异位表达的 RASGRP2（CalDAG-GEFI）在类风湿滑膜中导致破坏性关节炎的发生。

Ectopic RASGRP2 (CalDAG-GEFI) expression in rheumatoid synovium contributes to the development of destructive arthritis.

机构信息

Department of Rheumatology, Endocrinology and Nephrology, Faculty of Medicine and Graduate School of Medicine, Hokkaido University, Sapporo, Hokkaido, Japan.

Department of Orthopedic Surgery, Faculty of Medicine, Hokkaido University, Sapporo, Hokkaido, Japan.

出版信息

Ann Rheum Dis. 2018 Dec;77(12):1765-1772. doi: 10.1136/annrheumdis-2018-213588. Epub 2018 Aug 3.

DOI:10.1136/annrheumdis-2018-213588

PMID:30076153

Abstract

OBJECTIVES

Rheumatoid arthritis (RA) is an autoimmune polyarthritis, in which fibroblast-like synoviocytes (FLS) play a key role in cartilage and bone destruction through tumour-like proliferation and invasiveness. Considering still unsatisfactory remission rate in RA even under treatment with biological disease-modifying antirheumatic drugs, novel therapeutic strategy for treatment-resistant RA is still awaited. In this study, we analysed the expression and function of Ras guanine nucleotide-releasing proteins (RASGRPs), guanine exchange factors for small GTPase Ras, in FLS as a potential therapeutic target for RA.

METHODS

The expression of mRNA was quantified by a real-time PCR assay in FLS isolated from synovial tissue samples. RASGRP2 protein was also evaluated immunohistochemically. Then, we transiently transfected FLS with RASGRP2 expression vector and assessed their proliferation, adhesion, migration and invasion by cellular functional assays and downstream signalling activation using immunoblot. Finally, the therapeutic effect of RASGRP2 silencing was evaluated in type-II collagen-induced arthritis rats.

RESULTS

RASGRP2 was abundantly expressed in FLS from RA synovium, whereas scarcely found in those from osteoarthritis. Expression of RASGRP2 in RA-FLS was enhanced by transforming growth factor-beta. RASGRP2 activated RAP-1, subsequently affecting nuclear factor kappa-light-chain-enhancer of activated B cells pathway and actin dynamics in FLS. RASGRP2-overexpressed FLS had increased abilities of adhesion, migration and interleukin (IL)-6 production. Silencing of RASGRP2 using the intra-articular injection of -specific siRNAs dampened experimental arthritis in rats by inhibiting pannus formation.

CONCLUSIONS

RASGRP2 was identified to be involved in the pathogenesis of RA by promoting adhesion, migration and IL-6 production from FLS, proposed as a potential novel non-immunosuppressive therapeutic target for RA.

摘要

目的

类风湿关节炎（RA）是一种自身免疫性多关节炎，其中成纤维样滑膜细胞（FLS）通过类似于肿瘤的增殖和侵袭，在软骨和骨破坏中发挥关键作用。考虑到即使在使用生物疾病修正抗风湿药物治疗的情况下，RA 的缓解率仍不理想，因此仍需要针对治疗抵抗性 RA 的新治疗策略。在这项研究中，我们分析了 Ras 鸟嘌呤核苷酸释放蛋白（RASGRP）在 FLS 中的表达和功能，RASGRP 是 Ras 小 GTPase 的鸟嘌呤核苷酸交换因子，作为 RA 的潜在治疗靶点。

方法

通过实时 PCR 检测从滑膜组织样本中分离的 FLS 中 mRNA 的表达。还通过免疫组织化学评估 RASGRP2 蛋白。然后，我们通过细胞功能测定和使用免疫印迹法评估下游信号转导激活，将 RASGRP2 表达载体瞬时转染 FLS，评估其增殖、黏附、迁移和侵袭。最后，在 II 型胶原诱导的关节炎大鼠中评估 RASGRP2 沉默的治疗效果。

结果

RASGRP2 在 RA 滑膜的 FLS 中大量表达，而在骨关节炎的 FLS 中几乎检测不到。转化生长因子-β增强了 RA-FLS 中 RASGRP2 的表达。RASGRP2 激活 RAP-1，随后影响 FLS 中的核因子κB 轻链增强子的激活 B 细胞途径和肌动蛋白动力学。RASGRP2 过表达的 FLS 具有增强的黏附、迁移和白细胞介素（IL）-6 产生能力。使用关节内注射 - 特异性 siRNAs 沉默 RASGRP2 可通过抑制血管翳形成来抑制大鼠实验性关节炎。