α1抗胰蛋白酶角蛋白84和微管蛋白β链在类风湿关节炎滑膜组织中的高表达

Increased expression of alpha 1-antitrypsin, keratin type II cuticular Hb4 and tubulin beta chain in the synovial tissues of patients with rheumatoid arthritis

目的研究瓜氨酸化自身抗原在类风湿关节炎（RA）中的表达情况。方法分别对RA患者的滑膜和血清采用二维免疫印迹分析、基质辅助激光解析电离飞行时间质谱、免疫印迹、免疫组织化学和酶联免疫吸附试验（ELISA）法研究特异性瓜氨酸化蛋白的表达变化；通过ELISA法检测RA患者血清中潜在的自身抗原；采用单因素方差分析、LSD检验、Kruskal-Walls检验以及Spearman等级相关进行统计分析。结果研究发现RA患者的滑膜组织和血清存在α1，抗胰蛋白酶（A1AT）、纤维蛋白原β链（FIBB）、角蛋白84（KRT84）、微管蛋白β链（TBB）和波形蛋白（VIME）5种特异性表达蛋白；A1AT、角蛋白84和TBB在RA患者的滑膜和滑膜液中显著高表达，此外，RA患者血清中KRT84高表达。结论在RA中特异表达的自身抗原包括：A1AT、FIBB、KRT84、TBB和波形蛋白，已有研究证明了FIBB和波形蛋白为自身抗原，同时证明了我们研究的可行性和结果的可靠性。

Objective The present study investigated the expression of the citrullinated proteins in the synovium and serum of rheumatoid arthritis （RA） patients. Methods The expression of the citrullinated proteins in the synovium and serum of RA patients was analyzed by two-dimensional western blotting analysis （2-D WB）, mass spectrometry MALD1-TOF/TOF MS, western blotting, immunohistochemistry and ELISA. Then we analyzed the data with one-way ANOVA, LSD test, Krnskal-Walls test and Spearman correlation analysis. Results Alpha-l-antitrypsin （A1AT）, fibrinogen beta chain （FIBB）, keratin type II cuticular Hb4 （KRT84）, tubulin beta chain （TBB） and vimentin （VIME） were detected in RA serum and anti-citrulline antibody could be detected using 2-D WB. A1AT, KRT84 and TBB were expressed significantly in the synovial membranes and synovial fluids of RA patients. Furthermore, high levels of autoantibodies against KRT84 were detected in the blood of RA patients when compared with samples from the healthy controls. Conclusion Current study has identified novel autoantigens in RA, including A1AT, FIBB, KRT84, TBB and VIME using 2-D WB with purified RA sera and anti-citrulline antibody. FIBB and VIME have been confirmed to be autoantigens in the literature, this demonstrates the feasibility of our protocol and the reliability of our study results.