Novel Model for HIV Latency in Primary Memory T Cells

原代记忆 T 细胞中 HIV 潜伏期的新模型

• 批准号: 8082543
• 负责人: Eric M. Verdin
• 金额: $ 96.5万
• 依托单位: J. DAVID GLADSTONE INSTITUTES
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-30 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8082543
• 关键词: AIDS/HIV problem; Affect; Cell Maintenance; Cells; Excision; FOXO3A gene; Fluorescence Microscopy; Genetic Polymorphism; Genetic Transcription; HIV; HIV Infections; Highly Active Antiretroviral Therapy; Human; Immunology; In Vitro; Individual; Infection; Kinetics; Lead; Life; Lymphocyte; Lymphoid Cell; Maintenance; Memory; Microfluidics; Modeling; Molecular Virology; Nature; PI3K/AKT; Pathway interactions; Patients; Proteins; Recombinants; Research; Role; Signal Transduction; T memory cell; T-Lymphocyte; Techniques; Technology; Time; Transcriptional Activation; Transcriptional Regulation; Transformed Cell Line; abstracting; cellular targeting; drug abuser; in vitro Model; memory CD4 T lymphocyte; novel; prevent; protein activation; research study; transcription factor

DESCRIPTION (provided by applicant): The long-term persistence of HIV in a latent state in memory T cells in patients treated with HAART prevents the eradication of HIV and forces patients to remain on HAART for their whole life. While the transcriptional regulation of HIV has been extensively studied in transformed cell lines, our understanding of how latent HIV infection occurs in primary memory CD4 T lymphocytes is rudimentary. The purpose of this application is to develop new single cell technology to examine the transcriptional status of HIV in single primary lymphoid cells over time after an infection in vitro. These studies will bridge the two research fields of human immunology and HIV molecular virology. Understanding HIV latency in primary lymphocytes may lead to the identification of cellular proteins that control the entry of HIV in latency, the maintenance of latency or its reactivation. Such cellular targets could represent new avenues for the treatment of HIV/AIDS among drug abusers and possibly lead to the eradication of infection. I propose to use a novel live cell, time-lapse fluorescence microscopy combined with cell trapping via microfluidic chips and the use of HIV expressing recombinant fluorescent protein (destabilized GFP) to study the kinetics of HIV transcription at the single cell level. This novel technique will allow the fate of HIV expression to be followed in live individual cells over time. Human lymphoid cells will be activated in vitro, infected with an HIV expressing a fluorescent protein, activation signals will be removed and HIV transcription will be followed over time. We anticipate that HIV transcription will be restricted in a subset of cells returning to quiescence. The time separating removal of activation signal to infection is likely to be critical in allowing infection to proceed until HIV integration while restricting HIV transcriptional activation. The nature of the activation signal could also prove critical. Experiment will eventu

描述（由申请人提供）：接受HAART治疗的患者中，HIV在记忆T细胞中的潜伏状态长期持续存在，阻止了HIV的根除，并迫使患者终生接受HAART治疗。虽然HIV的转录调控已经在转化细胞系中得到了广泛的研究，但我们对初级记忆性CD 4 T淋巴细胞中潜伏性HIV感染如何发生的理解是初步的。本申请的目的是开发新的单细胞技术，以检查体外感染后单个初级淋巴细胞中HIV随时间的转录状态。这些研究将在人类免疫学和HIV分子病毒学两个研究领域之间架起桥梁。了解HIV在初级淋巴细胞中的潜伏期可能会导致识别控制HIV进入潜伏期、维持潜伏期或重新激活的细胞蛋白。这种细胞靶点可以成为治疗药物滥用者中艾滋病毒/艾滋病的新途径，并可能导致根除感染。我建议使用一种新的活细胞，延时荧光显微镜结合细胞捕获通过微流控芯片和使用的HIV表达重组荧光蛋白（不稳定的GFP）研究在单细胞水平的HIV转录的动力学。这项新技术将允许随着时间的推移在活的个体细胞中跟踪HIV表达的命运。人类淋巴细胞将在体外被激活，感染表达荧光蛋白的HIV，激活信号将被去除，HIV转录将随着时间的推移而被跟踪。我们预计，艾滋病毒的转录将被限制在一个子集的细胞恢复到静止状态。从激活信号的去除到感染的间隔时间可能是允许感染进行直到HIV整合的关键，同时限制HIV转录激活。激活信号的性质也可能被证明是至关重要的。实验将最终