Centre for Immune Regulation, Institute of Immunology, University of Oslo, Oslo University Hospital-Rikshospitalet, 0027 Oslo, Norway.
Rheumatol Int. 2012 May;32(5):1271-6. doi: 10.1007/s00296-010-1765-y. Epub 2011 Jan 26.
Isoform 4 of the human peptidylarginine deiminase (hPAD4) enzyme may be responsible for the citrullination of antigens in rheumatoid arthritis (RA) and has been shown to be itself the target of disease-specific autoantibodies. Here, we have tested whether the level of serum anti-hPAD4 antibodies in RA patients is stable over a period of 10 years and whether the antibodies influence hPAD4-mediated deimination of the small substrate N-α-benzoyl-L-arginine ethyl ester. RA sera (n = 128) obtained at baseline and after 10 years were assessed for anti-hPAD4 antibodies by a specific immunoassay. For 118 RA patients, serum anti-hPAD4 IgG levels were stable over 10 years. Seven patients who were negative for anti-PAD4 IgG at baseline had become positive after 10 years. Further, total IgG from selected RA patients and controls were purified, and a fraction was depleted for anti-hPAD4 antibodies. Kinetic deimination assays were performed with total IgG and depleted fractions. The k ( cat ) and K ( m ) values of hPAD4-mediated deimination of N-α-benzoyl-L-arginine ethyl ester were not affected by the depletion of the anti-hPAD4 antibodies from the total IgG pool. In conclusion, RA patients remain positive for anti-hPAD4 antibodies over time and some patients who are initially anti-hPAD4 negative become positive later in the disease course. The anti-hPAD4 antibodies did not affect the enzymatic activity of hPAD4 when the small substrate N-α-benzoyl-L-arginine ethyl ester was used. However, this finding may not exclude an effect of these autoantibodies on citrullination of protein substrates in RA.
人肽基精氨酸脱亚氨酶（hPAD4）同工酶 4 可能负责类风湿关节炎（RA）中抗原的瓜氨酸化，并且已经表明其本身是疾病特异性自身抗体的靶标。在这里，我们测试了 RA 患者的血清抗 hPAD4 抗体在 10 年内是否稳定，以及这些抗体是否影响 hPAD4 介导的小分子底物 N-α-苯甲酰-L-精氨酸乙酯的脱亚氨作用。通过特定的免疫测定法，评估了基线时和 10 年后获得的 128 例 RA 血清中的抗 hPAD4 抗体。对于 118 例 RA 患者，血清抗 hPAD4 IgG 水平在 10 年内保持稳定。基线时抗 hPAD4 IgG 阴性的 7 例患者在 10 年后转为阳性。此外，从选定的 RA 患者和对照者的血清中纯化总 IgG，并从总 IgG 池中耗尽抗 hPAD4 抗体的一个分数。使用总 IgG 和耗尽的分数进行动力学脱亚氨作用测定。hPAD4 介导的 N-α-苯甲酰-L-精氨酸乙酯脱亚氨作用的 k（cat）和 K（m）值不受从总 IgG 池耗尽抗 hPAD4 抗体的影响。总之，RA 患者随着时间的推移仍保持抗 hPAD4 抗体阳性，一些最初抗 hPAD4 阴性的患者在疾病过程中后来变为阳性。当使用小分子底物 N-α-苯甲酰-L-精氨酸乙酯时，抗 hPAD4 抗体并不影响 hPAD4 的酶活性。然而，这一发现并不能排除这些自身抗体对 RA 中蛋白质底物瓜氨酸化的影响。
