Single-cell sequencing of peripheral blood cells in SLE patients

SLE 患者外周血细胞的单细胞测序

• 批准号: 9522104
• 负责人: Chun Jimmie Ye
• 金额: $ 23.78万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-14 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9522104
• 关键词: Affect; Alleles; Antinuclear Antibodies; Autoimmune Diseases; B-Lymphocytes; Behavior; Biological Assay; Biological Markers; Blood Cells; CD4 Positive T Lymphocytes; Cell physiology; Cells; Collaborations; Collection; Computational algorithm; Data; Data Set; Dendritic Cells; Disease; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Determinism; Genetic Transcription; Genotype; Immune; Immune response; Immunologics; Individual; Inflammation; Interferon-beta; Interferons; Lead; Light; Lupus; Lupus Nephritis; Mediating; Methods; Microfluidics; Molecular; Monitor; Nephritis; Noise; Outcome; Pathogenicity; Pathway interactions; Patients; Peripheral Blood Mononuclear Cell; Regulator Genes; Role; Running; Sampling; Science; Severities; Severity of illness; Signal Pathway; Signal Transduction; Sorting - Cell Movement; Systemic Lupus Erythematosus; T-Lymphocyte; Technology; Time; Transcriptional Regulation; Variant; base; cell type; cost; design; disease phenotype; experimental study; genetic information; genetic variant; genome wide association study; genome-wide; innovation; instrument; molecular pathology; nanolitre; next generation sequencing; novel strategies; pleiotropism; programs; response; single cell sequencing; single-cell RNA sequencing; therapeutic development; transcription factor; transcriptome sequencing; transcriptomics; treatment response

Systemic lupus erythematosus (SLE) has long been characterized by the prototypical expression of type-1 interferon induced genes (IFIGs). However, because of the pleiotropic roles of many IFIGs, how different peripheral blood mononuclear cells (PBMCs) mediate type-1 interferon signaling to cause disease is largely unknown. Here, we propose to use droplet-based RNA-sequencing to study the interferon response of PBMCs from lupus patients and healthy controls. Leveraging naturally occurring “genetic barcodes”, we will develop a highly innovative multiplexing strategy that significantly increases the throughput, reduces the cost, and limits unwanted technical noise of current droplet-based RNA-sequencing technologies. We will develop computational algorithms for assigning individual cells to the donor of origin and removing unwanted droplets containing multiple cells. We will use the multiplexing strategy to generate a rich dataset (~250K total cells) that enables, for the first time, the unbiased characterization of the effects of interferon-beta on PBMCs without sorting. We will first compare PBMCs from 8 healthy controls, 8 lupus and 8 lupus nephritis patients, to identify cell-type-specific interferon-beta response signatures that is predictive of disease status and severity. These signatures could be used to better monitor lupus progression and treatment response. By profiling PBMCs from 64 genotyped lupus patients, we will then characterize how common genetic variants affect cell-type-specific responses to interferon-beta, including expression parameters (e.g. variance across single cells) impossible to obtain from bulk RNA-sequencing. These results could be compared to genetic variants associated with lupus identified by genome-wide association studies to better understand the molecular pathology of the disease.

系统性红斑狼疮（SLE）的特点是1型的原型表达 干扰素诱导基因（IFIG）。然而，由于许多IFIG的多效性作用， 外周血单个核细胞（PBMC）介导1型干扰素信号传导而引起疾病， 未知在这里，我们建议使用基于液滴的RNA测序来研究PBMC的干扰素应答 狼疮患者和健康对照组的数据。利用天然存在的“基因条形码”，我们将开发一种 高度创新的多路复用策略，可显著提高吞吐量，降低成本， 目前基于液滴的RNA测序技术的不需要的技术噪音。我们将开发 用于将单个细胞分配给来源供体并去除不需要的液滴的计算算法 含有多个细胞。我们将使用多路复用策略来生成一个丰富的数据集（约250 K总细胞）， 第一次能够无偏地表征干扰素-β对PBMC的作用， 分类我们将首先比较来自8名健康对照、8名狼疮和8名狼疮性肾炎患者的PBMC，以确定 细胞类型特异性干扰素-β反应特征，其可预测疾病状态和严重程度。这些 标记可用于更好地监测狼疮进展和治疗反应。通过分析PBMC 从64名基因型狼疮患者中，我们将描述常见的遗传变异如何影响 对干扰素-β的细胞类型特异性应答，包括表达参数（例如，单个细胞间的方差）， 细胞），这是不可能从批量RNA测序获得的。这些结果可以与遗传变异进行比较 通过全基因组关联研究确定与狼疮相关，以更好地了解分子 疾病的病理学。