Pathogenic T Cells in Chronic Beryllium Disease

描述（由申请人提供）：慢性铍病（CBD）是一种由工作场所铍暴露引起的肉芽肿性肺部疾病，其特征是肺中铍特异性CD4+ T细胞的积累。由于其独特的化学和物理性质，铍继续在高科技工业中得到利用。因此，CBD仍然是一个重要的公共卫生问题，超过100万美国工人接触过铍并有患病风险。由于存在已知抗原和可及的靶器官，CBD是免疫介导的器官破坏的重要模型。我们和其他人已经表明，最重要的HLA分子铍呈现是HLA- dp。在表达突变HLA-DP2分子的成纤维细胞中，铍的识别依赖于HLA-DP2链第69位（2Glu69）的谷氨酸残基。此外，T细胞对铍的识别发生在没有抗原处理的情况下，核磁共振明确显示可溶性HLA-DP2直接结合铍。当前应用的研究目标是阐明铍特异性CD4+ T细胞在HLA-DP2环境下识别铍的机制，并证明铍特异性记忆T细胞池的稳定性。2Glu69如何影响铍识别的最可能的可能性是：1)铍直接与HLA-DP2分子中2Glu69的羧酸盐结合，而肽只需要完成12TCR配体；2)2Glu69对铍呈递的影响是间接的，通过它影响可以与DP2结合的肽库，其中包括可以呈递铍的肽。我们的初步数据支持第一种假设，并表明我们可以在了解哪些肽对T细胞识别DP2-肽/Be2+复合物很重要之前研究铍与HLA-DP2的结合。第一个具体目标将提供明确的证据铍结合与x射线晶体学HLA-DP2有和没有铍。在第二个目标中，HLA-DP2突变体的产生将作为与第一个目标中的结构研究的功能关联。第三个目标将描述完成12TCR配体所需的肽，而最后一个特定目标将确定从铍致敏到疾病的进展是否与血液中铍特异性CD4+ T细胞频率增加有关。总之，这些研究将加强我们对抗原如何引起肉芽肿性炎症的理解，特别是金属抗原如何引发免疫反应。此外，它们将潜在地允许开发生物标志物来预测高风险受试者的疾病进展。这项转化性研究将利用职业性肺病患者的血液和肺标本，进一步了解铍诱导的导致致残性肺功能障碍的免疫机制。在以肺功能进行性丧失为特征的疾病中，识别疾病进展的潜在生物标志物将极大地促进这一患者群体的福利。

DESCRIPTION (provided by applicant): Chronic beryllium disease (CBD) is a granulomatous lung disorder caused by beryllium exposure in the workplace and is characterized by the accumulation of beryllium-specific CD4+ T cells in the lung. Due to its unique chemical and physical properties, beryllium continues to be utilized in high-technology industries. Thus, CBD remains an important public health concern with more than 1,000,000 US workers having been exposed to beryllium and at risk for disease development. With the presence of a known antigen and an accessible target organ, CBD is an important model of immune-mediated, organ destruction. We and others have shown that the most important HLA molecule for beryllium presentation is HLA-DP. Using fibroblasts expressing mutated HLA-DP2 molecules, beryllium recognition was dependent on the glutamic acid residue at position 69 (2Glu69) of the HLA-DP 2-chain. In addition, T cell recognition of beryllium occurred in the absence of antigen processing, and nuclear magnetic resonance definitively showed that soluble HLA-DP2 directly bound beryllium. The goals of the studies in the current application are to elucidate the mechanism by which beryllium-specific CD4+ T cells recognize beryllium in the context of HLA-DP2 and to demonstrate the stability of the beryllium-specific memory T cell pool. The most likely possibilities of how 2Glu69 influences beryllium recognition are that 1) beryllium directly binds to the carboxylate of 2Glu69 in HLA-DP2 molecules, with peptide(s) only required to complete the 12TCR ligand or that 2) the effect of 2Glu69 on beryllium presentation is indirect through its influence on the repertoire of peptides that can bind to DP2, among which are peptides that can present beryllium. Our preliminary data favor the first hypothesis and suggest that we can study beryllium binding to HLA-DP2 prior to knowing which peptides are important for T cell recognition of the DP2- peptide/Be2+ complex. The first specific aim will provide definitive proof of beryllium binding with x-ray crystallography of HLA-DP2 with and without beryllium. The generation of HLA-DP2 mutants in the second aim will serve as a functional correlate to the structural studies in Aim #1. The third aim will delineate which peptides are required to complete the 12TCR ligand while the final specific aim will determine whether progression from beryllium sensitization to disease is associated with an increased frequency of beryllium- specific CD4+ T cells in blood. Together, these studies will strengthen our understanding of how antigens cause granulomatous inflammation and specifically how metal antigens trigger an immune response. In addition, they will potentially allow the development of biomarkers to predict disease progression in high-risk subjects. PUBLIC HEALTH RELEVANCE This translational study will utilize blood and lung specimens from human subjects with an occupational lung disorder to further our understanding of the beryllium-induced immune mechanisms that lead to disabling lung dysfunction. In a disease characterized by progressive loss of lung function, the identification of potential biomarkers of disease progression will greatly advance the welfare of this patient population.