Single cell analysis of gene expression in human vascular cells

项目摘要 在我的临床训练中，我反复看到血管疾病对 病人们正处于他们的壮年时期。血管疾病，如冠状动脉疾病 (CAD)、中风、动脉夹层和偏头痛加在一起导致了超过一半的 美国的死亡和残疾。要根除血管疾病，重要的是 开发针对疾病开始部位的动脉细胞的新疗法。我的K08- 资助的研究一直是利用人类基因变异来识别这些新的途径 作为一个向导。最近，与多种血管疾病相关的基因座 通过全基因组关联研究(GWAS)确定。这些基因座代表着新的 治疗靶点，但其生物学机制在很大程度上仍未被探索。我有过 确认了染色体上的一种血管疾病相关变异的机制 6p24在远端调节内皮素-1的表达。这种非编码的调节作用 Variant是血管组织特有的，主要见于内皮细胞。是否其他 血管中的细胞负责该基因座的其他基因调控作用。 仍然不为人知。对于这个和其他血管疾病相关的基因座来说，它将是重要的 描述相关细胞类型或细胞中的基因调控效应 亚群。单细胞RNA测序的新方法 动脉壁中的全套细胞。基于液滴的单细胞RNA测序允许 分析数千个细胞并检测稀有细胞亚群。使用这个 方法，我已经鉴定了三种不同的内皮细胞亚群 暗示功能特化的转录签名。此R03应用程序 试图将这一发现扩展到人类血管组织。这项提议的目标是 确定单细胞RNA测序的最佳方法和确定标记 人类血管组织中内皮细胞的异质性。这将提供一份地图集 基于血管组织中转录签名的细胞类型，并作为 未来功能基因组学实验的基础资源。拥有更好的 了解动脉壁中的细胞类型和基因表达特征 有可能确定治疗和预防该病的新生物途径 血管疾病。

Project Summary In my clinical training, I repeatedly saw the devastating effects of vascular disease on patients in the prime of their lives. Vascular diseases such as coronary artery disease (CAD), stroke, arterial dissection, and migraine headache combine to cause over half the death and disability in the United States. To eradicate vascular disease it will be important to develop new treatments that target the arterial cells where the disease begins. My K08- funded research has been to identify these new pathways using human genetic variation as a guide. The loci associated with multiple vascular diseases have recently been identified through genome-wide association studies (GWAS). These loci represent new therapeutic targets, but their biological mechanisms remain largely unexplored. I have identified the mechanism by which a vascular disease associated variant on chromosome 6p24 distally regulates endothelin-1 expression. This regulatory effect of the non-coding variant is vascular tissue specific, and largely seen in endothelial cells. Whether other cells in the blood vessel are responsible for other gene regulatory effects for this locus remains unknown. For this and other vascular disease-associated loci it will be important to characterize the gene regulatory effects in the relevant cell type or cellular subpopulation. New methods in single cell RNA-sequencing all the identification of the full set of cells in the arterial wall. Droplet-based single cell RNA-sequencing allows for analysis of thousands of cells and detection of rare cellular subpopulations. Using this method, I have identified three distinct subpopulations of endothelial cells with transcriptional signatures that suggest functional specialization. This R03 application seeks to extend this finding to human vascular tissue. The goals of this proposal are to determine the best method for single cell RNA-sequencing and to determine the markers of endothelial cell heterogeneity in human vascular tissue. This will provide an atlas of cell types based on transcriptional signature in vascular tissue and serve as a foundational resource for future functional genomics experiments. With a better understanding of the cell types and gene expression signatures in the arterial wall it will be possible to identify new biological pathways for the treatment and prevention of vascular disease.