显微镜下型多血管炎患者血浆抗髓过氧化物酶抗体对髓过氧化物酶氧化活性的抑制作用

Inhibition of oxidative activity of myeloperoxidase by anti-myeloperoxidase antibodies from patients with microscopic polyangiitis

目的:本实验旨在研究用亲和层析提取的显微镜下型多血管炎(MPA)患者抗髓过氧化物酶(MPO)抗体对MPO氧化活性的抑制作用,并进一步研究MPO、铜蓝蛋白和抗MPO抗体之间的关系。方法:应用蛋白G亲和层析纯化11名抗MPO抗体阳性的显微镜下型多血管炎患者血浆及12名健康志愿者的血清中免疫球蛋白G(IgG)成分。进一步应用MPO亲和层析柱从患者IgG成分中提取抗MPO抗体。应用经典的MPO氧化活性测定方法分别测定在不同浓度的患者抗MPO抗体、不同浓度的铜蓝蛋白和正常人IgG存在条件下MPO酶的氧化活性,并应用酶联免疫吸附法测定铜蓝蛋白、MPO和抗MPO抗体之间的相互作用。结果:11名MPA患者中有7名患者的抗MPO抗体能够以非竞争性抑制的方式剂量依赖性地抑制MPO的活性。铜蓝蛋白能够以竞争性抑制的方式剂量依赖性和时间依赖性的抑制MPO的活性。抗MPO抗体能够抑制MPO和铜蓝蛋白之间的结合,最大可达到(75.4±11.6)%。结论:大多数MPA患者的抗MPO抗体能够抑制MPO的氧化活性;抗MPO抗体可以干扰MPO和铜蓝蛋白的结合。

Objective： To investigate the inhibitory effects on myeloperoxidase （MPO） oxidation activity by affinity-purified anti-MPO antibodies from patients with microscopic polyangiitis （MPA） and to further investigate the interaction between MPO, ceruloplasmin and anti-MPO antibodies. Methods： Human IgG fractions were purified from plasma of 11 patients with anti-MPO antibody positive MPA and sera of 12 normal controls. Anti-MPO antibodies were further purified from anti-MPO antibody containing IgG fractions using MPO affinity chromatography. The enzyme activity of MPO was measured, in the presence of anti-MPO antibodies and normal IgG preparations, using a classical MPO oxidation assay. Interaction between ceruloplasmin, MPO and anti-MPO antibodies was further investigated using ELISA. Results： Anti-MPO antibodies from 7/11 patients with MPA could inhibit the MPO activity as non-competitive inhibitors in a dose-dependent manner. Ceruloplasmin could competitively inhibit the oxidation activity of MPO in a dose-dependent and time-dependent manner. Anti-MPO antibodies could inhibit the binding between MPO and ceruloplasmin to a maximum of （75. 4 ± 11. 6 ）%. Conclusion： Anti-MPO antibodies, from the majority of patients with MPA, could inhibit the oxidation activity of MPO and interfere with the binding between MPO and ceruloplasmin.