HIV-induced immune activation impairs immune reconstitution through S1P

描述(申请人提供)：HIV-1感染者的持续免疫激活，其特征是促炎症细胞因子水平的增加，如IL-7、IL-6、干扰素-α、肿瘤坏死因子-β，以及包括CD69在内的激活标志物的表达，尽管成功地进行了抗逆转录病毒治疗，但与T细胞再生不足有关。缺乏T细胞再生的部分原因可能是造血干细胞进入胸腺的减少和原始T细胞从胸腺向外周流出的减少。然而，目前还没有可用的数据来阐明人类的这些过程。在小鼠的研究表明，神经鞘氨醇-1-磷酸(S1P)及其受体S1P-1(S1P-R1)在幼稚T细胞从胸腺输出到外周以及记忆T细胞从次级淋巴组织输出的过程中起着重要作用。也有证据表明，S1P通过胸腺内皮细胞和皮质上皮细胞控制CCL25等趋化因子的表达，从而控制前体进入小鼠胸腺。此外，S1P还可促进人CD34+细胞SDF-1β/CXCR4的迁移。综上所述，S1P在CD34+前体细胞进入胸腺以及幼稚T细胞从胸腺向外周流出的过程中起着重要作用。除了S1P受体的表达外，CD69分子还被发现在调节胸腺细胞外流方面发挥作用。CD69是已知的外周血细胞早期激活分子。在胸腺中，CD69在正选择和负选择中表达，但在最近的胸腺移民中不存在。我们在人类胸腺中的初步数据表明，只有成熟的CD69-胸腺细胞(CD3hiCD27+CD45RA+CD62L+CD69-)才能对S1P做出反应，并对S1P做出反应，这些细胞与幼小的外周血T细胞表型相同。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，在IL-7和干扰素-β作用下，CD34+祖细胞表面CD69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下调，从而干扰CD34+细胞进入和离开成熟的CD69-胸腺细胞到胸腺的外周。我们将使用我们已有的方法以及新的方法和独特的试剂来检验我们的假设。利用HIV感染的人源化小鼠模型和体外T细胞发育和迁移实验，研究HIV诱导的免疫激活S1P及其受体对CD34+造血干细胞进入胸腺的影响。2.探讨HIV感染是否通过S1P受体表达的改变影响胸腺至外周的NAVE T淋巴细胞亚群的外流。本研究提供了一种独特的协作方法来阐明HIV诱导的免疫激活在HIV-1感染中T细胞发育和外周原始T细胞重建中的作用。深入了解胸腺前体细胞进出胸腺的损伤机制，将有助于开发免疫调节性治疗干预措施，以对抗受损的T细胞发育和再生。 公共卫生相关性：HIV-1感染者的持续免疫激活，其特征是促炎症细胞因子水平和激活标志物的表达增加，尽管成功地进行了抗逆转录病毒治疗，但与T细胞再生不足有关。缺乏T细胞再生的部分原因可能是造血干细胞进入胸腺的减少和原始T细胞从胸腺向外周流出的减少，但目前还没有可用的数据来阐明人类的这些过程。本研究提供了一种独特的协作方法来阐明HIV-1感染中T细胞再生中影响造血干细胞进入和初始T细胞退出的因素的分子机制。

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.