Spatial Elucidation of Human Acute Kidney Injury and Chronic Kidney Disease using Imaging Mass Cytometry

摘要 急性肾损伤(AKI)是一个综合征术语，包括广泛的侮辱和致病因素 导致肾小球滤过率迅速减少的反应。尽管我们已经取得了重要的进展 在动物模型中我们对肾损伤的理解取得了进展，但关注的却少得多 人类不同类型AKI的发病机制和治疗。实现这一目标的最大障碍 据了解，为AKI进行的肾脏活检数量有限，组织数量很少。 取自肾活检。肾脏精准医学项目正在通过以下方式解决这个复杂的问题 招募愿意接受肾脏活检的AKI无私患者，以促进我们的 人类肾脏疾病的知识。这些活组织检查是用多个互补的 技术。我们建议使用成像质量细胞术来帮助提供高度详细的、定量的 肾活检切片中几乎所有细胞的细胞图，包括它们的分化 损伤和修复通路的状态和激活。结合细胞测序、代谢组学和 在KPMP指导下生成的蛋白质组数据，这将大幅增加我们的 对人类AKI和CKD的认识。 成像质量细胞术(IMC)使用高分辨率激光和质量细胞仪相结合来检测 多达42种不同的重金属结合抗体的存在、位置和数量 组织切片。我们已经成功地使用了IMC与22个重金属共轭验证面板 用于识别常驻肾细胞群、浸润性细胞群和细胞活化的抗体 利用FFPE档案人类肾脏组织的损伤状态，并开发了一种机器学习技术 称为肾脏MAPPS，可快速准确地识别、量化和定位其中约92%的细胞 活组织检查。我们现在建议将验证的抗体小组增加到&gt；30抗体，以允许 鉴定95%的细胞，改进细胞损伤和激活状态评估，并优化 IMC和肾脏-MAPPS分析流水线执行细胞位置的2D和3D定量评估， 人AKI和CKD活检组织中的细胞-细胞相互作用和细胞反应(SA1)。我们会 标准化已定义的工作流程协议，并结合关键的严格质量控制评估步骤 积分(SA2)，然后将此IMC工作流程应用于KPMP提供的肾脏样本，并与 KPMP中央枢纽和联盟成员制定准确的协议，以绘制 将scRNAseq/SnRNAseq数据、蛋白质组学数据和代谢组学数据添加到适当的细胞和 使用肾脏-MAPPS(SA3)的地点。

Abstract Acute kidney injury (AKI) is a syndromic term encompassing a wide range of insults and pathogenic responses that lead to a rapid reduction in glomerular filtration. Although we have made significant progress in our understanding of kidney injury in animal models, far less attention has been focused on the pathogenesis and treatment of the diverse types of human AKI. The largest barrier in achieving this knowledge is the limited number of kidney biopsies performed for AKI and the small amount of tissue obtained from renal biopsy. The Kidney Precision Medicine Project is tackling this complex issue by recruiting altruistic patients with AKI who are willing to have kidney biopsies in order to advance our knowledge of human kidney disease. These biopsies are being interrogated using multiple complementary technologies. We propose to use Imaging Mass Cytometry to help provide a highly detailed, quantitative cellular map of nearly all cells present in sections from those kidney biopsies, including their differentiation state and activation of injury and repair pathways. Combined with the cell sequencing, metabolomic, and proteomic data generated under KPMP guidance, this will provide a substantial increase in our understanding of human AKI and CKD. Imaging mass cytometry (IMC) uses a high-resolution laser combined with a mass cytometer to detect the presence, location and amount of up to 42 different heavy metal conjugated antibodies hybridized to a tissue section. We have successfully used IMC with a panel of 22 heavy metal conjugated validated antibodies to identify resident kidney cell populations, infiltrating cell populations, and cell activation and injury states using archival FFPE human kidney tissue, and developed a machine learning technique termed Kidney MAPPS to rapidly and accurately identify, quantify and localize ~92% of all cells in those biopsies. We now propose to increase that validated antibody panel to >30 antibodies that will allow identification of >95% of cells and improve cell injury and activation state assessment, and to optimize the IMC and Kidney-MAPPS analysis pipeline to perform 2D and 3D quantitative assessment of cell location, cell-cell interactions and cellular responses in human AKI and CKD biopsy tissues (SA1). We will standardize a defined work-flow protocol coupled with rigorous quality control assessment steps at key points (SA2), and then apply this IMC work-flow to kidney samples provided by KPMP and integrate with the KPMP Central Hub and consortium members to develop accurate protocols for mapping the scRNAseq/snRNAseq data, proteomics data and metabolomics data onto the appropriate cells and locations using Kidney-MAPPS (SA3).