The immunoregulatory role of Alveolar Macrophages in Chronic Beryllium Disease

项目概要/摘要： 本研究的目的是确定CD 55和JAK/STAT通路失调对人肺癌的影响。 肺泡巨噬细胞（AM）对慢性铍病（CBD）进展的影响。CBD是重要的 器官特异性免疫介导的疾病，以肉芽肿性肺部炎症、纤维化和 因终末期呼吸功能不全死亡。因此，CBD仍然是一个重要的公共卫生问题。 这种疾病的前兆（铍致敏; BeS）以约6-8%的速度进展为CBD 每一年。虽然Be通过含β Glu 69的HLA-DP分子与致病性 CD 4 + T细胞为DP 2与CBD和BeS的遗传连锁提供了解释，但这并不能解释DP 2与CBD和BeS的遗传连锁。 BeS向CBD的发展。此外，在没有BeS证据的Be-workers中发现高达40%的DP 2， CBD.因此，在BeS向CBD的进展中必须涉及额外的机制。我们的假设是 CBD AM下调免疫负调节基因CD 55及其通路，并过度表达 正调节激活JAK/STAT途径，其增强对Be的免疫应答， 从BeS到CBD。在目标1中，我们将确定AM上的CD 55是否是AM中的负调节因子。 BeS患者AM对Be的免疫应答。具体地，使用CD 55的过表达， 使用腺病毒过表达系统和激动性抗体减少CD 55基因或使用 腺病毒敲减系统和CD 55中和抗体，我们将确定改变 CD 55对TNF-α产生、表型（CD 16、CD 40、CD 80、CD 86、HLA- DR）、JAK/STAT活化和BeLPT，比较CBD、BeS和对照。最后，使用RNA-seq，我们将 研究与CBD相比BeS中CD 55下调相关的调控网络; 目的2：我们将确定JAK/STAT通路是否是AM对Be免疫应答的正调节因子 与BeS相比，来自CBD受试者的AM中的含量增加。具体而言，CBD的AM将 与来自BeS和对照受试者的那些相比，评估JAK/STAT的过表达， 使用腺病毒过表达系统或JAK/STAT途径基因的减少 使用腺病毒敲低系统和药理学JAK 1 - 3抑制剂对Be免疫应答的影响。我们 将评价TNF-α产生、表型（CD 16、CD 40、CD 80、CD 86、HLA-DR）和BeLPT的变化。 最后，使用RNA-seq，我们将定义与过度激活的调控网络。 JAK/STAT通路。在目标3中，我们将研究CD 55和JAK /STAT通路是否相关， 与BeS相比，CBD和从BeS到CBD的进展，使用纵向研究并验证这些 在更大的纵向人群中的发现，以确定这些基因和途径是否揭示了潜在的新的 CBD的诊断和预后的生物标志物。在这个项目结束时，我们将确定关键的监管 AM中与疾病相关的免疫缺陷介导的免疫失调的基因和途径 进展并揭示用于临床预后和诊断的潜在生物标志物。由此获得的结果 这项研究将使我们更了解CBD发展的因素，以及 治疗，并将作为一个模型的其他免疫相关的免疫反应和环境诱导的 慢性病

Project Summary/Abstract: The goal of this study is to determine the impact of CD55 and JAK/STAT pathway dysregulation in alveolar macrophages (AMs) on chronic beryllium disease (CBD) progression. CBD is an important organ-specific immune-mediated disease, characterized by granulomatous lung inflammation, fibrosis, and death, due to end-stage respiratory insufficiency. Thus, CBD remains an important public health concern. The precursor to this disease (beryllium sensitization; BeS) progresses to CBD at a rate of approximately 6-8% per year. Although the binding and presentation of Be via βGlu69-containing HLA-DP molecules to pathogenic CD4+ T cells provides an explanation for the genetic linkage of DP2 to CBD and BeS, this does not explain the progression of BeS to CBD. Furthermore, DP2 is found in up to 40% of Be-workers without evidence of BeS or CBD. Thus, additional mechanisms must be involved in progression of BeS to CBD. Our hypothesis is that CBD AMs downregulate the negative immunoregulatory gene CD55 and its pathway and overexpress the positive regulatory activating JAK/STAT pathway, which augments the immune response to Be and progression from BeS to CBD. In Aim 1, we will define if CD55 on AMs is a negative regulator in the immune response to Be in AMs from BeS patients. Specifically, using either the overexpression of the CD55 genes using a adenovirus overexpression system and agonistic antibody or the reduction of CD55 genes using adenovirus knockdown system and neutralizing antibody of CD55, we will determine the impact of alteration of CD55 on consequential functional changes in TNF-α production, phenotype (CD16, CD40, CD80, CD86, HLA- DR), JAK/STAT activation and BeLPT comparing CBD, BeS and controls. Finally, using RNA-seq, we will investigate the regulatory networks associated with the downregulation of CD55 in CBD compared to BeS; In Aim 2, we will determine if the JAK/STAT pathway is a positive regulator of the immune response to Be in AMs and is increased in AMs from subjects with CBD compared to BeS. Specifically, AMs from CBD will be compared to those from BeS and controls subjects, assessing either the overexpression of the JAK/STAT pathway genes using a adenovirus overexpression system or the reduction of the JAK/STAT pathway genes using adenovirus knockdown system and pharmacological JAK1- 3 inhibitors on the Be immune response. We will evaluate the changes in TNF-α production, phenotype (CD16, CD40, CD80, CD86, HLA-DR), and BeLPT. Finally, using RNA-seq we will define the regulatory networks associated with the overactivation of the JAK/STAT pathway. In Aim 3, we will investigate whether the CD55 and JAK /STAT pathway are associated with CBD and progression from BeS to CBD, compared to BeS, using a longitudinal study and validate these findings in a larger longitudinal population to determine if these genes and pathways reveal potential novel biomarkers of diagnosis and prognosis for CBD. At the end of this project we will determine the key regulatory genes and pathways of exposure-mediated immune dysregulation in AMs that are associated with disease progression and reveal potential biomarkers for clinical prognosis and diagnosis. The results obtained from this study will improve our understanding of factors involved in the development of CBD, as well as targets for therapy, and will serve as a model of other exposure-related immune responses and environmentally-induced chronic diseases.