The Role of Beta-Arrestins 1 and 2 in Rheumatoid Arthritis

描述（由申请人提供）：促炎细胞因子和趋化因子在类风湿关节炎（RA）中起关键作用。最近的研究已经解决了toll样受体（TLR）s在RA的发生和进展中的作用。各种tlr在关节炎动物模型中的重要作用已被证实。与健康供者相比，TLR4和内源性TLR4配体如透明质酸和热休克蛋白（HSP）22在RA患者滑膜组织中高表达，进一步暗示TLR4与RA发病机制有关。然而，调控内源性TLR4配体诱导的RA中TLR激活的信号通路尚不完全清楚。最近的研究表明，与TRAF6、I:B1和NF:B1p105相关的衔接蛋白2-阻滞蛋白1和2负性调节TLR信号。我们的数据首次证明，与野生型（WT）小鼠相比，2-阻滞蛋白2敲除（KO）小鼠在胶原抗体诱导的关节炎（CAIA）模型中表现出更严重的关节炎。此外，我们观察到，与对照小鼠相比，胶原诱导关节炎（CIA）小鼠的脾细胞和成纤维细胞样滑膜细胞（FLS）中的2-阻滞蛋白1和2的表达增加。研究还表明，与WT小鼠相比，LPS-和内源性TLR4配体低分子量透明质酸(lw - ha)-诱导的2-阻滞素2 KO小鼠脾细胞中TNF1、IL-6和IL-10的产生增加。这些发现使我们提出了一种假设，即上调2-阻滞蛋白1和2的表达，通过负向调节炎症细胞反应来抑制胶原诱导关节炎的炎症。具体目的是研究2-抑制蛋白1和2在胶原诱导关节炎炎症反应中的负调节作用。研究小鼠CIA模型中2-阻滞蛋白1和2在脾巨噬细胞、CD4+和CD8+ T淋巴细胞、B淋巴细胞、树突状细胞（DC）、多形核白细胞（PMN）和FLS中的表达与疾病严重程度的相关性。一旦鉴定出2-arrestins表达改变的特定免疫细胞，将使用慢病毒表达系统过度表达2-arrestins或用RNAi敲除2-arrestins来检测2-arrestins 1和2表达在这些细胞活化中的作用。了解调节促炎和抗炎基因表达的2-抑制因子依赖信号通路将为RA的发病机制提供新的见解，从而可以发展出创新的靶向干预措施。公共卫生相关性：本项目研究2-抑制蛋白1和2在胶原诱导关节炎炎症反应中的负调节作用。了解2-阻滞蛋白1和2表达与胶原诱导的小鼠关节炎模型中疾病严重程度及其细胞特异性的相关性，以及2-阻滞蛋白1和2表达在内源性toll样受体配体和其他刺激诱导的炎症反应中的作用，将为类风湿关节炎的发病机制提供新的见解，从而为创新的靶向干预提供可能。

DESCRIPTION (provided by applicant): Pro-inflammatory cytokines and chemokines play critical roles in rheumatoid arthritis (RA). Recent studies have addressed the role of toll-like receptors (TLR)s in the development and progression of RA. An important role of various TLRs in the animal model of arthritis has been demonstrated. TLR4 and endogenous TLR4 ligands such as hyaluronan and heat shock protein (HSP)22 are highly expressed in synovial tissue from RA patients compared with healthy donors further implicating TLRs in RA pathogenesis. However the signaling pathways that regulate endogenous TLR4 ligands-induced TLR activation in RA are not fully understood. Recent studies demonstrated that adaptor proteins 2-arrestin 1 and 2 associating with TRAF6, I:B1, and NF:B1p105 negatively regulate TLR signaling. Our data demonstrated for the first time that 2-arrestin 2 knockout (KO) mice exhibited more severe arthritis in the collagen antibody-induced arthritis (CAIA) model compared to wild type (WT) mice. Furthermore, we observed that 2-arrestin 1 and 2 expression are increased in splenocytes and fibroblast-like synoviocytes (FLS) in the collagen-induced arthritis (CIA) mice compared to the control mice. It was also demonstrated that LPS- and the endogenous TLR4 ligand low molecular weight hyaluronan (LMW-HA)- induced TNF1, IL-6 and IL-10 production were augmented in splenocytes from 2- arrestin 2 KO mice compared to WT mice. These findings led us to propose the hypothesis that up-regulation of 2-arrestin 1 and 2 expression suppresses inflammation in collagen-induced arthritis by negative regulation of inflammatory cell responses. The specific aim is to investigate the role of 2-arrestin 1 and 2 as negative regulators of the inflammatory response in collagen-induced arthritis. 2-arrestin 1 and 2 expression in splenic macrophages, CD4+ and CD8+ T lymphocytes, B lymphocytes, dendritic cells (DC)s, polymorphonuclear leukocyte (PMN)s and FLS correlation to the disease severity in the mouse CIA model will be examined. Once the specific immune cells that exhibit altered expression of 2-arrestins are identified, the role of 2-arrestin 1 and 2 expression on activation of these cells will be examined using the lentiviral expression system to overexpress 2-arrestins or knock down 2-arrestins with RNAi. Understanding 2-arrestins-dependent signaling pathways that regulate pro- and anti-inflammatory gene expression will provide novel insights into the pathogenesis of RA from which innovative targeted interventions can evolve. PUBLIC HEALTH RELEVANCE: This project investigates the role of 2-arrestin 1 and 2 as negative regulators of inflammatory response in collagen-induced arthritis. Understanding 2-arrestin 1 and 2 expression in correlation to the disease severity and its cellular specificity in the collagen-induced arthritis model in mice and role of 2-arrestin 1 and 2 expression on endogenous Toll-like receptor ligands and other stimuli induced inflammatory response will provide novel insights into the pathogenesis of rheumatoid arthritis from which innovative targeted interventions can evolve.