Rochas Caroline, Hillion Sophie, Saraux Alain, Mageed Rizgar A, Youinou Pierre, Jamin Christophe, Devauchelle Valérie
Université Européenne de Bretagne, Université de Brest, IFR 148 ScInBioS, and Laboratory of Immunology, Centre Hospitalier Universitaire, Brest Hôpital Morvan and Cavale Blanche, Brest, France.
Arthritis Rheum. 2009 May;60(5):1261-71. doi: 10.1002/art.24498.
B cells that accumulate in the synovial tissue of rheumatoid arthritis (RA) patients revise their receptors due to coordinate expression of recombination-activating gene 1 (RAG-1) and RAG-2 genes. The aim of this study was to determine the mechanisms that control this re-expression.
B cells from healthy control subjects were cocultured with fibroblast-like synoviocytes (FLS) from patients with RA and osteoarthritis (OA). Re-expression of RAG messenger RNA (mRNA) and proteins was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and indirect immunofluorescence. Activity of RAG enzymes was evaluated by flow cytometry to measure variations in immunoglobulin kappa and lambda light chain expression and by ligation-mediated-PCR to assess specific DNA breaks. Blocking antibodies, short hairpin RNA, and recombinant cytokine were used to identify the molecules involved in RAG re-expression.
RA FLS, but not OA FLS, induced B cells to re-express RAG mRNA and proteins. Enzymes were functional, since the kappa-to-lambda ratios decreased and specific DNA breaks were detectable after coculture with RA FLS. Transmembrane BAFF provided the first signal of RAG re-expression, since its down-regulation in RA FLS prevented RAG gene transcription in B cells. The failure of transmembrane BAFF from OA FLS to induce RAG suggests that a second signal was provided by RA FLS. Interleukin-6 (IL-6) is a candidate, since blockade of its receptors precluded transcription of RAG genes by RA FLS. Unless supplemented with IL-6, OA FLS were unable to induce RAG gene expression in normal B cells.
Two independent signals are required for the induction of RAG gene expression in B cells that infiltrate the synovium of patients with RA.
类风湿关节炎（RA）患者滑膜组织中积聚的B细胞由于重组激活基因1（RAG-1）和RAG-2基因的协同表达而对其受体进行重排。本研究的目的是确定控制这种重新表达的机制。
将健康对照受试者的B细胞与RA和骨关节炎（OA）患者的成纤维细胞样滑膜细胞（FLS）共培养。通过逆转录-聚合酶链反应（RT-PCR）和间接免疫荧光分析RAG信使核糖核酸（mRNA）和蛋白质的重新表达。通过流式细胞术测量免疫球蛋白κ和λ轻链表达的变化以及通过连接介导的PCR评估特定DNA断裂来评估RAG酶的活性。使用阻断抗体、短发夹RNA和重组细胞因子来鉴定参与RAG重新表达的分子。
RA FLS而非OA FLS诱导B细胞重新表达RAG mRNA和蛋白质。酶具有功能，因为与RA FLS共培养后κ/λ比值降低且可检测到特定的DNA断裂。跨膜BAFF提供了RAG重新表达的第一个信号，因为其在RA FLS中的下调阻止了B细胞中RAG基因的转录。OA FLS的跨膜BAFF未能诱导RAG表明RA FLS提供了第二个信号。白细胞介素-6（IL-6）是一个候选分子，因为阻断其受体可阻止RA FLS转录RAG基因。除非补充IL-6，OA FLS无法在正常B细胞中诱导RAG基因表达。
在浸润RA患者滑膜的B细胞中诱导RAG基因表达需要两个独立的信号。
