Immune Reconstitution

急性损失后T细胞群重建的生物学特征仍然不完全。通过小鼠模型，我们首先确定了T细胞免疫重建的两种主要途径，一种是经典的胸腺依赖途径，另一种是胸腺独立途径。然后，我们确定了T细胞表面标记物，通过重组T细胞群的表型分析，可以识别产生它们的途径，然后将这些信息应用于患者T细胞重组的表征。最初的工作确定了这种方法在年轻人类T细胞群再生研究中的适用性。最近的工作验证了该方法的有效性，并确定了两种主要途径中T细胞免疫重建在成年人中经过较长时间的过程。这项工作显示了胸腺在定量再生CD4+ T细胞中的重要作用。CD8+ T细胞可以通过外周扩增进行免疫重建，但CD4+和CD8+ T细胞都需要胸腺活性来维持或再生库多样性。这些发现导致研究重点转向了解控制胸腺功能的机制，以及包括疫苗策略在内的新治疗方法，以在再生免疫系统的背景下治疗癌症。目前已发展出四种胸腺调节模型；这项工作已经发展到两个新的研究项目领域——一个专注于胸腺功能的调节，另一个专注于将药物引入临床试验。在这个项目中解决的工作也导致了研究IL-7对胸腺细胞成熟的影响的努力。我们已经确定IL-7是胸腺生成的负调节因子，也是胸腺细胞发育所必需的。这些双重作用是剂量依赖性的，高浓度的负调节是通过控制Notch信号传导介导的，这是T/B谱系承诺的核心。此外，我们已经确定了可能调节胸腺的基因，并确定了一种名为Tbata（以前称为SPATIAL）的基因，这是一种在基质细胞内起作用的负调节作用。它似乎通过控制细胞周期，特别是通过调节Nedd8通路发挥作用。最近的研究结果表明，Tbata对细胞周期的调节超出了Nedd8，并涉及p53。研究了雄激素信号阻断和IGF-1调节胸腺功能的细胞和分子机制，并绘制了胸腺上皮细胞区室增殖调控点。一种新的转基因小鼠模型已经被开发来研究参与这种调节的信号通路。这种新模型涉及到在没有辅助组织损伤的情况下消耗外周T细胞室的能力。利用胸腺调控的一个关键控制点是胸腺上皮细胞增殖的控制这一信息，在离体细胞上以固定的时间间隔进行基因阵列，以鉴定活化的细胞分裂途径。与此同时，LC/MS也被用于鉴定可能作为这些已鉴定途径的配体的新蛋白质物种。已经发现了三种身份不明的蛋白质，并正在进行验证。如上所述，IL-7可能是胸腺功能的调节因子；通过表征IL-7受体在T细胞亚群中的调节作用，进一步研究了其在外周稳态中的作用。所有这些研究的目的都是为了了解T细胞稳态的生物学，以便开发新的治疗方法，以治疗T细胞群减少而导致免疫功能受损的患者。迄今为止，IL-7已被引入人类，一项雄激素阻断的临床试验正在进行中。雄激素阻断的试验显示出早期的积极结果，但随着试验的继续，这些结果需要得到证实。IL-7受体调节的研究表明，在亚群中存在一种复杂的差异信号调节模式，其明显效果有利于维持原始幼稚T细胞。这是特别有趣的，因为它为理解单个细胞因子如IL-7如何差异调节多个不同的T细胞亚群提供了基础。具体来说，初始CD4 T细胞与新近的胸腺移植物之间，以及CD4和CD8 T细胞之间存在差异调节。IL-7受体远端信号的差异似乎在机制上解释了这种差异调节。在人类中，我们注意到化疗后CD4恢复比CD8恢复更有限。其原因尚不清楚。我们现在正在研究小鼠模型中两种主要的稳态细胞因子在CD4和CD8亚群中的信号传导生物学，并发现了信号传导的一个新的复杂层面，有待表征。我们还观察到，在依赖于人类胸腺非依赖性途径的T细胞免疫重建中，这种扩张导致T细胞过早衰老，从而导致细胞衰老，这可能是移植后T细胞功能受损的原因。

The biology of reconstitution of T cell populations following acute loss remains incompletely characterized. Using murine models, we first identified two primary pathways of T cell immune reconstitution, the classic, thymic-dependent pathway, and a second, thymic-independent pathway. We then identified T cell surface markers which allowed identification, by phenotyping of reconstituted T cell populations, of the pathways which had given rise to them, and then applied this information to the characterization of T cell reconstitution in patients. Initial work established the applicability of this approach to the study of T cell population regeneration in humans who were young. Recent work verified validity of the approach and established the course of T cell immune reconstitution in adult humans over an extended period time for each of the two primary pathways. This work showed an essential role for the thymus in regenerating CD4+ T cells quantitatively. CD8+ T cells can numerically be reconstituted by peripheral expansion for immune reconstitution, but both CD4+ and CD8+ T cells require thymic activity for maintenance or regeneration of repertoire diversity. These findings have led in turn to a research emphasis on understanding mechanisms which control thymic function, and new treatments, including vaccine strategies, to treat cancer in the setting of a regenerating immune system. Four models of thymic regulation have been developed; this work has progressed to the developoment of two new project areas of research -- one focused on points of regulation of thymus function and one on introducing agents into clinical trials. The work addressed in this project has also led to efforts in investigating IL-7 effects on the maturation of thymocytes. We have identified IL-7 as a negative regulator of thymopoiesis as well as being essential for thymocyte development. These dual roles are dose