类风湿关节炎患者外周血单个核细胞肽酰基精氨酸脱亚氨酶4表达及组蛋白甲基化水平

Peptidyl arginine deiminase 4 participates in the pathogenesis of rheumatoid arthritis by influencing histone methylation

目的通过检测类风湿关节炎（RA）患者外周血单个核细胞（PBMC）肽酰基精氨酸脱亚氨酶4（PAD4）mRNA表达及组蛋白H3R17不对称二甲基化、H4R3对称二甲基化、H4R3单甲基化水平，探讨其可能的参与机制。方法采用实时荧光定量聚合酶链反应检测60例RA患者、40例健康人PBMCPAD4mRNA表达。Westernblot检测26例RA患者、12例骨关节炎（OA）患者、10例健康人H3R17不对称二甲基化、H4R3对称二甲基化、H4R3单甲基化水平。结果RA患者PAD4mRNA表达水平为34．6（16．7，70．8），高于健康人[20．6（11．1，51．8）；P〈0．05]。RA患者组蛋白H3R17不对称二甲基化水平（71．34±25．65）高于OA患者（37．18±18．62）和健康人（50．67±13．99），差异有统计学意义（P〈0．01），并与关节疼痛数、关节肿胀数呈正相关（r=0．418，P=0．034；r=0．402，P=0．042）；RA患者H4R3对称二甲基化水平（75．024-20．35）与OA患者（57．92-i-22．77）和健康人（68．374-17．57）比，差异无统计学意义（P〉0．05）；RA患者H4R3单甲基化水平（11．24±7．81）低于OA患者（32．77±30．77）和健康人（51．20±47．14），差异有统计学意义（P〈0．05）；H4R3单甲基化水平与PAIM呈负相关（r=－0．643，P〈0．01），H3R17不对称二甲基化水平、H4R3对称二甲基化水平与PAD4无相关性（r=－0．185，P：0．377；r=0．198，P=0．344）。结论RA患者组蛋白甲基化水平异常可能是RA发病的原因之一。PAD4可能通过影响组蛋白H4R3甲基化参与RA发病，有可能作为RA治疗的新靶点。

Objective To explore the probable function of peptidyl arginine deiminase 4 （ PAD4 ） in rheumatoid arthritis （RA）. Methods Real-time quantitative polymerase chain reaction （PCR） was used to determine the expression of PAD4 mRNA in peripheral blood mononucleated ceils （PBMCs） from 60 RA patients and 40 healthy individuals. Asymmetric di-methylation of histone H3R17, symmetric di-methylation and mono-methylation of H4R3 were semi-quantified by Western blotting in 12 patients with osteoarthritis （OA） ,26 patients with RA and 10 healthy controls. Results PAD4 mRNA in RA was significantly higher than that in healthy controls[34. 6（ 16. 7±70. 8） vs 20. 6（ 11.1, 51.8） ,P 〈0. 05]. The level of histone H3R17 asymmetric di-methylation in RA was significantly higher than that of OA or control groups（71.34± 25.65 vs 37.18 ＋ 18.62 vs 50. 67 ± 13.99,P 〈0. 01 ）, which was positively related to Tender joint count and Swollen joint count in 28 joints （ r = 0. 418, P = 0. 034 ; r = 0. 402, P = 0. 042）. The level of histone H4R3 symmetric di-methylation was similar in RA, OA and control groups （75.02 ± 20. 35 vs 57. 92 ± 22. 77 vs 68.37 ± 17.57, P 〉 0. 05 ）. The level of histone H4R3 mono-methylation in RA patients was significantly lower than that of OA patients and healthy individuals （ 11.24±7.81 vs 32. 77 ±30. 77 vs 51.20 ±47.14,P 〈 0. 05 ）. The level of histone H4R3 mono-methylation in RA patients was negatively correlated to PAD4 （r= -0. 643,P 〈 0. 01 ）. The level of histone H3R17 asymmetrie di-methylation and H4R3 symmetrie di-methylation was not assoeiated with PAD4 level in RA group （r = -0. 185, P = 0. 377; r =0. 198, P = 0. 344）. Conclusions The level of histone H3R17 asymmetrie di-methylation is signifieantly higher and the level of histone H4R3 mono-methylation is signifieantly lower in RA patients eomparing with OA and control groups. Abnormality of histone methylation may be one of the meehanisms for the development of RA. PAI）4 probably plays an important role in rheumatoid arthritis by influeneing histone methylation.