Sex-specific genomic mechanisms of transcriptional regulation in ME/CFS/SEID

 描述(由申请人提供)：目前对肉豆蔻性脑脊髓炎/慢性疲劳综合征/全身性劳力不耐受疾病(ME/CFS/SEID)的治疗完全依赖症状管理来提高生活质量，但没有解决疾病发生和发展的基本机制。为了更好地了解ME/CFS/SEID涉及的关键生物学靶点，本研究提案的主要目的是识别ME/CFS/SEID的男性特异性生物标记物和治疗靶点，并洞察特定性别疾病的发生和发展，这将导致更好的治疗干预。这项研究提案旨在使用我们最近为NIH资助的研究(1R01NS090200-01(Pi Fletcher))招募的患者的外周血单个核细胞(PBMC)。这些PBMC是从男性ME/CFS/SEID患者和匹配的健康对照组的三个时间点--运动前、最大摄氧量(VO2 Max)和运动后4小时分离出来的。这项建议的具体目标包括：具体目标1。确定代谢途径和可能影响男性ME/CFS/SEID的调节RNA。我们的假设是，未知的转录本和选择性剪接事件调节负责炎症反应、白细胞迁移和细胞发育调节的基因表达的变化。我们的目标是使用RNA-SEQ方法来识别已知和新的转录本的丰度，并在男性患者的PBMC中发现新的剪接事件。这将允许建立一组特定性别的ME/CFS/SEID候选成绩单。然后，我们将使用NanoString Technologies的nCounter系统验证它们，并创建一个可能用于ME/CFS诊断和管理的记录小组。具体目的2.探讨男性ME/CFS/SEID转录调控的可能机制。为了更好地了解男性ME/CFS/SEID患者基因表达变化的原因，我们旨在确定拷贝数变异(CNV)和基因组DNA甲基化的变化。我们将使用Agilent SurePrint G3人类基因组CGH+SNP微阵列来鉴定一组ME/CFS/SEID性别特异性候选CNV。我们将使用Illumina Infinium人类甲基化450珠芯片微阵列来评估ME/CFS/SEID患者和健康对照组之间甲基化模式的差异。特定目的3.比较男性和女性与ME/CFS之间受影响的代谢途径和转录调控机制。我们将比较ME/CFS/SEID患者和HC在男性中的差异，女性也有相同的差异(来自最近资助的拨款提案1 R15 NS087604-01A1，Pi Nathanson)。将对基因表达、选择性剪接、基因组DNA甲基化模式和CNV的结果进行比较。我们期望发现参与炎症反应和免疫系统过程的基因的调控方面的差异，特别是在细胞因子的产生和活性的调控方面。

 DESCRIPTION (provided by applicant): Current management of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome/Systemic Exertion Intolerance Disease (ME/CFS/SEID) relies solely on symptom management to improve quality of life but does not address the underlying mechanisms of disease onset and progression. To provide better insight into the key biological targets involved ME/CFS/SEID presentation, the main objective of this research proposal is to identify male-specific biomarkers and therapeutic targets of ME/CFS/SEID and provide insight into sex-specific disease onset and progression, which will lead to the better therapeutic intervention. This research proposal aims to use Peripheral Blood Mononuclear Cells (PBMC) from patients recruited for our recently NIH funded research (1R01NS090200-01 (PI Fletcher)). These PBMC have been isolated from male ME/CFS/SEID patients and matched healthy controls at three time points - before exercise challenge, at the peak of effort (VO2 max) and four hours after the peak effort. Specific Aims of this proposal include: Specific Aim 1. Identify metabolic pathways and possible regulatory RNAs affected in men with ME/CFS/SEID. Our hypothesis is that unknown transcripts and alternative splicing events regulate changes in expression of genes responsible for inflammatory response, leukocyte migration, and regulation of cell development. We aim to use an RNA-seq approach to identify the abundance of known and novel transcripts and discover new splicing events in PBMC of male patients. This will allow establishing a set of sex-specific ME/CFS/SEID candidate transcripts. We will then validate them using NanoString Technologies' nCounter system and create a panel of transcripts to potentially be used in the diagnosis and management of ME/CFS. Specific Aim 2. To investigate possible mechanisms of transcriptional regulation in men with ME/CFS/SEID. To better understand causes of changes in gene expression in male patients with ME/CFS/SEID, we aim to identify changes in copy number variation (CNV) and genomic DNA methylation. We will use Agilent SurePrint G3 Human Genome CGH+SNP microarrays to identify a set of ME/CFS/SEID sex specific candidate CNVs. We will use Illumina Infinium Human Methylation450 BeadChip microarrays to evaluate differences in the methylation patterns between ME/CFS/SEID patients and healthy controls. Specific Aim 3. Compare affected metabolic pathways and mechanisms of transcriptional regulation in men and women with ME/CFS. We will compare differences between ME/CFS/SEID patients and HC in men with the same differences in women (from the recently funded grant proposal 1 R15 NS087604-01A1, PI Nathanson). Comparisons will be done for the results in gene expression, alternative splicing, patterns on the genomic DNA methylation and CNVs. We expect to find differences in the regulation of genes involved in inflammatory response and immune system processes, particularly in the regulation of cytokines production and activity.