Plasmacytoid Dendritic Cells in HIV pathogenesis

描述（由申请人提供）：浆细胞样树突状细胞（pDC）是体内最有效的I型干扰素（IFN）产生细胞，被认为是“专业的IFN产生细胞”。hiv感染者的pDC在数量上和功能上都存在缺陷，这种功能障碍与进展为机会性感染密切相关。除了证明pDC在HIV感染中具有重要的保护作用外，过度的pDC IFN-a应答也与免疫激活和HIV发病机制有关。本应用中的实验是基于我们的观察，即病毒血症hiv感染者的外周血pDC不仅在数量上减少，而且在功能上也发生了改变。我们的研究表明，这种数量和功能障碍是由几个因素引起的，包括一些循环pDC的凋亡，外周细胞的激活和潜在的募集，以及外周pDC被骨髓新迁移的细胞取代。此外，我们发现pDC优先从活的病毒感染细胞和未感染细胞中摄取膜和细胞质片段，随后产生IFN-a，并在这种“蚕食”中成熟。然而，当与hiv感染的细胞相互作用时，这一过程被破坏，pDC与hiv感染的细胞融合。我们提出了三个具体目标来跟进这些观察结果：在第一个具体目标中，我们将研究pDC与hiv感染细胞融合的要求和后果。我们将确定不同的病毒株如何影响IFN-a的产生和pDC: hiv感染的细胞相互作用，无论是作为无细胞病毒还是作为病毒感染的细胞。这些病毒将包括从CHAVI获得的早期传播病毒以及慢性病毒。成像和传统的流式细胞术将是我们实验方法的核心。在第二个目标中，我们将通过三种不同的方法确定pDC:T细胞融合是否发生在体内和淋巴组织中：首先，我们将确定hiv感染患者外周血中是否有pDC:T细胞融合的证据；其次，我们将确定pDC: hiv感染的T细胞融合是否发生在扁桃体悬浮细胞和扁桃体器官培养中；第三，我们将与加州大学洛杉矶分校的Christel Uittenbogaart博士合作，确定hiv感染的HIS小鼠是否发生pDC:T细胞融合。在最后的具体目标中，我们将研究pDC在HIV感染和非HIV感染个体中不同表型亚群的功能意义，并研究HIV感染个体骨髓中的pDC。总之，本应用程序中提出的研究有望为人类pDC的生物学及其在HIV-1感染中的功能障碍机制提供有价值的新信息。

DESCRIPTION (provided by applicant): Plasmacytoid dendritic cells (pDC) are the most potent type I interferon (IFN)- producing cells in the body and are considered to be the "professional IFN producing cells". pDC become both numerically and functionally deficient in HIV-infected individuals, and this dysfunction is strongly correlated with progression to opportunistic infections. In addition to evidence for important protective roles of the pDC in HIV infection, an overzealous pDC IFN-a response has also been associated with immune activation and HIV pathogenesis. The experiments in this application are based on our observations that peripheral blood pDC in viremic HIV-infected-individuals are not only numerically depleted, but are also functionally altered. Our studies since the last competing submission of this grant indicate that this numerical and functional dysfunction is contributed to by several factors including apoptosis of some circulating pDC, activation and potentially recruitment out of the periphery, as well as replacement of peripheral pDC with newly emigrated cells from the bone marrow. In addition, we have discovered that pDC preferentially take-up pieces of membrane and cytoplasm from live, virus-infected cells vs. uninfected cells and subsequently produce IFN-a and mature in response to this "nibbling". Upon interaction with HIV-infected cells, however, this process is subverted, and pDC fuse with the HIV-infected cells. We propose three specific aims to follow-up on these observations: In the first specific aim, we will investigate the requirements for, and consequences of, pDC fusion with HIV-infected cells. We will determine how different viral strains affect IFN-a production and pDC:HIV-infected cell interactions, both as cell-free viruses and as virus-infected cells. Included in these viruses will be early transmitted viruses as well as chronic-stage viruses to be obtained from CHAVI. Imaging and traditional flow cytometry will be central to our experimental approaches. In the second aim, we will determine whether pDC:T cell fusion occurs in vivo and in lymphoid tissues using three distinct approaches: first we will determine whether there is evidence of pDC:T cell fusion in peripheral blood of HIV-infected patients; second, we will determine whether pDC:HIV-infected T cell fusion occurs in tonsil suspension cells and tonsil organ culture; third, in collaboration with Dr. Christel Uittenbogaart of UCLA, we will determine whether pDC:T cell fusion occurs in HIV-infected HIS mice. In the final specific aim, we will investigate the functional significance of the different phenotypic subpopulations of pDC in individuals with and without HIV infection and will investigate pDC in the bone marrow of HIV-infected individuals. Together, the studies proposed in this application are anticipated to provide valuable new information on the biology of human pDC and the mechanisms underlying their dysfunction in HIV-1 infection. PUBLIC HEALTH RELEVANCE: Plasmacytoid dendritic (pDC) cells are an important component of the innate immune response to HIV-1. In this continuing renewal application, we will follow-up on our interesting observations of aberrant pDC activation in HIV-infected individuals and pDC fusion with HIV-infected cells to determine how these factors impact HIV pathogenesis.