替罗非班抑制血小板与中性粒细胞结合干预抗体介导小鼠输血相关急性肺损伤  

Tirofiban interferes with antibody-mediated transfusion-related acute lung injury by inhibiting platelet-neutrophil binding

作　　者：袁欣 余云红 谯从超 孙盼[1] 苏娜[1] 蒋鹏 林方昭[1] YUAN Xin;YU Yunhong;QIAO Congchao;SUN Pan;SU Na;JIANG Peng;LIN Fangzhao(Institute of Blood Transfusion,Chinese Academy of Medical Science&Peking Union Medical College,Chengdu 610052,China)

机构地区：[1]中国医学科学院北京协和医学院输血研究所,四川成都610052

出　　处：《中国输血杂志》2022年第4期377-382,共6页Chinese Journal of Blood Transfusion

基　　金：国家自然科学基金(31801192);中国医学科学院医学与健康科技创新工程(2021-1-I2M-060)。

摘　　要：目的探讨血小板表面糖蛋白(GP)Ⅱb/Ⅲa受体拮抗剂替罗非班抑制血小板活化与中性粒细胞结合成聚集体,从而干预输血相关急性肺损伤(TRALI)的可行性。方法1)取8~10周龄野生型雄性Balb/c小鼠50只,随机均分为5个组,给每只小鼠腹腔注射脂多糖(LPS)0.1 mg/kg 18 h后,给3个组小鼠尾静脉分别注射4.5 mg/kg抗-MHC-Ⅰ或IgG2a同型对照抗体,其中1组在抗-MHC-Ⅰ注射前30 min注入0.5μg/g替罗非班,分别构建(标记)为TRALI模型组、替罗非班TRALI干预组、同型对照组;不做任何处理的1个组设为正常组;每只小鼠尾静脉仅注射0.5μg/g替罗非班的1组小鼠标记为替罗非班正常干预组。2)在各实验组小鼠抗体注射完成后观察小鼠2 h时,结束各组小鼠生命并取出其肺脏,比较肺干湿比、总蛋白、髓过氧化物酶(MPO)、炎性因子、HE染色定量结果差异,分析肺损伤的程度以及替罗非班的干预效果;流式细胞术检测各组小鼠全血血小板活化水平及免疫荧光(IF)定量血小板与中性粒细胞荧光密度,分析替罗非班对TRALI的作用机制。结果1)HE染色检测:替罗非班TRALI干预组与TRALI模型组小鼠肺损伤评分分别为0.1733±0.1204 vs 0.6633±0.1419(P<0.01);2)全血中血小板活化水平:替罗非班TRALI干预组TRALI模型组和正常组平均活化率(%)为5.767±3.224 vs 22.87±9.943 vs 5.070±2.234(P<0.01);3)IF检测:替罗非班TRALI干预组与TRALI模型组小鼠血小板中性粒细胞聚集体平均荧光密度为21.89±3.536 vs 32.77±0.9624(P<0.05)。结论血小板GPⅡb/Ⅲa特异性抑制剂替罗非班抑制了小鼠血小板与中性粒细胞结合,从而可能干预TRALI的进一步发展。Objective To explore the feasibility of tirofiban,a platelet surface glycoprotein(GP)Ⅱb/Ⅲa receptor antagonist intervene in transfusion-related acute lung injury(TRALI),by inhibiting platelet activation and by preventing platelet and neutrophil binding to form aggregates.Methods 1)Fifty wild-type male Balb/c mice,aged 8 to 10 weeks,were randomly divided into TRALI,normal,tirofiban TRALI intervention,isotype control and tirofiban normal intervention groups.In the TRALI model,tirofiban TRALI intervention and isotype control groups,each mouse was injected intraperitoneally with lipopolysaccharide(LPS)0.1 mg/kg,and after 18 h with 4.5 mg/kg anti-MHC-I or IgG2 a isotype control antibody,in which 0.5μg/g tirofiban was injected 30 min before anti-MHC-I injection,and was labeled as tirofiban TRALI intervention.The group without any treatment was set as normal group.The tirofiban normal intervention group was injected with only 0.5μg/g tirofiban into the tail vein,30 min before the injection of anti-MHC-I.2)After antibody injection,the mice were observed for 2 h,then executed with their lungs removed,and the extent of lung injury and the intervention effect of tirofiban were analyzed by comparing the differences in lung dry to wet ratio,total protein,myeloperoxidase(MPO),inflammatory factors and quantitative results of HE staining.The platelet activation level in whole blood and immunofluorescence(IF)quantification of platelet and neutrophil fluorescence were detected by flow cytometry to analyze the mechanism of tirofiban on TRALI.Results 1)The indexes of lung injury in the tirofiban TRALI intervention group and TRALI model group for HE staining were 0.6633±0.1419 vs.0.1733±0.1204(P<0.05),respectively;2)Platelet activation levels(%)in whole blood in the TRALI group,normal group and tirofiban TRALI intervention group were 22.87±9.943 vs 5.070±2.234 vs 5.767±3.224(P<0.05),respectively.3)The mean fluorescence density of platelet neutrophil aggregates for IF detection in the tirofiban intervention group and TRALI

关 键 词：输血相关急性肺损伤 聚集体 血小板-中性粒细胞 GPⅡb/Ⅲa受体拮抗剂 替罗非班 

分 类 号：R457.13R563[医药卫生—治疗学] R9732[医药卫生—临床医学]