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Abstract：
The aim of the present study was to establish a non-invasive, fast and robust enzymatic assay to confirm fatty acid oxidation defects (FAOD) in humans following informative newborn-screening or for selective screening of patients suspected to suffer from FAOD.
The reliability of this method was tested in whole blood from FAOD patients with specific enzymatic defects. Whole blood samples were assayed in 30 medium chain- (MCADD, age 0 to 17 years), 6 very long chain- (VLCADD, age 0 to 4 years), 6 long chain hydroxy- (LCHAD, age 1 to 6 years), 3 short chain- (SCADD, age 10 to 13 years) acyl-CoA-dehydrogenase- and 2 primary carnitine transporter deficiencies (CTD, age 3 to 5 years). Additionally, 26 healthy children (age 0 to 17 years) served as controls. Whole blood samples were incubated with stable end-labeled palmitate; labeled acylcarnitines were analyzed by tandem mass spectrometry and compared with controls and between patient groups (Mann-Whitney Rank Sum Test). Concentrations of specific labeled acylcarnitine metabolites were compared between particular underlying MCADD- (ANOVA), VLCADD- and LCHADD- genetic variants (descriptive data analysis).
11 different acylcarnitines were analyzed. MCADD- (C8-, C10-carnitine, C8/C10- and C8/C4-carnitine), VLCADD- (C12-, C14:1-, C14:2-carnitine, C14:1/C12- and C14:2/C12-carnitine), LCHADD (C16-OH-carnitine) as well as CTD- deficiency (sum of all acylcarnitines) samples could be clearly identified and separated from control values as well as other FAOD, whereas the sum of all acylcarnitines was not conclusive between FAOD samples. Furthermore, C4- (SCADD), C14- (VLCADD) and C14-OH-carnitines (LCHADD) were discriminating between the FAOD groups. Metabolic parameters did not differ significantly between underlying MCADD variants; similar results could be observed for VLCADD- and LCHADD- variants.
This functional method in whole blood samples is relatively simple, non-invasive and little time consuming. It allows to identify MCADD-, VLCADD-, LCHADD- and carnitine transporter deficiencies. The genetic phenotypes of one enzyme defect did not result in differing acylcarnitine patterns in MCADD, VLCADD or LCHADD in vitro.
The reliability of this method was tested in whole blood from FAOD patients with specific enzymatic defects. Whole blood samples were assayed in 30 medium chain- (MCADD, age 0 to 17 years), 6 very long chain- (VLCADD, age 0 to 4 years), 6 long chain hydroxy- (LCHAD, age 1 to 6 years), 3 short chain- (SCADD, age 10 to 13 years) acyl-CoA-dehydrogenase- and 2 primary carnitine transporter deficiencies (CTD, age 3 to 5 years). Additionally, 26 healthy children (age 0 to 17 years) served as controls. Whole blood samples were incubated with stable end-labeled palmitate; labeled acylcarnitines were analyzed by tandem mass spectrometry and compared with controls and between patient groups (Mann-Whitney Rank Sum Test). Concentrations of specific labeled acylcarnitine metabolites were compared between particular underlying MCADD- (ANOVA), VLCADD- and LCHADD- genetic variants (descriptive data analysis).
11 different acylcarnitines were analyzed. MCADD- (C8-, C10-carnitine, C8/C10- and C8/C4-carnitine), VLCADD- (C12-, C14:1-, C14:2-carnitine, C14:1/C12- and C14:2/C12-carnitine), LCHADD (C16-OH-carnitine) as well as CTD- deficiency (sum of all acylcarnitines) samples could be clearly identified and separated from control values as well as other FAOD, whereas the sum of all acylcarnitines was not conclusive between FAOD samples. Furthermore, C4- (SCADD), C14- (VLCADD) and C14-OH-carnitines (LCHADD) were discriminating between the FAOD groups. Metabolic parameters did not differ significantly between underlying MCADD variants; similar results could be observed for VLCADD- and LCHADD- variants.
This functional method in whole blood samples is relatively simple, non-invasive and little time consuming. It allows to identify MCADD-, VLCADD-, LCHADD- and carnitine transporter deficiencies. The genetic phenotypes of one enzyme defect did not result in differing acylcarnitine patterns in MCADD, VLCADD or LCHADD in vitro.
