEL E SAWAYA
• 金额: $ 36.91万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-30 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8512828
• 关键词: Acquired Immunodeficiency Syndrome; Animal Model; Astrocytes; Axonal Transport; BCL2 gene; BDNF gene; Behavioral; Biological Assay; Biological Models; Blood - brain barrier anatomy; Body Fluids; CREB1 gene; Calcium; Cell Line; Cells; Communication; Data; Dementia; Detection; Development; Disease; Encephalopathies; Endoplasmic Reticulum; Enzyme-Linked Immunosorbent Assay; Gene Expression; Gene Targeting; Genes; Genome; HIV; HIV Envelope Protein gp120; HIV-1; Highly Active Antiretroviral Therapy; Hippocampus (Brain); Homeostasis; Human; Immunohistochemistry; In Vitro; Incidence; Individual; Infection; Inflammatory; Latent Virus; Lead; Length; Measures; Membrane; Methods; MicroRNAs; Microglia; Mitochondria; Modeling; Molecular; Mus; Nerve Degeneration; Neurites; Neurocognitive; Neuronal Dysfunction; Neuronal Plasticity; Neurons; Outcome Study; Oxidative Stress Pathway; Pathogenesis; Pathway interactions; Patients; Pharmaceutical Preparations; Prevalence; Proteins; Q-Type Calcium Channels; RNA; Rattus; Recombinants; Regulation; Role; Serum; Small RNA; Source; Stress; Synaptic plasticity; Testing; Therapeutic Agents; Transgenic Mice; Validation; Viral; Viral Load result; Viral Proteins; Virion; Virus; Virus Diseases; Western Blotting; base; brain tissue; cytokine; endoplasmic reticulum stress; extracellular; in vivo; macrophage; mitochondrial dysfunction; mutant; neuron loss; neurotoxic; new therapeutic target; prevent; release factor

DESCRIPTION (provided by applicant): Even in the HAART era where the viral load is below detection levels, the prevalence of HIV-1 associated neurocognitive disorders (HAND) remains high due to many reasons such as latent virus reactivation and drugs inability to efficiently cross the blood brain barrier (BBB). Therefor, it is important to understand the mechanisms leading to neuronal deregulation in HIV-1-infected patients in the HAART era. The lack of productive infection of neurons by HIV-1 suggests that viral and cellular proteins with neurotoxic activities that are released from HIV-1 infected target cells, or reservoirs cells for latent active virus, cause this neuronal deregulation. The viral protein R (Vpr) is one of the proteins encoded by HIV-1 and has been shown to alter the expression of various important cytokines and inflammatory proteins in infected and uninfected cells. The mechanisms and the cellular factors used by Vpr to cause neuronal damage remain unclear. Using human neuronal cell line, SH-SY5Y, we found that Vpr can be taken up by neurons (immunohistochemistry and Western blot analysis), which gave us the rationale to measure the amount of Vpr in neurons (HPCE) (almost 100 pg). We also demonstrated that Vpr deregulates calcium homeostasis, promotes endoplasmic reticulum (ER)-calcium release and stress, activation of the oxidative stress pathway, mitochondrial dysfunction and axonal transport alteration. These effects were specifically noted with the full length Vpr but not with Vpr mutant (R73A). In search for the cellular factors involved, we performed microRNA and gene array assays using RNA collected from primary human cultures of neurons and/or SH-SY5Y-treated with recombinant Vpr protein. Interestingly, Vpr deregulates the levels of several microRNA (e.g. miR-34a) and their target genes (e.g. CREB), which could lead to neuronal dysfunctions. These factors were deregulated in human brain tissues of HIV-infected patients and in the brain tissues of Vpr-transgenic mice. Interestingly, these factors were also deregulated in neuronal cells treated with Tat, gp120 or supernatant collected from infected cells. Therefore, we propose a comprehensive study utilizing molecular, virological, and cellular approaches to unravel the mechanisms and identify the cellular factors used by Vpr as well as its interplay with microRNAs to cause neuronal dysfunction. These studies will be validated in an animal model (Vpr-transgenic mice). The outcome of these studies will advance the understanding of HIV-1 pathogenesis and will decipher the mechanisms used by Vpr that lead to neuronal degeneration even in the HAART era.

描述（由申请人提供）： 即使在HAART时代，病毒载量低于检测水平，HIV-1相关的神经认知疾病（手）的患病率仍然很高，这是由于许多原因（例如潜在病毒重新激活）和药物无法有效地越过血液脑屏障（BBB）。因此，重要的是要了解导致HAART时代HIV-1感染患者神经元失调的机制。 HIV-1缺乏神经元的生产性感染表明，病毒和细胞蛋白具有神经毒性活性，这些活性是从HIV-1感染靶标释放的 细胞或潜在活性病毒的储层细胞会导致这种神经元失调。病毒蛋白R（VPR）是由HIV-1编码的蛋白质之一，已被证明会改变感染和未感染细胞中各种重要的细胞因子和炎症蛋白的表达。 VPR所用的机制和细胞因子导致神经元损伤尚不清楚。 使用人类神经元细胞系SH-SY5Y，我们发现VPR可以通过神经元（免疫组织化学和蛋白质印迹分析）来吸收，这使我们有理由衡量神经元（HPCE）中VPR量（几乎100 pg）。我们还证明了VPR会消除钙稳态，促进内质网（ER） - 钙释放和应激，氧化应激途径的激活，线粒体功能障碍和轴突转运的改变。这些效应是用全长VPR特别指出的，但没有使用VPR突变体（R73A）。为了搜索所涉及的细胞因子，我们使用了从神经元原代培养物和/或用重组VPR蛋白进行的SH-SY5Y处理的RNA进行了microRNA和基因阵列测定。有趣的是，VPR会消除几种microRNA（例如miR-34a）及其靶基因（例如CREB）的水平，这可能导致神经元功能障碍。这些因素在HIV感染患者和VPR-转基因小鼠的脑组织的人脑组织中受到失调。有趣的是，这些因素在用TAT，GP120或从感染细胞中收集的上清液处理的神经元细胞中也解除了管制。 因此，我们提出了一项全面的研究，利用分子，病毒学和细胞方法来揭示机制，并确定VPR使用的细胞因子以及与microRNA的相互作用来引起神经元功能障碍。这些研究将在动物模型（VPR-转基因小鼠）中进行验证。 这些研究的结果将提高人们对HIV-1发病机理的理解，并破译VPR所使用的机制，即使在HAART时代也会导致神经元变性。