肌酸激酶MM同工酶在新生儿杜氏肌营养不良症筛查中的应用

Application of creatine kinase isoenzyme MM in screening neonatal Duchenne muscular dystrophy

目的评估肌酸激酶MM同工酶(CK-MM)用于新生儿杜氏肌营养不良症(DMD)筛查的可行性。方法收集男性新生儿滤纸干血片样本7035份,采用时间分辨荧光法检测CK-MM,对筛查阳性的样本进行DMD基因检测。对时间分辨荧光法检测CK-MM的性能(正确度、精密度、线性范围等)进行验证。采用受试者工作特征(ROC)曲线评价CK-MM筛查DMD的效能。结果时间分辨荧光法检测CK-MM的批内CV、批间CV和平均偏移均<10%,线性回归方程为Y=1.0458X+1.9296(r=0.9999,P<0.01)。DMD患儿CK-MM浓度明显高于正常对照组(P<0.0001),是正常对照组的67倍。7035例男性新生儿筛查阳性6例(0.085%),确诊2例,DMD基因检测结果显示均为半合变异,基因型分别为c.1990C>T(p.Gln664*)和c.7657C>T(p.Arg2553*)。ROC曲线分析结果显示,CK-MM诊断DMD的曲线下面积(AUC)为1.0,最佳临界值为762 ng/mL,敏感性为100%,特异性为99.94%。结论以滤纸干血片为样本检测CK-MM能有效筛查出DMD患儿,且检测方法简便,值得临床推广。

Objective To evaluate the feasibility of creatine kinase isoenzyme MM(CK-MM)in neonatal Duchenne muscular dystrophy(DMD)screening.Methods Totally,7035 dried blood spots of male newborns were collected.CK-MM was determined by time-resolved fluorescence method,and the positive results was determined by DMD gene determination further.The performance(accuracy,precision,linear range and so on)of CK-MM determined by time-resolved fluorescence method was verified.The efficiency of CK-MM in screening DMD was evaluated by receiver operating characteristic(ROC)curve.Results The within-run coefficient of variation(CV),the between-run CV and average bias of CK-MM determined by time-resolved fluorescence method were<10%.The linear regression equation was Y=1.0458X+1.9296(r=0.9999,P<0.01).The concentration of CK-MM in children with DMD was higher than that in healthy control group(P<0.0001),which was 67 times higher than that of healthy control group.Among the 7035 male newborns,6(0.085%)cases were positive after screening,and 2 cases were confirmed.DMD gene determination results showed that all of them were hemizygous mutation,and their genotypes were c.1990C>T(p.Gln664*)and c.7657C>T(p.Arg2553*).ROC curve analysis showed that the area under curve(AUC)of CK-MM for the diagnosis of DMD was 1.0,the optimal cut-off value was 762 ng/mL,the sensitivity was 100%,and the specificity was 99.94%.Conclusions Using dried blood spots to determine CK-MM can effectively screen the children with DMD,and this method is simple and worthy of clinical application.