Cellular Dynamics at the Synovium-Bone interface in RA

 描述（由申请人提供）：本临床和技术联合研究中心（CTRS）申请的重点是应用新型组织形态学和转录组分析，以确定靶组织和充分表征的患者人群中类风湿性关节炎（RA）中淋巴细胞（B细胞和T细胞）与骨稳态之间的相互作用。我们计划开发和优化策略，对疾病子集进行分类，定义治疗反应的生物标志物，并确定新的治疗靶点。迄今为止，RA研究的重点是确定个体免疫细胞群或参与破骨细胞（OC）分化和活化或成骨细胞（OB）功能的信号通路的异常，但很少有研究组对疾病中的多个相关细胞群以及靶组织中淋巴细胞和骨隔室之间的相互作用进行综合评估。尽管先前的方法已经产生了关于免疫发病机制的重要线索，但该领域的主要局限性包括缺乏靶向适当治疗策略的生物标志物，以及目前可用的治疗未能在大多数患者中实现低疾病活动性和/或限制进行性关节损伤。我们 研究小组已经进行了多项观察，将B细胞与关节损伤联系起来，包括B细胞耗竭作为治疗的功效和B细胞聚集体在很少研究的靶组织中的存在，所述靶组织包括滑膜、软骨下骨和骨髓（BM），其中OB和OC发展，与T细胞和OC活化以及最近的OB功能障碍有关。我们小组的额外的精液观察已经确定了RA中的单核细胞功能障碍，并表征了OC分化和活化中的关键下游信号通路。然而，异常B-T细胞相互作用和其他途径（包括单核细胞和细胞因子活化）在介导靶组织关节破坏中的作用和精确机制相对未被探索，并且是鉴定新治疗靶点的成熟领域。迄今为止，由于缺乏相关靶组织的获取途径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，这一评估受到阻碍。我们可以克服这些局限性，因为流变学和骨科之间的独特合作，可以从未经治疗的RA患者中获得前所未有的手术和活检样本，以及罗切斯特大学（UR）肌肉骨骼研究中心（CMSR）内的强大技术，这些技术已经在肌肉骨骼疾病的研究中得到了很好的验证。在该提案中，我们将（1）整合治疗和未治疗RA患者的临床、实验室和超声（US）数据，并建立通过US引导下穿刺活检获取滑膜组织的最佳方法，（2）使用新型组织形态计量工具自动定量表征关节中相关免疫和骨细胞的定位，和（3）通过滑膜中离散分选和捕获的细胞群的RNA测序来确定细胞功能。使用新型组织形态学工具和离散细胞群的RNA测序来表征骨-血管翳交界处关键细胞群的位置并在该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鉴定。

 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.