Epigenetic Alterations in IPF Fibroblastic Foci

描述（由申请人提供）： 特发性肺纤维化（IPF）是一种没有明确病因的致命疾病。由于发病率随着年龄的增长而增加，因此环境影响可能很重要。最近的数据表明IPF中发育基因标签的再表达。表观遗传调控，包括DNA甲基化和组蛋白修饰，通常发生在发育过程中，并响应于衰老和环境刺激，导致基因表达的可遗传变化，而不依赖于基因本身的任何变异。富含胞嘧啶-鸟嘌呤（CpG岛）的基因启动子区域的DNA甲基化和组蛋白修饰都在基因沉默中发挥重要作用;这些范例在发育期间的细胞分化中很重要，并且在许多癌症中的肿瘤抑制因子沉默中至关重要。表观遗传修饰是潜在可逆的，因此是开发新疗法的有吸引力的靶点。尽管越来越多的证据表明表观遗传调控可能在纤维化疾病中发挥重要作用，但很少有研究探讨IPF的表观遗传调控。我们的实验室已经确定，抑制成纤维细胞纤维化表型的Thy-1在IPF的成纤维细胞病灶中不存在，其缺失可能是由Thy-1启动子区域的甲基化引起的，并且是可逆的。我们实验室的其他初步数据表明，已知在IPF中下调的其他基因的表观遗传沉默。我们假设表观遗传变化改变了IPF的基因表达谱，促进了纤维化。这项研究将确定DNA甲基化是IPF发病机制中的一个重要因素，并为进一步探索这种致命疾病的表观遗传学改变提供支持，从而为开发新的治疗方法奠定基础。该提案将利用肺组织研究联盟（LTRC）提供的已充分表征的组织，与慢性阻塞性肺疾病（COPD）相比，对IPF中DNA甲基化的总体和基因特异性改变进行表征，以确定表观遗传学改变是IPF的重要致病机制。本研究的目的是：1。通过以下方式定义与COPD和对照肺组织相比，IPF中DNA甲基化与基因表达的关系：a）IPF、COPD和对照组织中基因表达的微阵列分析，B）这些样品中的甲基化阵列分析，和c）通过甲基化特异性PCR确认选定的基因;和2.通过以下方式测量IPF和COPD组织中整体DNA甲基化的区域变化：a）抗甲基胞嘧啶的定量分析和IHC，以及B）局部组织水平DNA甲基转移酶（DNMT）和其他DNA甲基化调节因子的分析。这些研究将确定表观遗传学改变，特别是DNA甲基化变化在IPF中发生的程度。这一重要且以前未探索的范例可能会对这种毁灭性疾病的发病机制产生重要的额外见解，并确定IPF和其他纤维化疾病的新治疗靶点。公共卫生相关性：特发性肺纤维化（IPF）是一种致命的不治之症，影响超过100，000名美国人，其原因不明，并在肺部产生进行性瘢痕。在人类发育过程中，有一种控制基因表达的生物学机制，称为表观遗传调控，越来越多地发现它在癌症和其他疾病中异常。该项目将利用肺组织研究联盟（LTRC）提供的充分表征的IPF组织，以确定表观遗传异常在多大程度上导致IPF中观察到的不受控制的瘢痕形成，从而为开发这种毁灭性疾病的新疗法开辟道路。(End摘要）

DESCRIPTION (provided by applicant): Idiopathic Pulmonary Fibrosis (IPF) is a deadly disease without a clearly-defined etiology. Because the incidence increases with age, it is plausible that environmental effects may be important. Recent data indicate re-expression of developmental gene signatures in IPF. Epigenetic regulation, including DNA methylation and histone modification, which often occurs during development and in response to aging and environmental stimuli, results in heritable changes in gene expression independent of any variation in the genes themselves. DNA methylation at gene promoter regions rich in cytosine-guanine (CpG islands) and histone modifications both play important roles in gene silencing; these paradigms are important in cellular differentiation during development, and are critical in silencing of tumor suppressors in many cancers. Epigenetic modifications are potentially reversible and therefore attractive targets for development of novel therapies. Although there is increasing evidence that epigenetic regulation may play an important role in fibrotic disease, few studies have explored epigenetic regulation in IPF. Our lab has established that Thy-1, which suppresses the fibrotic phenotype of fibroblasts, is absent in fibroblastic foci of IPF, that its absence can be caused by methylation at the Thy-1 promoter region and is reversible. Additional preliminary data from our lab demonstrates epigenetic silencing of other genes known to be downregulated in IPF. We hypothesize that epigenetic changes alter the gene expression profile in IPF, promoting fibrogenesis. This study will establish DNA methylation as an important factor in the pathogenesis of IPF, and provide support for further exploration of epigenetic alterations in this deadly disease, thus setting the stage for development of novel therapeutic approaches. This proposal will leverage the availability of well-characterized tissues available through the Lung Tissue Research Consortium (LTRC) to characterize global and gene-specific alterations in DNA methylation in IPF in comparison to chronic obstructive pulmonary disease (COPD), in order to establish epigenetic alterations as an important pathogenic mechanism in IPF. The aims of the study will be to: 1. Define the relationship of DNA methylation to gene expression in IPF in comparison to COPD and control lung tissue, by: a) microarray analysis of gene expression in IPF, COPD, and control tissues, b) methylation array analysis in these samples, and c) confirmation of selected genes by methylation-specific PCR; and 2. Measure regional alterations of global DNA methylation in IPF and COPD tissues, by: a) quantitative analysis and IHC for anti-methylcytosine, and b) analysis of DNA methyltranserases (DNMTs) and other regulators of DNA methylation at the local tissue level. These studies will define the extent to which epigenetic alterations, specifically changes in DNA methylation, occur in IPF. This important and previously unexplored paradigm is likely to yield significant additional insight into the pathogenesis of this devastating disease, as well as to define novel therapeutic targets for IPF and other fibrotic disorders. PUBLIC HEALTH RELEVANCE: Idiopathic Pulmonary Fibrosis (IPF) is a deadly incurable disease affecting over 100,000 Americans, the cause of which is unknown, and which produces progressive scarring in the lungs. There is a biological mechanism for controlling expression of genes during human development known as epigenetic regulation, which is increasingly found to be abnormal in cancer and other diseases. This project will take advantage of the well characterized IPF tissues available through the Lung Tissue Research Consortium (LTRC) in order to determine the extent to which epigenetic abnormalities are responsible for the uncontrolled scarring seen in IPF, thus leading the way to development of novel therapies for this devastating disease. (End of Abstract)