Using miRNAs to elucidate the cellular sources of HIV-1

描述(由申请人提供)：在急性和随后的持续性感染期间，允许细胞群体对HIV-1病毒血症和总病毒载量的体内相对贡献尚不清楚。更好地了解组织巨噬细胞和CD4+T细胞亚群的作用将有助于更全面地了解艾滋病毒的发病机制，并有可能刺激细胞特异性抗病毒方法的发展，这些方法可能会减少与当前抗病毒疗法相关的副作用。到目前为止，了解体内不同细胞间隔的影响的方法一直是有限的和费力的。建议在这里开发的新方法将允许该领域清楚地定义这些细胞对病毒载量和细胞因子/趋化因子变化的贡献。在这些组织培养和外植体研究中，我们建议开发一种新的创新技术来控制HIV的趋向性，目的是描述特定细胞类型对HIV感染和发病的贡献，并提供在体内使用时可以为HIV细胞储存库的影响提供重要见解的原则证据。我们将通过利用内源性miRNAs的细胞特异性表达来设计限制在髓系或淋巴系特异性培养中的重组HIV毒株。通过将完美的miRNA靶点整合到HIV基因组中，这些HIV的表达被限制在目标细胞亚型中，从而使我们能够确定这些细胞因功能表型丧失而做出的贡献。通过限制HIV在各种不同的造血细胞亚群中的复制，我们将确定这如何在原发体外感染的背景下对病毒和宿主生物学产生影响。

DESCRIPTION (provided by applicant): The relative in vivo contribution of permissive cell populations to HIV-1 viremia and total virus load during acute and subsequent persistent infections is poorly understood. A better understanding of the contribution of tissue macrophages and CD4+ T cell subpopulations will allow a more complete picture of HIV pathogenesis and potentially stimulate the development of cell-specific antiviral approaches that are likely to decrease side effects that are associated with current antiviral therapies. To date, approaches to understand the impact of the different cellular compartments in vivo have been limited and laborious. The novel approach proposed to be developed here will allow the field to clearly define the contribution of these cells to virus load and cytokine/chemokine changes. In these tissue culture and explant studies, we propose to develop a new and innovative technology to control the tropism of HIV for the purposes of delineating the contribution of specific cell types for HIV infection and pathogenesis and provide proof-of-principal that when used in vivo can provide important insights into impact of the cellular reservoirs of HIV. We will engineer recombinant HIV strains that are restricted in either myeloid- or lymphoid-specific cultures by harnessing cell- specific expression of endogenous miRNAs. Through incorporation of perfect miRNA target sites into the HIV genome, expression of these HIVs are restricted in targeted cellular subtypes, thereby allowing us to ascertain the contribution of these cells by a loss of function phenotype. By restricting HIV replication in a variety of different hematopoietic cellular subsets, we will determine how this impacts both virus and host biology in the context of primary ex vivo infections.