A＆R：瓜氨酸化Hsp90α/β可能是RA相关肺间质病变的自身抗体标记物

部分类风湿关节炎（RA）患者可出现包括间质性肺炎（ILD）在内的关节外症状。由于目前尚缺乏评估RA患者合并发生ILD风险的方法，所以发现一种独特的RA-相关ILD（RA-ILD）血清标记物显得尤为重要。针对这一问题，来自美国迈阿密米勒医学院的Lisa Harlow等人进行了一项研究。研究结果发表于2013年4月的《关节炎与风湿病》（ARTHRITIS & RHEUMATISM）杂志上。研究发现，瓜氨酸化Hsp90α/β可能成为一种有效的RA-ILD的生物标记物。
研究收集了合并或不合并ILD的RA患者的血清，使用K562细胞（源于1个慢性髓性白血病患者的细胞系）提取物对瓜氨酸及非瓜氨酸化的蛋白进行免疫沉淀。通过不同免疫沉淀可鉴定出RA-ILD患者血清中的瓜氨酸化蛋白，并对此蛋白进行质谱分析。将这些候选蛋白作为抗原底物，并从RA、RA-ILD、混合性结缔组织病（MCTD）或特发性肺间质纤维化（IPF）患者中获取高效筛选的血清，对两者的反应进行ELISA分析。
免疫沉淀的差异及此后的质谱分析结果显示瓜氨酸化的Hsp90α和瓜氨酸化的Hsp90β是RA-ILD患者的候选自身抗原。ELISA分析中的抗原底物包括非瓜氨酸化及瓜氨酸化的Hsp90亚型，研究结果显示瓜氨酸化的Hsp90优先识别RA-ILD患者的血清，与单纯RA（未合并ILD）、MCTD及IPF患者相比，其具有中度敏感性（20-30%）及高度特异性（＞95%）。
本研究表明，基于免疫沉淀差异所识别的瓜氨酸化蛋白提取物可作为一种新自身抗原。应用此技术鉴定出的瓜氨酸化Hsp90α和Hsp90β，是将RA-ILD从RA-未合并ILD、MCTD、IPF中鉴别出的一种有效自身抗体靶点。所以，抗瓜氨酸化Hsp90α/β自身抗体可能成为一种有效的RA-ILD的生物标记物。
与RA相关的拓展阅读：

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Identification of citrullinated hsp90 isoforms as novel autoantigens in rheumatoid arthritis-associated interstitial lung disease.
OBJECTIVE
Subsets of patients with rheumatoid arthritis (RA) develop extraarticular complications that include interstitial lung disease (ILD). Because standardized algorithms for identification of RA patients at risk of developing clinically significant ILD are lacking, the purpose of this study was to elucidate unique serologic markers of RA-associated ILD (RA-ILD).
METHODS
Sera from RA patients with ILD and from RA patients without ILD were used to immunoprecipitate citrullinated and uncitrullinated proteins derived from K562 cell extracts. Mass spectrometry was performed to facilitate identification of citrullinated proteins differentially immunoprecipitated by RA-ILD patient sera. These candidate proteins were then used as substrate antigens in custom enzyme-linked immunosorbent assays (ELISAs) for high-throughput screening of sera obtained from cohorts of patients with RA, RA-ILD mixed connective tissue disease (MCTD), or idiopathic pulmonary fibrosis (IPF).
RESULTS
Differential immunoprecipitation and subsequent mass spectrometric sequencing identified citrullinated Hsp90α and citrullinated Hsp90β as candidate autoantigens in patients with RA-ILD. ELISAs incorporating uncitrullinated and citrullinated isoforms of Hsp90 as substrate antigens demonstrated that sera from patients with RA-ILD preferentially recognized citrullinated versions of Hsp90 with moderate sensitivity (range 20-30%) and great specificity (>95%) relative to sera derived from patients with RA alone (without ILD), patients with MCTD, or patients with IPF.
CONCLUSION
These studies demonstrate the utility of a novel autoantigen discovery method based on differential immunoprecipitation of citrullinated protein extracts. Application of these techniques identified citrullinated versions of Hsp90α and Hsp90β as autoantibody targets distinguishing RA-ILD from RA without ILD, MCTD, and IPF, suggesting that anti-citrullinated Hsp90α/β autoantibodies may serve as effective biomarkers for the potentially devastating pulmonary manifestations of RA-ILD.