PATHOGENESIS OF BONE EROSION IN RHEUMATOID ARTHRITIS

描述（由申请人提供）：类风湿性关节炎（RA）是一种慢性炎症性疾病，通常会导致关节软骨和骨骼的严重破坏。大量证据表明，RA的骨侵蚀是由破骨细胞（OCs）产生的。破骨细胞分化和激活的一个重要因素是NF-KB配体的受体激活剂（RANKL），它通过特定的细胞表面受体NF-KB受体激活剂（RANK）介导其作用。最近，一种新的RANKL分泌形式（secRANKL）被发现，它似乎是由一个独特的启动子调控的。本提案中概述的研究旨在验证RANKL在RA骨侵蚀部位的局部表达增强的假设，以及骨侵蚀部位细胞中RANKL表达的调节是局灶性骨质流失的关键决定因素。特异性目的1将验证来自RA滑膜的细胞产生RANKL在破骨细胞骨侵蚀的发病机制中至关重要的假设。这个目标将解决以下问题：RA骨侵蚀部位RANKL的确切细胞来源是什么？RANKL在哪里表达与OPG表达和RANK阳性OC前体相关？回收的人体组织将首先用于回答这些问题。我们将在两种炎症性关节炎小鼠模型（胶原诱导关节炎（CIA）和KJBxN模型）中确认细胞表达谱，并确定这些因子的时间表达。来自RA滑膜的分散细胞将用于确定RANKL亚型在相关细胞类型中的表达。最后，将在体外共培养破骨细胞形成模型中确定secRANKL亚型在破骨细胞形成中的活性。特异性目的2将验证T细胞衍生的RANKL是RA骨侵蚀所必需的假设。为了明确确定T细胞来源的RANKL在骨侵蚀中的作用，将在T细胞提供RANKL的唯一来源的小鼠和缺乏T细胞的基因工程小鼠品系中产生关节炎。特异性目标3将确定在RA中存在的细胞类型中负责构成和诱导膜结合RANKL异构体表达的调控元件，并将测试NFAT转录因子家族在诱导调节memRANKL基因中至关重要的假设。对局灶性RA骨侵蚀中RANKL表达细胞类型的RT-PCR分析显示，两种已知RANKL亚型的组成型和诱导型表达存在差异。初步数据还证明了nfat在T细胞中调控RANKL的潜在作用。该目的将提供RANKL基因表达在RA骨侵蚀细胞类型中的调控数据，并可能导致新的治疗策略，以防止该疾病的骨破坏。

DESCRIPTION (provided by applicant): Rheumatoid arthritis (RA) is a chronic inflammatory disease that often produces severe destruction of articular cartilage and bone. Considerable evidence indicates that bone erosions in RA are produced by osteoclasts (OCs). An essential factor for the differentiation and activation of osteoclasts is receptor-activator of NF-KB ligand (RANKL), which mediates its effects via a specific cell surface receptor, receptor-activator of NF-kB (RANK). Recently, a novel secreted form of RANKL (secRANKL) has been identified which appears to be regulated by a unique promoter. The studies outlined in this proposal are designed to test the hypothesis that that there is enhanced local expression of RANKL at sites of bone erosion in RA and that regulation of RANKL expression in cells at sites of bone erosion is a critical determinant of focal bone loss. Specific Aim 1 will test the hypothesis that RANKL production by cells derived from RA synovium is critical in the pathogenesis of osteoclastic bone erosion. This Aim will address the following questions: What are the exact cellular sources of RANKL at sites of bone erosion in RA? Where is RANKL expressed in relation to OPG expression and to RANK positive OC precursors? Retrieved human tissues will be utilized initially to answer these questions. Cellular expression profiles will be confirmed and the temporal expression of these factors will be determined in two murine models of inflammatory arthritis: collagen-induced arthritis (CIA), and the KJBxN model. Dispersed cells from RA synovium will be used to determine the expression of RANKL isoforms in relevant cell types. Finally, the activity of the secRANKL isoform in osteoclastogenesis will be determined in an in vitro co-culture model of osteoclastogenesis. Specific Aim 2 will test the hypothesis that T cell-derived RANKL is required for bone erosion in RA. Arthritis will be generated in mice in which T cells provide the only source of RANKL, and in genetically engineered mouse strains lacking T cells, in order to definitively determine the role of T cell-derived RANKL in bone erosion. Specific Aim 3 will identify regulatory elements responsible for the constitutive and inducible expression of the membrane-bound RANKL isoform in cell types present in RA, and will test the hypothesis that the NFAT family of transcription factors is critical in the inducible regulation of the memRANKL gene. RT-PCR analysis of RANKL expressing cell-types present in focal RA bone erosions demonstrates differential constitutive and inducible expression of the two known RANKL isoforms. Preliminary data also demonstrate a potential role of NFATs in RANKL regulation in T cells. This Aim will provide data on the regulation of RANKL gene expression in cell types present in bone erosion in RA and may lead to new therapeutic strategies for preventing bone destruction in this disease.