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: 9372979
• 负责人: Chun Jimmie Ye
• 金额: $ 19.81万
• 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/9372979
• 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; 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.

全身性红斑狼疮（SLE）长期以来一直以1型的典型表达为特征 干扰素诱导的基因（IFIG）。但是，由于许多IFIG的多效性角色，多么不同 外周血单核细胞（PBMC）介导1型干扰素信号引起疾病，在很大程度上是 未知。在这里，我们建议使用基于液滴的RNA测序来研究PBMC的干扰素响应 来自狼疮患者和健康对照。利用自然发生的“遗传条形码”，我们将发展一个 高度创新的多路复用策略可显着增加吞吐量，降低成本和限制 当前基于液滴的RNA序列技术的技术噪声。我们将发展 用于分配单个单元格的计算算法并去除不需要的液滴 包含多个细胞。我们将使用多路复用策略来生成一个丰富的数据集（约250k总单元格） 首次启用干扰素β对PBMC的影响的公正表征 排序。我们将首先比较来自8个健康对照组，8个狼疮和8名狼疮患者的PBMC，以鉴定 细胞型特异性干扰β反应签名，可预测疾病状况和严重程度。这些 可以使用签名来更好地监测狼疮的进展和治疗反应。通过分析PBMC 从64名基因分型狼疮患者中，我们将表征常见的遗传变异如何影响 细胞类型对干扰素β的反应，包括表达参数（例如，单一方差 细胞）不可能从散装RNA测序中获得。可以将这些结果与遗传变异进行比较 与全基因组关联研究确定的狼疮有关，以更好地了解分子 疾病的病理。