HIV-induced immune activation impairs immune reconstitution through S1P

描述（由申请人提供）：HIV-1感染个体的持续免疫激活，其特征是促炎细胞因子水平升高，如IL-7、IL-6、IFN-？肿瘤坏死因子- ? ?尽管抗逆转录病毒治疗成功，但包括CD69在内的激活标记物的表达与T细胞再生的缺乏有关。缺乏T细胞再生的部分原因可能是由于造血干细胞进入胸腺和幼稚T细胞从胸腺向外周的出口减少。然而，没有数据可以阐明人类的这些过程。小鼠实验表明，鞘氨醇-1-磷酸（S1P）及其受体之一S1P受体-1 （S1P- r1）在幼年T细胞从胸腺向外周的输出以及记忆T细胞从次要淋巴组织的输出中起着重要作用。也有证据表明，S1P控制胸腺内皮细胞和皮质上皮细胞表达CCL25等趋化因子，从而控制前体进入小鼠胸腺。此外，S1P可增强SDF-1?？/CXCR4在人CD34+细胞中的迁移综上所述，这些数据表明S1P在CD34+前体进入胸腺以及幼稚T细胞从胸腺向外周的输出中起着重要作用。除了S1P受体的表达外，CD69分子还被发现在调节胸腺细胞的分泌中发挥作用。CD69被认为是外周细胞的早期激活分子。在胸腺中，CD69在阳性和阴性选择中表达，但在最近的胸腺迁移中不存在。我们在人类胸腺中的新初步数据表明，只有成熟的CD69-胸腺细胞（CD3hiCD27+CD45RA+ CD62L+CD69-），它们在表型上与幼稚的外周血T细胞相同，才能对S1P产生反应，并在响应S1P时迁出胸腺。这种作用可被S1P-R激动剂FTY720抑制。有趣的是，我们的初步数据还显示，暴露于IL-7和IFN-后，CD69在CD34+祖细胞上表达上调。因此，基于文献中的小鼠数据和我们对人CD34+细胞和胸腺细胞的初步数据，我们假设HIV-1诱导的免疫激活增加了CD69的表达，导致S1P-R1的下调，从而干扰了CD34+细胞进入胸腺和成熟CD69-胸腺细胞从胸腺向外周的出口。我们将使用我们现有的方法，以及新的方法和独特的试剂来检验我们的假设。实验将在hiv感染的人源化小鼠模型和体外T细胞发育和迁移实验中进行，具体目的如下：1。探讨hiv诱导的S1P及其受体免疫激活对CD34+造血干细胞进入胸腺的影响。2. 通过改变S1P受体的表达，确定HIV感染是否会影响T淋巴细胞亚群从胸腺向外周的输出。目前的提议代表了一种独特的合作方法来阐明HIV诱导的免疫激活在HIV-1感染中T细胞发育和外周幼稚T细胞重构中的作用。对初生T细胞进入胸腺和离开胸腺的前体损伤机制的深入了解，将有助于开发免疫调节治疗干预措施，以抵消受损的T细胞发育和再生。

DESCRIPTION (provided by applicant): Persistent immune activation in HIV-1 infected individuals, characterized by increased levels of pro- inflammatory cytokines, such as IL-7, IL-6, IFN-?, TNF-??, and expression of activation markers, including CD69, is associated with a lack of T cell regeneration despite successful antiretroviral therapy. Lack of T cell regeneration is likely due in part to a decrease in entry of hematopoietic stem cells into the thymus and egress of naive T cells from the thymus to the periphery. However, there are no data available elucidating these processes in humans. From studies in the mouse it has become apparent that Sphingosine-1-phosphate (S1P) and one of its receptors, S1P receptor-1 (S1P-R1) play essential roles in egress of naive T cells from the thymus to the periphery as well as egress of memory T cells from the secondary lymphoid tissues. There is also evidence that S1P controls expression of chemokines like CCL25 by thymic endothelial and cortical epithelial cells and thereby entry of precursors into the murine thymus. In addition, S1P enhances SDF-1??/CXCR4 migration of human CD34+ cells. Together these data suggest that S1P plays an important role in entry of CD34+ precursors into the thymus as well as egress of naive T cells from the thymus to the periphery. In addition to S1P receptor expression, the CD69 molecule has been found to play a role in regulating thymocyte egress. CD69 is known as an early activation molecule in the periphery. In the thymus CD69 is expressed during positive and negative selection, but is not present on recent thymic emigrants. Our novel preliminary data in the human thymus show that only mature CD69- thymocytes (CD3hiCD27+CD45RA+ CD62L+CD69-), which are phenotypically identical to naive peripheral blood T cells, can respond to S1P and emigrate out of the thymus in response to S1P. This effect is inhibited by FTY720, an S1P-R agonist. Interestingly, our preliminary data also shows that CD69 is upregulated on CD34+ progenitor cells after exposure to IL-7 and IFN-??. Thus, based on murine data in the literature and our preliminary data with human CD34+ cells and thymocytes we hypothesize that HIV-1 induced immune activation increases expression of CD69 leading to downregulation of S1P-R1, thereby interfering with entry of CD34+ cells into and egress of mature CD69- thymocytes out of the thymus to the periphery. We will use our established methods as well as novel approaches and unique reagents to test our hypothesis. Experiments will be performed with HIV-infected humanized mouse models and in vitro T cell development and migration assays as proposed in the following specific aims: 1. To investigate the effect of HIV-induced immune activation on S1P and its receptors on entry of CD34+ hematopoietic stem cells into the thymus. 2. To determine whether HIV infection affects egress of nave T lymphocyte subsets from the thymus to the periphery through changes in S1P receptor expression. The present proposal represents a unique collaborative approach to elucidate the role of HIV induced immune activation on T cell development and reconstitution of peripheral naive T cells in HIV-1 infection. Insights into the mechanisms underlying the impairment of precursor entry into and exit of naive T cells out of the thymus will enable the development of immunomodulatory therapeutic interventions to counteract impaired T cell development and regeneration. PUBLIC HEALTH RELEVANCE: Persistent immune activation in HIV-1 infected individuals, characterized by increased levels of pro- inflammatory cytokines and expression of activation markers, is associated with a lack of T cell regeneration despite successful antiretroviral therapy. Lack of T cell regeneration is likely due in part to a decrease in entry of hematopoietic stem cells into the thymus and egress of naive T cells from the thymus to the periphery, but there are no data are available elucidating these processes in humans. The present proposal represents a unique collaborative approach to elucidate molecular mechanisms governing factors involved in entry of hematopoietic stem cells and exit of naive T cells in T cell regeneration in HIV-1 infection.