Release of Active Peptidyl Arginine Deiminases by Neutrophils Can Explain Production of Extracellular Citrullinated Autoantigens in Rheumatoid Arthritis Synovial Fluid.
Release of Active Peptidyl Arginine Deiminases by Neutrophils Can Explain Production of Extracellular Citrullinated Autoantigens in Rheumatoid Arthritis Synovial Fluid.
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DOI：
10.1002/art.39313
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发表时间：
2015-12
期刊：
影响因子：
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通讯作者：
Scheel-Toellner D
中科院分区：
文献类型：
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作者：
Spengler J;Lugonja B;Ytterberg AJ;Zubarev RA;Creese AJ;Pearson MJ;Grant MM;Milward M;Lundberg K;Buckley CD;Filer A;Raza K;Cooper PR;Chapple IL;Scheel-Toellner D
在大多数类风湿性关节炎（RA）患者中，抗体特异性识别肽基精氨酸脱亚胺酶（PAD）产生的瓜氨酸化自身抗原。中性粒细胞表达高水平的PAD，并在疾病发作期间在RA患者的滑液（SF）中积聚。进行这项研究是为了验证这样的假设：由NETosis（基因组DNA-蛋白质复合物的挤出，称为中性粒细胞细胞外陷阱[NET]）或坏死诱导的中性粒细胞死亡可能有助于炎症关节中自身抗原的产生。在RA患者、骨关节炎（OA）患者和银屑病关节炎（PsA）患者的SF中定量细胞外DNA。通过Western印迹、质谱、免疫荧光染色和PAD活性测定研究PAD从中性粒细胞的释放。在RA和OA患者的SF中评估PAD 2和PAD 4蛋白表达以及PAD酶活性。RA SF中细胞外DNA的表达水平显著高于OA SF（P < 0.001）和PsA SF（P < 0.05），且其表达水平与RA SF中中性粒细胞浓度和PAD活性相关。在体外实验中，坏死的中性粒细胞释放的可溶性细胞外DNA少于NETotic细胞（P < 0.05）。RA SF中PAD活性高于OA SF（P < 0.05）。瓜氨酸化蛋白PAD 2和PAD 4被发现附着在NET上，并在上清液中自由扩散。在中性粒细胞发生NETosis或坏死的上清液中检测到PAD酶活性。中性粒细胞死亡释放活性PAD亚型进入SF是RA细胞外自身抗原产生的合理解释。
In the majority of patients with rheumatoid arthritis (RA), antibodies specifically recognize citrullinated autoantigens that are generated by peptidylarginine deiminases (PADs). Neutrophils express high levels of PAD and accumulate in the synovial fluid (SF) of RA patients during disease flares. This study was undertaken to test the hypothesis that neutrophil cell death, induced by either NETosis (extrusion of genomic DNA–protein complexes known as neutrophil extracellular traps [NETs]) or necrosis, can contribute to production of autoantigens in the inflamed joint. Extracellular DNA was quantified in the SF of patients with RA, patients with osteoarthritis (OA), and patients with psoriatic arthritis (PsA). Release of PAD from neutrophils was investigated by Western blotting, mass spectrometry, immunofluorescence staining, and PAD activity assays. PAD2 and PAD4 protein expression, as well as PAD enzymatic activity, were assessed in the SF of patients with RA and those with OA. Extracellular DNA was detected at significantly higher levels in RA SF than in OA SF (P < 0.001) or PsA SF (P < 0.05), and its expression levels correlated with neutrophil concentrations and PAD activity in RA SF. Necrotic neutrophils released less soluble extracellular DNA compared to NETotic cells in vitro (P < 0.05). Higher PAD activity was detected in RA SF than in OA SF (P < 0.05). The citrullinated proteins PAD2 and PAD4 were found attached to NETs and also freely diffused in the supernatant. PAD enzymatic activity was detected in supernatants of neutrophils undergoing either NETosis or necrosis. Release of active PAD isoforms into the SF by neutrophil cell death is a plausible explanation for the generation of extracellular autoantigens in RA.