Neural-derived extracellular vesicles microRNAs in bipolar disorder: a peripheral window into the brain

项目摘要/摘要 双相情感障碍是一种慢性的、通常是严重的精神疾病，有很强的遗传基础。然而，即使是 随着分子遗传学研究的重大进展，与这种疾病相关的少数遗传变异 只解释了其遗传性的一小部分，这导致了表观遗传学参与的假说 BD的病理生理学改变。虽然最近的研究试图识别表观遗传生物标记物 BD，大多数结果是有限的，因为表观遗传改变的组织特异性和对 通过微阵列进行候选基因或全基因组研究。自从寻找生物标记物以来 反复关注不一定反映大脑变化的外周测量，有意义 与生物学和临床相关的发现受到了极大的阻碍。这项研究将填补这一重要空白 通过测量大脑释放的血浆来源的细胞外小泡(EVS)中的表观遗传标记。 具体地说，我们将关注最近提出的microRNAs和其他非编码RNAs 介导BD的重要机制，并可能整合基因和环境刺激。值得注意的是，microRNAs- 填充的电动汽车是由神经组织释放的，作为细胞间通讯的一种方式，可能是关键角色 在将表观遗传标记转移到生殖细胞的过程中，最终有助于代际和跨代 BD的传播。我们的工作假设是BD患者会表现出特定的miRNAs和 血浆神经源性EVS中其他非编码RNA转录本与对照组的比较。为了测试这一点，我们首先将 通过分析60名健康对照和60名BD患者的血液样本鉴定BD中神经特异性EVS microRNAs I型患者(已在我科收集和储存)(目标1)。在神经源性电动汽车之后 免疫沉淀和鉴定，RNA将被分离，下一代测序文库将被 在Illumina NextSeq仪器上制备和测序，单端读数为1x75bp，测序速度约为 每个样本的深度为1,000-1,500万次读取。显著改变的miRNAs将通过实时聚合酶链式反应和 差异表达转录本的假定生物学相关性将通过功能途径进行评估 分析。此外，还将使用miRNA表达数据建立机器学习模型，以便预测 无论个体样本属于BD组还是对照组，允许建立临床 有用的预测性生物签名，可在现场立即产生影响。在第二步，我们将调查 神经元EVS转录组标志物与临床、神经解剖学和神经认知的相关性 可用于每个受试者的参数，最终目标是确定新的- 确定疾病内表型的EVS标记物(目标2)。这种脑特异性标志物的鉴定 可能会为BD的科学研究开辟新的途径。

Project Summary/Abstract Bipolar disorder is a chronic and often severe psychiatric disorder with a strong genetic basis. However, even with significant progress made by molecular genetic studies, the few genetic variants associated with the disorder explain only a small portion of its heritability, which led to the hypothesis of the involvement of epigenetic alterations in BD’s pathophysiology. While recent studies have attempted to identify epigenetic biomarkers in BD, most results are limited due to the tissue specificity of epigenetic alterations and the biased assessments of candidate genes or genome-wide investigations through microarrays. Since the search for biomarkers have repetitively focused on peripheral measures that do not necessarily reflect brain alterations, significant biologically- and clinically-relevant findings have been significantly hindered. This study will fill this important gap by measuring epigenetic markers in plasma-derived extracellular vesicles (EVs) released by the brain. Specifically, we will focus on microRNAs and other non-coding RNAs that have been recently proposed to mediate important mechanisms in BD and may integrate gene and environment stimuli. Of note, microRNAs- filled EVs are released by the neural tissue as a method of cell-to-cell communication and may be the key players in transferring epigenetic markers to germ cells, ultimately contributing to the inter- and transgenerational transmission of BD. Our working hypothesis is that patients with BD will show alterations in specific miRNAs and other non-coding RNA transcripts in plasma neural-derived EVs compared to controls. To test this, we will initially identify neural-specific EVs microRNAs in BD by analyzing blood samples from 60 healthy controls and 60 BD type I patients (already collected and stored in our Department) (Aim 1). After neural-derived EVs immunoprecipitation and characterization, RNA will be isolated and next generation sequencing libraries will be prepared and sequenced on an Illumina NextSeq instrument with 1x75 bp single-end reads at an approximate depth of 10-15 million reads per sample. Significantly altered miRNAs will be validated by real-time PCR and the putative biological relevance of the differentially expressed transcripts will be assessed by functional pathway analyses. In addition, a machine learning model will be built using the miRNA expression data in order to predict whether an individual sample belongs to the BD or control group, allowing for the establishment of a clinically useful predictive biosignature that can have immediate impact in the field. On a second step we will investigate the correlation between neuronal EVs transcriptome markers with clinical, neuroanatomical, and neurocognitive parameters available for each subject, with the ultimate goal of identifying the clinical relevance of the newly- identified EVs markers in endophenotypes of illness (Aim 2). The identification of such brain-specific markers will likely open new avenues for scientific investigation of BD.