Immune Reconstitution

急性丧失后T细胞群体重建的生物学特征仍不完全清楚。利用小鼠模型，我们首先确定了T细胞免疫重建的两条主要途径，经典的胸腺依赖途径和第二条胸腺非依赖途径。然后，我们确定了T细胞表面标志物，通过对重组的T细胞群体进行表型鉴定，确定了导致它们的途径，然后将这些信息应用于患者T细胞重建的特征。这项工作从数量上说明了胸腺在再生CD4+T细胞中的重要作用。CD8+T细胞可以通过外周扩增进行数量上的重组，以进行免疫重建，但CD4+和CD8+T细胞都需要胸腺活动来维持或再生谱系的多样性。这些发现反过来又导致了一项研究重点，即了解控制胸腺功能的机制，以及在再生免疫系统的背景下治疗癌症的新疗法。这项工作已经发展成两个新的研究项目领域--一个集中在胸腺功能的调节点上，另一个集中在将药物引入临床试验中。该项目所涉及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是胸腺生成的负性调节因子，也是胸腺细胞发育所必需的。这些双重作用是剂量依赖的，在较高浓度下的负调节是通过控制Notch信号介导的--Notch信号是T/B谱系承诺的核心。在高剂量下，IL-7有利于B细胞的发育，从而将胸腺转化为B细胞器官。此外，我们还努力识别可能调节胸腺的基因，并确定了一种名为Tbata(以前是空间的)的基因的特征，它是一种在基质细胞间隔内发挥作用的负调控因子。它似乎通过控制细胞周期发挥作用，特别是通过调节Nedd8途径。最近的结果表明，Tbata对细胞周期的调控超出了Nedd8的范围，并涉及到P53。这是由于Tbata编码的蛋白质参与了一个控制基因表达的大型多蛋白质复合体。有趣的是，这可能反映在雌性和雄性小鼠胸腺生成的差异上。这种差异现在已经从多个实验中清晰可见，其机制正在研究中。实验正在进行中，以确定可能具有临床适用性的Tbata功能阻滞剂。IL-7作为胸腺功能的可能调节因子的作用如上所述；通过表征T细胞亚群中IL-7受体的调节，进一步研究了其在外周稳态中的作用。IL-7受体调控的研究显示了亚群之间差异信号调节的复杂模式，其明显的效果是有利于原始原始T细胞的维持。这是特别有意义的，因为它为了解单个细胞因子(如IL-7)如何以不同方式调节多个不同的T细胞亚群提供了基础。具体地说，幼稚的CD4T细胞和记忆的CD4T细胞之间存在着不同的调节。IL-7受体远端信号的差异似乎是这种差异调节的机械原因。我们现在正在研究CD8初始记忆亚群和CD8记忆亚群中两种主要的动态平衡细胞因子对信号的生物学作用，并再次发现在T细胞受体周转水平上的差异。这些结果现已发表。我们还观察到，在人类依赖胸腺非依赖性途径的T细胞免疫重建中，这种扩张导致T细胞过早老化，从而导致细胞衰老，这可能是移植后T细胞功能损害的一个因素。在研究小鼠模型中的干细胞隔室时，我们发现Flt3配体通过与CXCR4和骨髓壁龛相互作用来调节胸腺前体细胞和造血干细胞。最后，在慢性移植物抗宿主病(GVHD)小鼠模型中，使用重氢葡萄糖评估T细胞亚群动力学的一系列实验获得了关于这些亚群的信息，并演变成成功地成像该疾病中快速增殖的细胞的努力。这项合作工作已经进展到对其他快速增殖的细胞，即肿瘤细胞的成像。该成像具有足够的敏感性和特异性，可供临床参考。它还可能通过基于增殖活性选择性摄取同位素而在疾病治疗中具有治疗应用。

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. 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 led in turn to a research emphasis on understanding mechanisms which control thymic function, and new treatments to treat cancer in the setting of a regenerating immune system. This work has progressed to the development 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. At high doses, IL-7 favors B cell development and so turns the thymus towards a B cell-poietic organ. 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. This is due to involvement of the protein encoded by Tbata in a large multi-protein complex which controls gene expression. Interestingly, this may be reflected in differences of thymopiesis in female versus male mice. The difference is now clear from multiple experiments, and the mechanism is under investigation. Experiments are in progress to identify blockers of Tbata function that might have clincial applicability. 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 characterizing IL-7 receptor regulation among T cell subsets. 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 and memory CD4 T cells. Differences in signaling distal to the receptor for IL-7 appear to mechanistically account for this differential regulation. We are now addressing the biology of signaling by the two primary homeostatic cytokines in the CD8 naive versus CD8 memory subsets in murine models and have found again differences at the level of T cell receptor turnover. These results are now published. 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 for years following transplant. Investigating the stem cell compartment in murine models, we showed that FLT3 ligand regulates thymic precursor cells and hematopoietic stem cells through interactions with CXCR4 and the marrow niche. Finally, a series of experiments using deuterated glucose to assess T cell subsets kinetics in the pathogenesis of chronic GVHD in murine models yielded information on such subsets and evolved into efforts to successfully image rapidly proliferating cells in that disease. This collaborative work has progressed to imaging of other rapidly proliferating cells, namely neoplastic cells. This imaging has sufficient sensitivity and specificity to be of clinical interest. It also may have therapeutic application in the treatment of disease by selective uptake of isotope based on proliferative activity.