Development of a diagnostic platform for PTSD blood biomarkers

开发 PTSD 血液生物标志物诊断平台

• 批准号: 10455593
• 负责人: Michael S Breen
• 金额: $ 21.13万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10455593
• 关键词: Binding Sites; Biological; Biological Markers; Blood specimen; CD14 gene; CD19 gene; CD8B1 gene; Cells; Clinical; DNA Binding; Data; Development; Dexamethasone; Diagnostic; Discrimination; Disease; Etiology; Exposure to; Flow Cytometry; Future; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Glean; Glucocorticoid Receptor; Glucocorticoids; Goals; Hematological Disease; Heterogeneity; Hormones; Immune; Immune system; Impairment; Individual; Individual Differences; Inflammatory; Laboratory Finding; Light; Measures; Mediating; Modeling; Molecular; NCAM1 gene; Neurosecretory Systems; Outcome; Participant; Pathway interactions; Patients; Pattern; Peripheral; Peripheral Blood Mononuclear Cell; Post-Traumatic Stress Disorders; Predisposition; Production; Recovery; Regulation; Regulator Genes; Regulatory Element; Research; Resolution; Signal Transduction; Site; Stress; System; Techniques; Technology; Therapeutic; Tissues; Translating; Trauma; Variant; Veterans; Work; biological adaptation to stress; biomarker development; case control; cell type; chromatin immunoprecipitation; combat veteran; cytokine; density; diagnostic biomarker; diagnostic platform; diagnostic screening; differential expression; gene expression variation; genome-wide; hypothalamic-pituitary-adrenal axis; in vitro Model; indexing; insight; liquid biopsy; novel; post-trauma; receptor binding; receptor expression; resilience; response; stress reactivity; transcriptome; transcriptome sequencing; transcriptomics; trauma exposure; traumatic event

PROJECT SUMMARY Post-traumatic stress disorder (PTSD) is a disorder of stress reactivity with clinical manifestations that are evident when patients are triggered following exposure to a traumatic event. Impaired sensitivity to glucocorticoids (GCs) and alterations in peripheral blood mononuclear cell (PBMC) gene expression profiles, including genes implicated in GC signaling and inflammatory cytokine production, are at the crossroads of peripheral and central susceptibility pathways and represent promising PTSD biomarkers. However, their small case-control effect sizes have been a significant barrier to the development of actionable diagnostic biomarkers for the disorder. Notably, such studies have been performed post-trauma under baseline conditions and do not fully recapitulate the molecular response to adverse pathophysiological triggers, such as exposure to stress hormones. We developed a new in vitro model that sensitizes PBMCs to the synthetic GC, dexamethasone (DEX). Our model captures differences between individuals in sensitivity to GCs and significantly amplifies PTSD gene expression effect sizes into a range that may facilitate the development of actionable biomarkers. Still, cell type-specific differences in GC sensitivity have not been fully appreciated in PTSD and represent an important step forward for the development of translatable biomarkers and therapeutics. Here, we propose to combine our in vitro model with single-cell cellular indexing of transcriptomes and epitomes by sequencing (CITE-seq) and chromatin immunoprecipitation (ChIP)-seq to investigate the single-cell transcriptional responses to GC activation in PTSD. Specifically, we will investigate Aim 1) cell type-specific transcriptional differences using the CITE-seq approach on vehicle and DEX treated PBMCs derived from trauma-exposed combat veterans with PTSD(+; n=20) and without PTSD(-; n=20), as well as healthy trauma-free participants (HC; n=20). In doing so, we will study ~600,000 single cells (~5,000 cells per condition for each individual). Flow-cytometry will quantify the density and proportion of GR expression on five major immune cell types. Because the GR has site-specific DNA binding activity that differs between cells and conditions, we will Aim 2) perform anti-GR ChIP-seq data on five major immune cell subsets to identify genome-wide, cell type-specific GR binding sites. Integrating CITE-seq, GR expression and ChIP- seq across DEX exposed immune cell subsets will provide more nuanced and accurate insights into the cellular and molecular perturbations associated with altered GC sensitivity in PTSD. We have already measured genome-wide RNA-seq in bulk PBMCs treated with DEX and uncovered robust transcriptional differences distinguishing PTSD+ from PTSD-. Successful completion of this proposal will uncover cell type- specific transcriptional responses that will serve as definitive guides for the development of biomarkers for PTSD stress responsivity, vulnerability and resiliency.

项目概要 创伤后应激障碍（PTSD）是一种应激反应性障碍，其临床表现为 当患者在经历创伤事件后被触发时，这一点很明显。敏感性受损 糖皮质激素（GC）和外周血单核细胞（PBMC）基因表达谱的改变， 包括与 GC 信号传导和炎症细胞因子产生有关的基因，都处于十字路口 外周和中枢易感性途径，代表有前途的 PTSD 生物标志物。然而，他们的小 病例对照效应大小一直是开发可操作诊断的重大障碍 该疾病的生物标志物。值得注意的是，此类研究是在基线条件下进行的创伤后研究 并且没有完全概括对不良病理生理触发因素（例如暴露）的分子反应 压力荷尔蒙。我们开发了一种新的体外模型，使 PBMC 对合成 GC 敏感， 地塞米松（DEX）。我们的模型捕捉了个体对 GC 敏感性的差异 显着地将 PTSD 基因表达效应大小放大到可能促进发展的范围 可操作的生物标志物。尽管如此，GC 敏感性的细胞类型特异性差异尚未得到充分认识 PTSD 代表了可翻译生物标志物和药物开发的重要一步 疗法。在这里，我们建议将我们的体外模型与单细胞细胞索引相结合 通过测序 (CITE-seq) 和染色质免疫沉淀 (ChIP)-seq 获得转录组和表位组 研究 PTSD 中 GC 激活的单细胞转录反应。具体来说，我们将调查 目标 1) 使用 CITE-seq 方法对载体和 DEX 处理的细胞类型特异性转录差异 PBMC 源自患有 PTSD（+；n=20）和无 PTSD（-；n=20）的遭受创伤的退伍军人，如 以及健康无创伤的参与者（HC；n=20）。在此过程中，我们将研究约 600,000 个单细胞（约 5,000 每个人每个条件的细胞）。流式细胞术将量化 GR 的密度和比例 五种主要免疫细胞类型的表达。因为 GR 具有不同的位点特异性 DNA 结合活性 在细胞和条件之间，我们的目标是 2) 对五个主要免疫细胞亚群进行抗 GR ChIP-seq 数据 识别全基因组、细胞类型特异性的 GR 结合位点。整合 CITE-seq、GR 表达和 ChIP- 对 DEX 暴露的免疫细胞子集进行 seq 将提供更细致、更准确的见解 与 PTSD 中 GC 敏感性改变相关的细胞和分子扰动。我们已经 测量了用 DEX 处理的批量 PBMC 的全基因组 RNA-seq 并发现了强大的转录 区分 PTSD+ 和 PTSD- 的差异。成功完成该提案将揭示细胞类型- 特定的转录反应将作为开发生物标志物的明确指南 PTSD 压力反应性、脆弱性和弹性。