The Single Cell Landscape of Early Human Diabetic Nephropathy

项目摘要/摘要 糖尿病肾病是终末期肾病的主要原因，也是发病率的主要因素。 和死亡率。我们已经成功地进行了单核RNA测序(SnRNAseq)和单 5例健康人和8例糖尿病患者肾组织细胞核ATAC测序结果分析 人糖尿病早期肾脏细胞类型特异性转录和染色质构象的研究 疾病。糖尿病近端小管差异表达的转录本显示 糖异生基因和糖皮质激素信号通路的丰富。这项建议旨在 集成SnRNAseq和SnATACseq以确定在 调节糖异生的糖皮质激素敏感基因的增强子和启动子区。我们会 随后，用糖尿病损伤的体外模型和糖尿病小鼠模型验证我们在体内的发现。 这项建议建立在首席研究员以前的研究经验和临床培训的基础上。 目前，帕克·威尔逊博士将25%的时间用于肾脏和分子病理学临床服务。 剩下的75%分配给本杰明·汉弗莱斯博士的实验室进行基础研究。威尔逊医生有一种 与Humphreys博士建立了指导关系，并发表了他对SnRNAseq数据的分析 发表在《美国国家科学院院刊》上的人类糖尿病肾脏。此外，威尔逊博士还有一位最近被接受的第一作者 描述多模式SnRNAseq和SnATACseq整合在健康成人肾脏中的手稿 通讯。这些发现为这一应用程序提供了基础，该应用程序将专注于扩展Dr。 威尔逊在单细胞方法、生物信息学分析和糖尿病肾损伤模型方面的科学技能。 职业发展目标将通过汉弗莱斯博士的指导和一项咨询来实现 在染色质构象调控、生物信息学和糖尿病肾病方面具有专长的委员会。Dr。 威尔逊将承担研究伦理、科学交流和拨款写作方面的教学课程， 分子生物学和高级计算机编程，以加深他现有的知识基础。这项工作将 在华盛顿大学举行，这所大学有着指导成功的内科科学家的悠久历史。 完成这项职业发展奖将为威尔逊博士的职业发展奠定坚实的基础 独立性和R01级资金。

Project Summary/Abstract Diabetic kidney disease is the leading cause of end-stage renal disease and a major contributor to morbidity and mortality. We have successfully performed single nucleus RNA sequencing (snRNAseq) and single nucleus ATAC sequencing (snATACseq) on five healthy control and eight diabetic kidney samples to measure the cell-type-specific transcriptional and chromatin conformational profile of early human diabetic kidney disease. The differentially expressed transcripts in the diabetic proximal tubule showed upregulation of gluconeogenic genes and enrichment of pathways involved in corticosteroid signaling. This proposal aims to integrate snRNAseq and snATACseq to determine whether there are changes in chromatin accessibility in the enhancer and promoter regions of corticosteroid-sensitive genes that regulate gluconeogenesis. We will subsequently validate our in vivo findings with an in vitro model of diabetic injury and a diabetic mouse model. This proposal builds on the principal investigator’s previous research experience and clinical training. Currently, Dr. Parker Wilson is spending 25% of his time on the renal and molecular pathology clinical services with the remaining 75% allocated to basic research in Dr. Benjamin Humphreys’ laboratory. Dr. Wilson has an established mentoring relationship with Dr. Humphreys and has published his analysis of snRNAseq data from human diabetic kidney as a first author in PNAS. In addition, Dr. Wilson has a recently-accepted co-first author manuscript describing multimodal snRNAseq and snATACseq integration in the healthy adult kidney in Nature Communications. These findings provide the foundation for this application, which will focus on expanding Dr. Wilson’s scientific skills in single cell methods, bioinformatics analysis and models of diabetic kidney injury. The career development goals will be achieved through mentoring by Dr. Humphreys and an advisory committee with expertise in regulation of chromatin conformation, bioinformatics and diabetic nephropathy. Dr. Wilson will undertake didactic coursework in research ethics, scientific communication and grant writing, molecular biology and advanced computer programming to further his existing knowledgebase. The work will take place at Washington University, which has a strong history of mentoring successful physician-scientists. Completion of this career development award will build a solid foundation for Dr. Wilson as he pursues independence and R01-level funding.