摘要：
本研究的目的是建立一种非侵入性的，在提供信息的新生儿筛查或对怀疑患有FAOD的患者进行选择性筛查后，快速可靠的酶促测定可确认人类的脂肪酸氧化缺陷（FAOD）。
在具有特定酶缺陷的FAOD患者的全血中测试了该方法的可靠性。在30个中链中测定全血样品-(MCADD，年龄0至17岁)，6非常长的链条-（VLCADD，年龄0至4岁)，6长链羟基-(LCHAD，1至6岁)，3短链-(SCADD,年龄10至13岁)酰基-CoA-脱氢酶-和2个初级肉碱转运蛋白缺乏症(CTD，3至5岁)。此外,26名健康儿童（0至17岁）作为对照。将全血样品与稳定的末端标记的棕榈酸酯一起孵育;通过串联质谱法分析标记的酰基肉碱,并与对照和患者组之间进行比较(Mann-Whitney等级和检验)。在特定的潜在MCADD-（ANOVA）之间比较了特定标记的酰基肉碱代谢物的浓度，VLCADD-和LCHADD-遗传变异(描述性数据分析)。
分析了11种不同的酰基肉碱。MCADD-(C8-,C10-肉碱,C8/C10-和C8/C4-肉碱)，VLCADD-(C12-,C14:1-,C14:2-肉碱,C14：1/C12-和C14：2/C12-肉碱），LCHADD（C16-OH-肉碱）以及CTD-缺乏（所有酰基肉碱的总和）样品可以清楚地识别并与对照值以及其他FAOD分离。而FAOD样品之间所有酰基肉碱的总和并不是决定性的。此外，C4-(SCADD),C14-（VLCADD）和C14-OH-肉碱（LCHADD）在FAOD组之间有区别。代谢参数在潜在的MCADD变体之间没有显着差异;对于VLCADD-和LCHADD-变体可以观察到类似的结果。
这种在全血样本中的功能方法相对简单，非侵入性和很少的时间消耗。它允许识别MCADD-，VLCADD-,LCHADD-和肉碱转运蛋白缺乏。一种酶缺陷的遗传表型并没有导致MCADD中不同的酰基肉碱模式，体外VLCADD或LCHADD。
在具有特定酶缺陷的FAOD患者的全血中测试了该方法的可靠性。在30个中链中测定全血样品-(MCADD，年龄0至17岁)，6非常长的链条-（VLCADD，年龄0至4岁)，6长链羟基-(LCHAD，1至6岁)，3短链-(SCADD,年龄10至13岁)酰基-CoA-脱氢酶-和2个初级肉碱转运蛋白缺乏症(CTD，3至5岁)。此外,26名健康儿童（0至17岁）作为对照。将全血样品与稳定的末端标记的棕榈酸酯一起孵育;通过串联质谱法分析标记的酰基肉碱,并与对照和患者组之间进行比较(Mann-Whitney等级和检验)。在特定的潜在MCADD-（ANOVA）之间比较了特定标记的酰基肉碱代谢物的浓度，VLCADD-和LCHADD-遗传变异(描述性数据分析)。
分析了11种不同的酰基肉碱。MCADD-(C8-,C10-肉碱,C8/C10-和C8/C4-肉碱)，VLCADD-(C12-,C14:1-,C14:2-肉碱,C14：1/C12-和C14：2/C12-肉碱），LCHADD（C16-OH-肉碱）以及CTD-缺乏（所有酰基肉碱的总和）样品可以清楚地识别并与对照值以及其他FAOD分离。而FAOD样品之间所有酰基肉碱的总和并不是决定性的。此外，C4-(SCADD),C14-（VLCADD）和C14-OH-肉碱（LCHADD）在FAOD组之间有区别。代谢参数在潜在的MCADD变体之间没有显着差异;对于VLCADD-和LCHADD-变体可以观察到类似的结果。
这种在全血样本中的功能方法相对简单，非侵入性和很少的时间消耗。它允许识别MCADD-，VLCADD-,LCHADD-和肉碱转运蛋白缺乏。一种酶缺陷的遗传表型并没有导致MCADD中不同的酰基肉碱模式，体外VLCADD或LCHADD。
