Murine Model System for HIV Pathogenesis

描述（由申请人提供）：尽管在揭示人类细胞中HIV复制周期的机制方面取得了进展，但我们对病毒如何在体内传播、如何引起免疫缺陷以及为什么免疫应答通常不能有效地中和它的理解有限。这些问题很难在人类甚至非人灵长类动物中进行研究。我们的主要目标是产生一个小鼠模型系统，将忠实地再现HIV- 1与其人类宿主的相互作用。这将允许相对便宜的检查病毒的传播，复制和发病机制的各个方面与众多的变量，它将允许候选人的研究宿主的遗传因素在艾滋病毒疾病。开发这样一个模型需要更好地了解病毒是如何传播的，以及限制HIV在鼠细胞中复制的物种特异性限制。体外研究表明，树突状细胞（DC）在HIV的传播和复制中具有重要作用。DC内吞HIV颗粒，长时间携带HIV，并增强反式T细胞的感染。DC-SIGN是HIV内吞作用的关键DC受体，HIV内吞作用是DC：T细胞共培养物中有效复制HIV所需的过程。HIV与DC相互作用的重要性尚未在体内测试。具体目标1将是使用基因修饰的小鼠来确定在暴露于粘膜内的DC后，HIV是否可以传播到引流淋巴组织中的细胞。我们将研究DC-SIGN转基因小鼠与SCID-Hu小鼠和表达人CD 4、CCR 5和细胞周期蛋白T1的小鼠的组合。尽管来自多重转基因动物的T细胞和巨噬细胞可以感染HIV，但它们不能释放感染性颗粒。具体目标2将采用遗传学方法来鉴定增强HIV组装和/或从鼠细胞释放的人类基因，并且将允许HIV在鼠模型中更好地复制。具体目标3是确定APOBEC 3G（一种靶向HIV Vif蛋白降解的抗病毒蛋白）是否限制原代小鼠细胞中的复制。如果是这样，我们将开发克服其活动的方法。Vif诱导人而非小鼠APOBEC 3G的降解。RNA干扰和基因靶向将用于减少小鼠酶的表达，我们将用Vif改造HIV，使其能够结合并诱导小鼠apobec 3g的降解。

DESCRIPTION (provided by applicant): Despite progress in unraveling the mechanism of the HIV replication cycle in human cells, we have limited understanding of how virus is transmitted in vivo, how it causes immunodeficiency, and why immune responses are generally ineffective in neutralizing it. These issues are difficult to study in humans and even in non-human primates. Our broad aim is to generate a murine model system that will faithfully reproduce the interactions of HIV- 1 with its human host. This would allow for relatively inexpensive examination of various aspects of viral transmission, replication, and pathogenesis with numerous variables, and it would permit studies of candidate host genetic factors in HIV disease. Development of such a model will require a better understanding of how virus is transmitted and of species-specific restrictions that limit HIV replication in murine cells. In vitro studies suggest that dendritic cells (DC) have important roles in HIV transmission and replication. DC endocytose HIV particles, harbor HIV for long periods of time, and enhance infection of T cells in trans. DC-SIGN is a key DC receptor for HIV endocytosis, a process required for efficient HIV replication in DC:T cell co-cultures. The importance of HIV interaction with DC has not yet been tested in vivo. Specific Aim 1 will be to use genetically modified mice to determine if HIV can be transmitted to cells in draining lymphatic tissues after exposure to DC within mucosa. We will examine mice transgenic for DC-SIGN in combination with SCID-Hu mice and mice expressing human CD4, CCR5, and cyclin T1. Although T cells and macrophages from the multiply transgenic animals can be infected with HIV, they fail to release infectious particles. Specific Aim 2 will be to employ genetic approaches to identify human genes that enhance HIV assembly and/or release from murine cells and that will permit better replication of HIV in the murine model. Specific Aim 3 will be to determine if APOBEC3G, an antiviral protein targeted for degradation by the HIV Vif protein, restricts replication in primary mouse cells. If so, we will develop means to overcome its activity. Vif induces degradation of human, but not mouse, APOBEC3G. RNA interference and gene targeting will be used to reduce expression of the mouse enzyme, and we will engineer HIV with Vif that can bind to and induce degradation of mouse apobec3g.