RITP患者血浆中利福平依赖性抗血小板抗体的特性及其作用机制  

Study on the characteristics and mechanisms of rifampicin dependent antiplatelet antibody in plasma of patients with RITP

作　　者：孙平 吴广胜[2] 梁迅[3] 

机构地区：[1]锦州医科大学附属第一医院血液科,辽宁锦州121000 [2]石河子大学医学院附属第一医院血液科,新疆石河子832008 [3]锦州医科大学基础医学院生物化学与分子生物学教研室,辽宁锦州121000

出　　处：《解放军医学杂志》2017年第5期413-419,共7页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金面上项目(81370619);锦州医科大学校长基金-奥鸿博泽科技创新基金(AH2014010)~~

摘　　要：目的研究利福平诱导的免疫性血小板减少症(RITP)患者血浆中的利福平依赖性抗体(Rd-Ab)的特性及其诱导出血的可能机制,并探索治疗RITP药物的效果和作用特征。方法提取RITP患者血浆,通过流式细胞术、单克隆抗体俘获血小板抗原(MAIPA)法及血小板聚集试验检测血清中Rd-Ab的特性。取NOD/SCID小鼠,建立RITP模型,观察抗体对血小板的破坏机制,评价糖皮质激素和静脉注射免疫球蛋白(IVIG)单独或联合使用对保护血小板的疗效。结果体外实验证实RITP患者血清中存在利福平依赖性抗体(Rd-Ab),该抗体与血小板的结合可被抗糖蛋白Ⅱb/Ⅲa单抗AP2完全阻断,证明其与糖蛋白Ⅱb/Ⅲa的结合位点是钙离子依赖性抗原决定簇。血小板聚集试验显示,Rd-Ab可抑制血小板聚集。小鼠体内实验结果显示,Rd-Ab可致小鼠体内人血小板被迅速清除,单用糖皮质激素可阻断Rd-Ab诱导的人血小板清除,延长血小板平均寿命(MPL),该作用在24h后起效,72h疗效达最佳,而IVIG可即刻起效。与RITP小鼠模型组相比(MPL 1.1±0.2h),糖皮质激素单独治疗组和IVIG单独治疗组的MPL分别延长至4.7±0.7h和4.2±1.1h(P<0.05),两药联合治疗组血小板MPL达6.2±1.5h(P<0.05)。结论 Rd-Ab与血小板膜糖蛋白Ⅱb/Ⅲa上的钙离子依赖性抗原决定簇结合,从而抑制血小板聚集。Rd-Ab可导致小鼠血中人血小板的迅速清除,糖皮质激素和IVIG干预均可在一定程度上抑制血小板清除,IVIG起效更快,但两药联合效果更好。Objective Drug-induced immune thrombocytopenia （DITP） is a major adverse drug effect mediated by drugdependent antibodies. It can be actuated by a wide range of medications, including Rifampicin. Methods Rifampicin-dependent antibody （rd-Ab） from a rifampicin-treated tuberculosis patient with thrombocytopenia （RITP） was characterized by in vitro and in vivo assays, which includes flow cytometry, monoclonal antibody immobilization of platelet antigens （MAIPA） assay, platelet aggregation assay and RITP mouse model. Results The rd-Ab could bind glycoprotein （GP） Ⅱ b/Ⅲ a, as shown by the MAIPA assay. The binding of rd-Ab could be blocked by AP2, a monoclonal antibody that binds a calcium dependent site on GP Ⅱ b/Ⅲ a complex and inhibits ADP-induced platelet aggregation. These results suggested that rd-Ab binds the same site or proximal sites on the GP Ⅱ b/Ⅲ a complex as AP2. Furthermore, the rd-Ab inhibited platelet aggregation in vitro. In an established NOD/SCID mouse model of RITG the rd-Ab caused a rapid clearance of human platelets （MPL=1.1 ＋ 0.2h）. The in vivo platelet loss could be partially prevented by treatments with intravenous immunoglobulin （IVIG） or corticosteroids （MPL=4.7 ± 0.7h and 4.2 ± 1.1h, P〈0.05）. Slowing of platelet clearance was observed immediately upon IVIG treatment, while upon corticosteroid treatment at 24h, and reaching its peak at 72h. Combination of IVlG and corticosteroid treatments was more efficacious （MPL=6.2 ± 1.5h, P〈0.05）. Conclusions 1. The rd-Ab binds to calcium-dependent-epitopes on the calcium dependent site of GP Ⅱ b/Ⅲ a complex, which causes platelet damage and inhibits platelet aggregation. 2. Either IVIG or corticosteroid alone can partially block platelet clearance by rd-Ab. Slowing of platelet clearance is observed immediately upon IVIG treatment, or upon corticosteroid treatment several days later, indicating that IVIG may be more suitable for acutely raising the platelet count than corticosteroid treatment

关 键 词：利福平 血小板减少症 糖蛋白Ⅱb/Ⅲa 模型 动物 

分 类 号：R743.3[医药卫生—神经病学与精神病学]