Role of gp120 in Neuropathogenesis in a New Model of HIV Infection in Mice

描述（由申请人提供）：我们最近建立了一种在健康免疫正常小鼠中感染HIV-1的新模型。EcoHIV是一种携带MLV包膜的嵌合HIV-1，可介导HIV-1进入小鼠细胞，经静脉或腹腔注射后，小鼠会持续感染，产生针对病毒蛋白的抗体，并在脾脏、腹膜巨噬细胞和PBL中存在病毒。很大一部分接种过疫苗的动物显示出病毒早期渗入大脑的证据，MCP-1、STAT1和IL-1b等基因的表达升高，这些基因与人类脑组织炎症有关。相比之下，相对较少的动物显示出明显的脑部疾病的脑损伤。因此，EcoHIV在小鼠中具有神经嗜性和神经致病性，但它不表达HIV-1 gp120，并且似乎在小鼠中引起减毒的“临床前”脑疾病。HIV-1 gp120被认为通过与巨噬细胞、星形胶质细胞和神经元表面受体的相互作用参与神经发病。初步研究表明，携带天然gp120的HIV-1在培养的小鼠星形胶质细胞中诱导的炎症分子范围比EcoHIV大。我们假设EcoHIV表达的gp120片段，如V3，将促进hiv -1介导的小鼠神经发病。本文提出的探索性研究将从两个方面来评估这一假设。在Aim 1中，我们将构建一系列嵌合EcoHIV，将其插入到EcoHIV的MLV Env中，其V3周围的HIV-1 gp120区域越来越大，包括那些负责与表面受体相互作用的区域。与EcoHIV相比，将测试嵌合体在体外和体内的感染性，以及在培养的小鼠巨噬细胞和星形胶质细胞中诱导炎症蛋白的能力。我们构建的第一个这样的嵌合体EcoHIV-V3在体内被发现具有传染性。在Aim 2中，我们将比较选择的表达gp120的嵌合体和EcoHIV在小鼠神经发病机制中的诱导作用。我们将确定体内病毒感染和神经侵袭的动力学，大脑中HIV-1神经发病机制的选定分子标记的表达，以及神经病理学的发展。所提出的研究可能有助于确定HIV-1 gp120在HIV-1介导的小鼠神经发病机制中的作用，区别于缺乏gp120的病毒基因组的贡献。该研究采用动物模型、动物组织的分子和病理分析、分子病毒学和重组DMA技术。在更大的公共卫生背景下，我们的研究将建立这种HIV-1感染动物模型在HIV-1发病机制研究中的实用性，包括在中枢神经系统中的研究，HIV-1疫苗的开发，以及方便廉价的体内抗病毒化合物测试。

DESCRIPTION (provided by applicant): We have recently established a novel model for HIV-1 infection in healthy immunocompetent mice. Mice inoculated i.v. or i.p. with EcoHIV, a chimeric HIV-1 carrying MLV envelope that mediates HIV-1 entry into mouse cells, become persistently infected, with production of antibodies to viral proteins and presence of virus in the spleen, peritoneal macrophages, and PBL. A large proportion of inoculated animals show evidence of early virus infiltration into the brain and elevated expression of genes such as MCP-1, STAT1, and IL-1b, which are associated with inflammation in brain tissue in humans. In contrast, relatively few animals displayed brain lesions indicative of overt brain disease. Thus EcoHIV is neurotropic and neuropathogenic in mice but it does not express HIV-1 gp120 and it appears to cause attenuated, "preclinical" brain disease in mice. HIV-1 gp120 is thought to contribute to neuropathogenesis through interaction with surface receptors on macrophages, astrocytes, and neurons. Preliminary studies indicate that HIV-1 carrying native gp120 induces a wider range of inflammatory molecules in mouse astrocytes in culture than EcoHIV. We hypothesize that expression of selected gp120 segments, such as V3, by EcoHIV will facilitate HIV-1-mediated neuropathogenesis in mice. The exploratory studies proposed here will evaluate this hypothesis in two Aims. In Aim 1, we will construct a series of chimeric EcoHIV with increasingly large HIV-1 gp120 regions around V3, including those responsible for interaction with surface receptors, inserted into MLV Env of EcoHIV. The chimeras will be tested for infectivity in vitro and in vivo and for their ability to induce inflammatory proteins in mouse macrophages and astrocytes in culture compared to EcoHIV. The first such chimera we constructed, EcoHIV-V3, was found to be infectious in vivo. In Aim 2, we will compare selected gp120- expressing chimeras and EcoHIV for induction of neuropathogenesis in mice. We will determine the kinetics of virus infection and neuroinvasion in vivo, the expression of selected molecular markers of HIV-1 neuropathogenesis in the brain, and the development of neuropathology. The proposed studies may help to define the role of HIV-1 gp120 in HIV-1-mediated neuropathogenesis in mice in distinction from contribution of the viral genome lacking gp120. The studies employ an animal model, molecular, and pathological analyses of animal tissues, and molecular virology and recombinant DMA technologies. In the larger public health context, our studies will establish the utility of this animal model of HIV-1 infection in research on HIV-1 pathogenesis, including that in the central nervous system, development of HIV-1 vaccine, and convenient inexpensive testing of antiviral compounds in vivo.