Modulation of Dendritic Cells against HIV-1

调节树突状细胞对抗 HIV-1

• 批准号: 7847595
• 负责人: Dinesh K. Singh
• 金额: $ 3.69万
• 依托单位: NORFOLK 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 2010-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7847595
• 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 cell response; 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; 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）：目前还没有艾滋病毒/艾滋病疫苗。抗逆转录病毒疗法（ART）不能从感染个体中根除HIV，因为病毒储库的存在和ART抗性株的发展。树突状细胞（DC）与HIV宿主的发育和传播有关。现有数据表明，HIV颗粒被DC-SIGN捕获，并在pH较温和的内体中路由，以逃避溶酶体降解。一些实验室已经报道了使用针对DC-SIGN的siRNA来减少DC中的HIV感染。但目前尚不清楚DC中参与HIV转运的细胞内分子。四跨膜蛋白如CD 9、CD 63和CD 81在内体中与HIV-1颗粒共定位，但它们在这种环境中的确切作用尚未完全阐明。因此，本项目的目标是通过用HIV-DNA疫苗致敏来调节DC，并评估DC-SIGN、CD 63、CD 81和CD 9在DC内细胞内运输中的作用。核心假设是，由于DC-SIGN、CD 63、CD 81和CD 9参与了HIV向易感T细胞的捕获、内化和传播，因此这些分子的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。目的1探讨HIV DNA疫苗对DC捕获和杀伤病毒的调节作用。目的二是研究DC-SIGN对HIV病毒体向T细胞的传递的RNAi干扰作用。目的3将评估CD 9、CD 63和CD 81在DC内HIV内部贩运中的作用。该方法是通过HIV DNA转染来致敏DC，并通过RNAi干扰来敲低DC-SIGN、CD 9、CD 63和CD 81的表达，以阐明它们在T细胞的内化、细胞内定位和反式感染中的确切作用。我们将采用免疫荧光共聚焦显微镜、电子显微镜、gp 120 Env ELISA、Western blot、RT PCR和共培养试验来获得我们的结果。我们预期HIV- DNA引发将激活DC通过抗原加工和呈递途径捕获和加工HIV。还预期四跨膜蛋白的RNAi沉默将使HIV转向溶酶体降解。我们预计，这一战略将防止艾滋病毒宿主的产生和传播。这些结果可能对理解艾滋病毒的发病机制和传播产生深远而重大的影响。