Establishing a Single-Cell Proteomic Atlas for Normal and Osteoarthritic Articular Cartilage

摘要 尽管已经提出了多种途径和靶点来治疗骨性关节炎，但临床上药物失败率 审判次数之多令人震惊。成功有限的原因包括发现疾病的时间较晚 以及对患者之间的分子异质性缺乏了解。在这项建议中，我们的目标是 充分利用最新开发的单细胞蛋白质组技术，即质量细胞术(CyTOF)，它可以 在单个细胞中同时检测40-80个蛋白质，目的是鉴定不同的细胞 骨性关节炎软骨中的亚群。尽管软骨是一个相对简单的组织，只有一种单细胞类型 包裹在其分泌的细胞外基质(ECM)中的不同程度的退变与 每个骨性关节炎患者都建议，在单细胞水平上了解这个组织(和其他关节组织)可以提供 对骨性关节炎病理和患者异质性的新见解。这将是对单细胞转录的补充 数据，还有一个优势，即蛋白质组快照还可以识别活跃的信号通路 已识别的亚群。单细胞蛋白质组学方法特别适用于强有力的鉴定 很难从RNA测序数据中辨别的稀有细胞群。在这项提案中，我们将建立 用精制的稀土金属板标记的一大批骨关节炎软骨样本的单细胞图谱 Aim1中的抗体用于识别骨关节炎软骨中不同的亚群。在目标2中，我们将测试是否调制 两个新发现的稀有亚群在创伤后骨性关节炎小鼠模型中具有治疗作用 跟踪他们的动态和疾病的进展。在目标3中，我们将分析药物治疗如何影响软骨。 亚群及其在不同患者中的串扰特别要区分是均匀的还是异质的 患者队列中的反应。总的来说，拟议的研究将在识别小说方面产生影响 骨关节炎的再生和病理细胞群，并测试它们的调节治疗潜力。

Abstract Although multiple pathways and targets have been proposed for OA treatment, the rate of drug failure in clinical trials has been astoundingly high. The reasons for the limited success include the late detection of the disease and a lack of understanding of the molecular heterogeneity between patients. In this proposal, we aim to capitalize on the newly developed single-cell proteomic technique, mass cytometry (CyTOF) that allows detection of 40-80 proteins simultaneously in single cells, with the aim of identifying the diverse cellular subpopulations in OA cartilage. Although cartilage is a relatively simple tissue, with a single cell type being encapsulated in its secreted extracellular matrix (ECM), the variable degree of degeneration associated with each OA patient suggests that understanding this tissue (and other joint tissues) at a single cell level can provide novel insights into both OA pathology and patient heterogeneity. This will compliment single-cell transcriptomic data, with the additional advantage that the proteomic snapshot can also identify active signaling pathways in the identified subpopulations. The single-cell proteomic approach is especially pertinent in robustly identifying rare cell populations that are difficult to discern from RNA-sequencing data. In this proposal, we will establish single cell profiles of a large cohort of OA cartilage samples using a refined panel of rare earth metal labeled antibodies in Aim1 to identify distinct subpopulations in OA cartilage. In aim 2, we will test if the modulation of two newly identified rare subpopulations would be therapeutic in a mouse model of post-traumatic OA as well as follow their dynamics with disease progression. In Aim 3, we will analyze how drug treatments affect the cartilage subpopulations and their crosstalk in different patients especially to discern between a uniform or heterogenous response among the patient cohort. Collectively, the proposed studies will be impactful in identifying novel regenerative and pathological cell populations in OA and testing the therapeutic potential of their modulation.