基于细胞计数试剂盒8的药物淋巴细胞刺激试验在急性药物性肝损伤诊断中的应用  

Application of drug lymphocyte stimulation test using cell counting kit-8 assay in diagnosis of acute drug-induced liver injury

作　　者：范作鹏[1] 梁珊[1] 聂巍[1] 仇丽霞[1] 陈杰[1] 张晶[1] 金荣华[1] 胡中杰[1] 

机构地区：[1]首都医科大学附属北京佑安医院丙肝与中毒性肝病科,北京100069

出　　处：《临床肝胆病杂志》2016年第8期1562-1565,共4页Journal of Clinical Hepatology

基　　金：首都临床特色应用研究专项(Z121107001012086);中国肝炎防治基金会天晴肝病研究基金(TQGB20140052);中国初级保健基金会佑安肝病艾滋病基金(YNKT20160016)

摘　　要：目的建立一种基于细胞计数试剂盒8（CCK-8）法的药物淋巴细胞刺激试验（DLST）并应用于急性药物性肝损伤（DILI）的诊断。方法筛选2011年1月-2014年12月北京佑安医院收治的急性DILI患者,收集导致肝损伤的可疑药物,分离外周血单个核细胞（PBMC）,经体外培养后,与可疑药物共孵育,采用CCK-8法检测淋巴细胞的增殖程度。每次实验设阳性对照组,以植物血凝素进行刺激;同时以健康人的PBMC暴露于相同药物,同步检测作为阴性对照。将DLST结果引入RUCAM评分系统,评估对急性DILI诊断敏感性的影响。计数资料组间比较采用χ2检验。结果预实验确定了CCK-8法DLST的实验条件。共纳入61例急性DILI患者,其中52例75种药物完成了基于CCK-8法的DLST检测,48例70种药物结果可靠,共19例22种药物DLST阳性,患者和药物的阳性率分别为39.6%和31.4%,特异性为93.8%,38.2%（13/34）的中药和25.0%（9/36）的西药的DLST结果为阳性,刺激指数中位数为2.29（1.81~14.20）。将DLST结果引入RUCAM评分系统,可将评估结果为＂极可能＂和＂很可能＂的药物由74.7%提高至92.0%（χ2=8.112,P=0.004）。结论利用CCK-8法可进行DLST,将DLST结果引入RUCAM评分系统可显著提高对DILI诊断的敏感性。与经典DLST相比,这种基于CCK-8法的DLST实验室要求低,操作简便、快捷,且花费更低,可在临床进一步使用验证。Objective To develop a novel drug lymphocyte stimulation test（ DLST） using cell counting kit- 8（ CCK- 8） assay,and to investigate its application in the diagnosis of acute drug- induced liver injury（ DILI）. Methods The patients with acute DILI who were admitted to Beijing You An Hospital from January 2011 to December 2014 were screened,and the suspected drugs for liver injury were collected. The patients＇ peripheral blood mononuclear cells（ PBMCs） were isolated,cultured in vitro,and then incubated with the suspected drugs. The CCK- 8 assay was used to measure the proliferation of lymphocytes. A positive control group was established for each experiment and was stimulated by phytohemagglutinin. Meanwhile,PBMCs from healthy subjects were enrolled as the negative control group,and were exposed to the same drugs and measured simultaneously. The results of DLST were introduced into the Roussel Uclaf Causality Assessment Method（ RUCAM） scoring system to assess their effects on the sensitivity of the diagnosis of acute DILI. The chi- square test was used for comparison of categorical data between groups. Results Preliminary experiments were performed to determine the experimental conditions of DLST based on CCK- 8 assay. A total of 61 patients with acute DILI were enrolled. DLST based on CCK- 8 assay was completed in 52 patients who received a total of 75 drugs,and the results were identified to be reliable in 48 patients with 70 drugs. A total of 19 patients with22 drugs achieved positive results of DLST,with positive rates of patients and drugs being 39. 6% and 31. 4%,respectively,and a specificity of 93. 8%. Thirteen（ 38. 2%） out of 34 traditional Chinese medicines and 9（ 25%） out of 36 Western medicines achieved positive results of DLST,and the median stimulation index was 2. 29（ 1. 81- 14. 20）. After the results of DLST were introduced into the RUCAM scoring system,the proportion of drugs assessed as＂highly probable＂or ＂probable＂were increased from 74. 7% to 92. 0%（ χ2=

关 键 词：药物性肝损伤 淋巴细胞活化 细胞计数 诊断 

分 类 号：R575[医药卫生—消化系统]