【摘要】 目的　研究被自由基氧化修饰的 β2糖蛋白 1(β2 GP1)在自身免疫疾病中自身抗体产生中的作用。方法　采用高氯酸沉淀 ,两步肝素连接的交联琼脂糖凝胶层析结合葡聚糖凝胶 G- 10 0分离纯化大鼠 β2 GP1;应用自由基氧化修饰大鼠 β2 GP1;氧化修饰的 β2 GP1(oxβ2 GP1)、β2 GP1分别加与不加心磷脂 (CL)及 CL 免疫大鼠 ,EL ISA方法检测免疫大鼠血清中抗 β2 GP1抗体和抗心磷脂抗体 (ACA)。结果　两步亲和层析结合凝胶过滤层析有效地分离纯化了大鼠 β2 GP1,蛋白纯度达 90 .78% ;通过羰基含量测定证实了自由基对 β2 GP1的氧化修饰 ;oxβ2 GP1加 CL、β2 GP1加 CL、oxβ2 GP1免疫组与空白对照组、实验对照组比较 ACA水平有显著升高 (P<0 .0 0 1) ,oxβ2 GP1加 CL 免疫组与 β2 GP1加 CL 免疫组比较 ACA水平差异有显著性 (P<0 .0 5 ) ;oxβ2 GP1免疫组与oxβ2 GP1加 CL 免疫组比较 ACA水平差异不显著 (P>0 .0 5 ) ;oxβ2 GP1加 CL、oxβ2 GP1免疫组与对照组比较 ,抗oxβ2 GP1抗体水平有显著升高 (P<0 .0 5 )。结论　两步肝素连接的交联琼脂糖凝胶层析结合葡聚糖凝胶 G- 10 0能有效地分离纯化 β2 GP1;oxβ2 GP1诱导了 ACA和抗 oxβ2 GP1抗体的产生 ;β2 GP1可能是自身免疫疾病发生过程重要的辅助因子或靶抗原更多还原

【Abstract】 OBJECTIVE To explore the role of oxidative modified beta2 glycoprotein1 (β2GP1) by free radicals in auto antibodies production associated with autoimmune disease (AID). METHODS The rat β2GP1 was purified by sequential perchloric acid precipitation, heparin SepharoseCL 6B affinity chromatography and Sephadex G100 gel filter chromatography. β2GP1 was oxidized and modified with free radicals. Using β2GP1, oxidative modified β2GP1 (oxβ2GP1) plus cardiolipin (CL) and oxβ2GP1, β2GP1, CL, respectively, to immunize the rats and to detect anti β2GP1 antibodies and anti cardiolipin antibodies (ACA) levels by ELISA. RESULTS The rat β2GP1 was isolated and purified effectively, the purity was 90.78%. The oxβ2GP1 was confirmed by significant increase in carbonyl group formation. ACA levels were significantly higher in rat serum immunized by oxβ2GP1, oxβ2GP1 plus CL compared with that of control groups (P<0.001), immunized by oxβ2GP1 plus CL compared with β2GP1 plus CL (P<0.05), but immunized by oxβ2GP1 in comparison with oxβ2GP1 plus CL, ACA levels were not significanly distinguished. Anti oxβ2GP1 antibody levels were significantly higher in rat serum immunized by oxβ2GP1 plus CL or oxβ2GP1 compared with control groups (P<0.05). CONCLUSIONS The rat β2GP1 effectively purified by three steps affinity chromatography. The oxβ2GP1 induced to generate both ACA and anti oxβ2GP1 antibodies. β2GP1 is possible an important cofactor leading to AID and the most common antigenic target of autoimmune antibodies. The free radical damages to β2GP1 are possible important pathological process in the pathogenesis and mechanisms responsible for AID.更多还原