Cellular Dynamics at the Synovium-Bone interface in RA

 描述(由申请人提供)：这一联合临床和技术研究网站(CTRS)应用的重点是应用新的组织形态计量学和转录组分析，以确定目标组织和具有良好特征的患者群体中淋巴细胞(B细胞和T细胞)和类风湿关节炎(RA)中的骨稳态之间的相互作用。我们计划制定和优化战略，对疾病亚类进行分类，定义治疗反应的生物标记物，并确定新的治疗目标。到目前为止，RA的研究主要集中在确定单个免疫细胞群体的异常或参与破骨细胞(OC)分化和激活或成骨细胞(OB)功能的信号通路，但很少有小组提供了对疾病中多个相关细胞群体以及靶组织中淋巴细胞和骨间隔之间的相互作用的综合评估。虽然以前的方法已经产生了关于免疫发病机制的重要线索，但该领域的主要限制包括缺乏针对适当治疗策略的生物标志物，以及目前可用的治疗方法未能在大多数患者中实现较低的疾病活动性和/或限制进行性关节损害。我们的 研究小组曾多次观察到B细胞与关节损伤之间的关系，包括B细胞去除作为治疗方法的疗效，以及很少被研究的靶组织中B细胞聚集体的存在，这些组织包括滑膜、软骨下骨和骨髓(BM)，在这些组织中OBS和OCS发生，与T细胞和OC激活以及最近的OB功能障碍有关。我们小组的其他开创性观察定义了类风湿性关节炎的单核细胞功能障碍，并确定了OC分化和激活的关键下游信号通路。然而，异常的B-T细胞相互作用和其他途径(包括单核细胞和细胞因子激活)在介导靶组织关节破坏中的作用和确切机制相对未被探索，并且是识别新的治疗靶点的成熟领域。到目前为止，由于无法接触到相关的靶组织，以及需要新的技术方法来更好地定义细胞和细胞因子网络，这一评估受到了阻碍。我们可以克服这些限制，因为风湿学和骨科之间的独特合作，可以获得前所未有的手术和活组织检查样本，来自未经治疗的RA患者，以及罗切斯特大学(UR)肌肉骨骼研究中心(CMSR)内已经得到很好验证的肌肉骨骼疾病研究的强大技术。在这项建议中，我们将(1)整合已治疗和未治疗的RA患者的临床、实验室和超声(US)数据，并建立通过超声引导的针刺活检获取滑膜组织的最佳方法，(2)使用用于自动定量的新型组织形态计量学工具来表征相关免疫细胞和骨细胞在关节中的定位，以及(3)通过对滑膜中离散的、分类的和捕获的细胞群体进行RNA测序来确定细胞功能。使用新的组织形态计量学工具和离散细胞群体的RNA测序，确定关键细胞群体在骨-血管膜连接处的位置和在这个3D环境中定义细胞功能的强大双重方法将更好地反映潜在的疾病发病机制，允许更精确的患者分层和新靶点的识别。

 DESCRIPTION (provided by applicant): The focus of this Combined Clinical and Technology Research Site (CTRS) application is to apply novel histomorphometric and transcriptome analytics to define the interplay between lymphocytes (B cells and T cells) and bone homeostasis in rheumatoid arthritis (RA) within target tissue and well-characterized patient populations. We plan to develop and optimize strategies to categorize subsets of disease, define biomarkers of treatment response, and identify novel treatment targets. Research to date in RA has focused on defining abnormalities in individual immune cell populations or signaling pathways involved in osteoclast (OC) differentiation and activation or osteoblast (OB) function, but very few groups have provided an integrated assessment of the multiple relevant cell populations in the disease and the interactions between the lymphocyte and bone compartments in target tissue. Although prior approaches have yielded important clues regarding immune pathogenesis, major limitations in the field include the lack of biomarkers to target appropriate treatment strategies and the failure of currently available treatments to achieve low disease activity and/or limit progressive joint damage in the majority of patients. Our group has made multiple observations linking B cells to joint damage including the efficacy of B cell depletion as a treatment and the presence of B cell aggregates in rarely studied target tissue including the synovium, subchondral bone, and bone marrow (BM) where OBs and OCs develop, associated with T cell and OC activation and more recently OB dysfunction. Additional seminal observations by our group have defined monocyte dysfunction in RA and characterized key down-stream signaling pathways in OC differentiation and activation. However, the role and precise mechanisms of aberrant B-T cell interactions and other pathways (including monocyte and cytokine activation) in mediating joint destruction in the target tissue are relatively unexplored and a ripe area for identification of new treatment targets. This assessment to date has been hampered by lack of access to relevant target tissue and the need for novel technologic approaches to better define cellular and cytokine networks. We can surmount these limitations given a unique collaboration between Rheumatology and Orthopedics with access to unprecedented surgical and biopsy samples from untreated RA patients and powerful technologies within the Center for Musculoskeletal Research (CMSR) at the University of Rochester (UR) already well-validated for the study of musculoskeletal diseases. In this proposal we will (1) Integrate clinical, laboratory, and ultra-sound (US) data on treated and untreated RA patients and establish optimal methods for acquisition of synovial tissue by US-guided needle biopsy, (2) Characterize the localization of relevant immune and bone cells in the joint using novel histomorphometric tools for automated quantitation, and (3) Define cell function by RNA sequencing of discrete sorted and captured cell populations in synovium. The powerful dual approach of characterizing the location of key cell populations at the bone-pannus junction and defining cell function within this 3D milieu using novel histomorphometric tools and RNA sequencing of discrete cell populations will better reflect underlying disease pathogenesis, allowing more precise patient stratification and identification of novel targets.