dependent and negative regulation at higher concentrations is mediated through control of Notch signaling -- which is central to T/B lineage commitment. Additionally, we have worked to identify genes which might regulate the thymus, and have characterized a gene called Tbata (previously SPATIAL) which is a negative regulator acting within the stromal cell compartment. It appears to exert its effect through control of cell cycle, specifically through regulation of the Nedd8 pathway. Recent results indicate that regulation of cell cycle by Tbata extends beyond Nedd8 and involves p53. The cellular and molecular mechanisms by which androgen signaling blockade and IGF-1 modulate thymus function were characterized and map points of regulation of proliferation to the epithelial cell compartment of the thymus. A new transgenic murine model has been developed to investigate signaling pathways involved in this regulation. This new model involves the ability to deplete the peripheral T cell compartment without ancillary tissue injury. Using the information that a key control point of thymus regulation is the control of thymic epithelial cell proliferation, gene arrays are being carried out at fixed time intervals on isolated cells to identify activated cell pathways of division. In parallel, LC/MS is being used to identify new protein species that may act as ligands for those identified pathways. Three unidentified proteins have been found and are being validated. The role of IL-7 as a possible regulator of thymus function was noted above; its role in peripheral homeostasis has been further investigated by charactreizing IL-7 receptor regulation among T cell subsets. The purpose in all of these studies is to understand the biology of T cell homeostasis in order to develop new approaches to therapy for patients in whom T cell populations are depleted with consequences of impaired immunity. To date, IL-7 has been introduced in humans and a clinical trial with androgen blockade is in progress. The trial with androgen blockage shows early positive results which need to be confirmed as the trial continues. The work with IL-7 receptor modulation has shown a complex pattern of differential signaling regulation among subsets with the apparent effect of favoring the sustaining of primitive naive T cells. This is of special interest because it provides a basis for understanding how a single cytokine such as IL-7 can differentially regulate multiple distinct T cell subsets. Specifically, there is differential regulation between naive CD4 T cells versus recent thymic emigrants, and also between CD4 and CD8 T cells. Differences in signaling distal to the receptor for IL-7 appear to mechanistically account for this differential regulation. In humans, we noted that CD4 recovery was more limited than CD8 recovery following chemotherapy. Reasons for this are not known. We are now addressing the biology of signaling by the two primary homeostatic cytokines in the CD4 versus the CD8 subsets in murine models and have found a new layer of complexity in signaling that remains to be characterized. We have also made the observation that in T cell immune reconstitution reliant on the thymus-independent pathway in humans, that such expansion results in premature aging of the T cells with resultant cellular senescence, a likely contributor to T cell functional impairment following transplant.