Modulation of Dendritic Cells against HIV-1

调节树突状细胞对抗 HIV-1

• 批准号: 7667422
• 负责人: Dinesh K. Singh
• 金额: --
• 依托单位: WINSTON-SALEM STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2009-08-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7667422
• 关键词: AIDS prevention; Acquired Immunodeficiency Syndrome; Affect; Antigen Presentation Pathway; Antigens; Biological Assay; Body part; CD4 Positive T Lymphocytes; CD81 gene; CD8B1 gene; Cells; Cervix Uteri; Coculture Techniques; Confocal Microscopy; DNA; DNA Vaccines; Data; Dendritic Cells; Development; Disputes; Electron Microscopy; Endosomes; Enzyme-Linked Immunosorbent Assay; Genome; Goals; HIV; HIV Antigens; HIV Envelope Protein gp120; HIV Infections; HIV vaccine; HIV-1; Immunity; Individual; Infection; Lectin; Macaca; Modeling; Morbidity - disease rate; Organ; Outcome; Pathogenesis; Pathway interactions; Phase; Play; Process; RNA; RNA Interference; Reporting; Research; Role; Route; Sentinel; Small Interfering RNA; Source; Staging; Surface; Synapses; T-Lymphocyte; Testing; Therapeutic Intervention; Time; Transfection; Vaccine Antigen; Vaccines; Vagina; Vesicle; Viral; Viral Antigens; Viral Proteins; Virion; Virus; Western Blotting; antiretroviral therapy; efficacy testing; knock-down; mortality; particle; pathogen; prevent; resistant strain; response; simian human immunodeficiency virus; synaptogenesis; therapy resistant; trafficking; transmission process; vaccine efficacy

DESCRIPTION (provided by applicant): Currently, a vaccine for HIV/AIDS is not available. The antiretroviral therapies (ART) fail to eradicate HIV from infected individuals because of the existence of viral reservoirs and development of ART-resistant strains. Dendritic cells (DCs) are implicated in the development and spread of HIV reservoirs. Available data suggest that HIV particles are captured by DC-SIGN and routed in endosomes with milder pHs to evade lysosomal degradation. Internalized particles are subsequently transmitted to T-cells in trans. A few labs have reported using siRNA against DC-SIGN to diminish HIV infection in DCs. But the intracellular molecules participating in trafficking of HIV within DCs are not clear. Tetraspanins such as CD9, CD63 and CD81 colocalize with HIV-1 particles in endosomes, but their exact role in this milieu has not been fully elucidated. Therefore, the goals of this project are to modulate DCs by priming with an HIV-DNA vaccine and evaluate roles of DC-SIGN, CD63, CD81, and CD9 in intracellular trafficking within DCs. The central hypothesis is that, since DC-SIGN, CD63, CD81 and CD9 participate during capture, internalization and transmission of HIV to susceptible T cells, RNAi interference of these molecules will help to elucidate their roles. The central hypothesis will be tested by three specific aims. Aim 1 will evaluate modulation of DCs with HIV-DNA vaccine priming on capture and destruction of virus. Aim 2 will evaluate RNAi interference of DC-SIGN on transmission of HIV virions to T-cells. Aim 3 will evaluate the roles of CD9, CD63 and CD81 in internal trafficking of HIV within DCs. The approach is to prime DCs by HIV DNA transfection, and knock down expressions of DC-SIGN, CD9, CD63 and CD81 by RNAi interference to elucidate their exact roles in, internalization, intracellular localization and trans infection of T-cells. We will employ immunofluorescent confocal microscopy, electron microscopy, gp120 Env ELISA, Western blot, RT PCR and coculture assays to obtain our results. We expect that HIV- DNA priming will activate DCs to capture and process HIV via antigen processing and presentation pathways. It is also expected that RNAi silencing of tetraspanins will divert HIV to lysosomal degradation. We anticipate that this strategy will prevent creation and transmission of HIV reservoirs. These outcomes could have a profound and significant impact on understanding the pathogenesis and spread of HIV.

描述（由申请人提供）：目前尚无艾滋病毒/艾滋病疫苗。由于病毒库的存在和抗逆转录病毒疗法（ART）病毒株的发展，抗逆转录病毒疗法（ART）无法从感染者体内根除艾滋病毒。树突状细胞 (DC) 与 HIV 病毒库的形成和传播有关。现有数据表明，HIV 颗粒被 DC-SIGN 捕获，并进入具有较温和 pH 值的内体中，以避免溶酶体降解。内化的颗粒随后被反式传输至 T 细胞。一些实验室报告使用针对 DC-SIGN 的 siRNA 来减少 DC 中的 HIV 感染。但参与 DC 内 HIV 运输的细胞内分子尚不清楚。 CD9、CD63 和 CD81 等四跨膜蛋白在内体中与 HIV-1 颗粒共定位，但它们在此环境中的确切作用尚未完全阐明。因此，该项目的目标是通过 HIV-DNA 疫苗启动来调节 DC，并评估 DC-SIGN、CD63、CD81 和 CD9 在 DC 内细胞内运输中的作用。中心假设是，由于 DC-SIGN、CD63、CD81 和 CD9 参与了 HIV 捕获、内化和传播到易感 T 细胞的过程，这些分子的 RNAi 干扰将有助于阐明它们的作用。中心假设将通过三个具体目标进行检验。目标 1 将评估 HIV-DNA 疫苗对 DC 的调节作用，以捕获和破坏病毒。目标 2 将评估 DC-SIGN 对 HIV 病毒颗粒向 T 细胞传播的 RNAi 干扰。目标 3 将评估 CD9、CD63 和 CD81 在 DC 内 HIV 内部贩运中的作用。该方法是通过 HIV DNA 转染来引发 DC，并通过 RNAi 干扰敲低 DC-SIGN、CD9、CD63 和 CD81 的表达，以阐明它们在 T 细胞的内化、细胞内定位和反感染中的确切作用。我们将采用免疫荧光共聚焦显微镜、电子显微镜、gp120 Env ELISA、Western blot、RT PCR 和共培养测定来获得我们的结果。我们预计 HIV-DNA 启动将激活树突状细胞，通过抗原加工和呈递途径捕获和处理 HIV。还预计四跨膜蛋白的 RNAi 沉默将使 HIV 转向溶酶体降解。我们预计这一策略将防止艾滋病毒储存库的产生和传播。这些结果可能对了解艾滋病毒的发病机制和传播产生深远而重大的影响。