Abstract：
The recombinant human proteoglycan aggrecan-G1 domain (rhG1)-induced arthritis (GIA) mouse model is a complex model of rheumatoid arthritis (RA). In GIA, autoimmune arthritis is induced by repeated intraperitoneal immunization of genetically susceptible BALB/c mice with the rhG1 antigen emulsified in the adjuvant dimethyldioctadecylammonium (DDA). This article describes the steps for producing and purifying the rhG1 antigen, the immunization protocol, methods for following the clinical picture of arthritis, and the evaluation of relevant laboratory parameters. In this model, the autoimmune arthritis develops stepwise, similar to RA: First is the preclinical stage (after the first immunization, days 0-20) with no sign of inflammation but detectable T and B cell activation; next, the stage of early arthritis (after the second immunization, days 21-41), where the first definitive signs of arthritis appear together with autoantibody production; and then the severe late-stage arthritis (after the third immunization, after day 42), which presents with massive inflammation of the limbs, leading to cartilage and bone destruction and finally ankylosis. The protocols described here provide sufficient information for investigators to use the GIA model to study different aspects of autoimmune arthritis. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Induction of recombinant human proteoglycan aggrecan-G1 domain (rhG1)-induced arthritis (GIA) Support Protocol 1: Production of rhG1-Xa-mFc2a fusion protein with CHOK1 mammalian expression system Support Protocol 2: Purification of the rhG1-Xa-mFc2a fusion protein by affinity chromatography Support Protocol 3: Preparation of DDA adjuvant Support Protocol 4: Clinical assessment of arthritis Support Protocol 5: Measurement of serum antibody levels and cytokines Support Protocol 6: Measurement of rhG1-induced proliferation and cytokine production in spleen cell culture Support Protocol 7: Histological assessment of arthritic limbs Support Protocol 8: Evaluation of arthritis with micro-computed tomography.
摘要：
重组人蛋白聚糖聚集素-G1结构域(rhG1)诱导的关节炎(GIA)小鼠模型是类风湿性关节炎(RA)的复杂模型。在GIA,自身免疫性关节炎是通过在佐剂二甲基二十八烷基铵（DDA）中乳化的rhG1抗原对遗传易感的BALB/c小鼠进行反复腹膜内免疫诱导的。本文介绍了rhG1抗原的生产和纯化步骤,免疫方案，方法跟踪关节炎的临床表现，以及相关实验室参数的评估。在这个模型中,自身免疫性关节炎逐步发展，与RA相似：首先是临床前阶段(第一次免疫后，第0-20天)，没有炎症迹象，但可检测到T和B细胞活化；接下来，早期关节炎的阶段（第二次免疫后，第21-41天），其中关节炎的第一个明确迹象与自身抗体产生一起出现；然后是严重的晚期关节炎（第三次免疫后，在第42天之后)，表现为四肢的大量炎症，导致软骨和骨骼破坏，最后导致强直。本文描述的方案为研究人员使用GIA模型研究自身免疫性关节炎的不同方面提供了足够的信息。©2024作者WileyPeriodicalsLLC出版的当前协议。基本方案:诱导重组人蛋白聚糖聚集蛋白-G1结构域(rhG1)-诱导的关节炎(GIA)支持方案1:用CHOK1哺乳动物表达系统产生rhG1-Xa-mFc2a融合蛋白支持方案2:通过亲和层析纯化rhG1-Xa-mFc2a融合蛋白支持方案3:DDA佐剂支持方案4:关节炎的临床评估支持细胞因子的产生方案4:用计算机骨断层摄影术
