Pettazzoni Magali, Froissart Roseline, Pagan Cécile, Vanier Marie T, Ruet Séverine, Latour Philippe, Guffon Nathalie, Fouilhoux Alain, Germain Dominique P, Levade Thierry, Vianey-Saban Christine, Piraud Monique, Cheillan David
Service de Biochimie et Biologie Moléculaire Grand Est, Unité Médicale Pathologies Métaboliques, Erythrocytaires et Dépistage Périnatal, Centre de Biologie et de Pathologie Est, Hospices Civils de Lyon, Bron, France.
Unité Mixte de Recherche 5305, Centre National de la Recherche Scientifique (CNRS) Université Claude Bernard Lyon 1, Lyon, France.
PLoS One. 2017 Jul 27;12(7):e0181700. doi: 10.1371/journal.pone.0181700. eCollection 2017.
The biological diagnosis of sphingolipidoses currently relies on the measurement of specific enzymatic activities and/or genetic studies. Lysosphingolipids have recently emerged as potential biomarkers of sphingolipidoses and Niemann-Pick type C in plasma.
We developed a sensitive and specific method enabling the simultaneous quantification of lysosphingolipids by LC-MS/MS: lysoglobotriaosylceramide for Fabry disease, lysohexosylceramide (i.e. lysoglucosylceramide and/or lysogalactosylceramide) for Gaucher and Krabbe diseases, lysosphingomyelin and its carboxylated analogue lysosphingomyelin-509 for Niemann-Pick type A or B, and C diseases, lysoGM1 ganglioside for GM1gangliosidosis and lysoGM2 ganglioside for GM2 gangliosidosis.
The diagnostic performances were validated in plasma samples analysing a large series of patients affected with sphingolipidoses and Niemann-Pick type C disease (n = 98), other inborn errors of metabolism (n = 23), and controls (n = 228). The multiplex measurement of lysosphingolipids allowed the screening of Fabry (including female patients and late-onset variants), Gaucher and infantile Krabbe, Niemann-Pick type A/B and C diseases with high sensitivity and specificity. LysoGM1 and LysoGM2 were elevated in most of the patients affected with GM1 and GM2 gangliosidosis respectively. In amniotic fluid supernatant from pregnancies presenting non-immune hydrops fetalis (n = 77, including previously diagnosed Gaucher (n = 5), GM1 gangliosidosis (n = 4) and galactosialidosis (n = 4) fetuses) and from normal pregnancies (n = 15), a specific and dramatic increase of lysohexosylceramide was observed only in the Gaucher amniotic fluid samples.
This multiplex assay which allows the simultaneous measurement of lysosphingolipids in plasma modifies the diagnostic strategy of sphingolipidoses and Niemann-Pick type C. Furthermore, in pregnancies presenting non-immune hydrops fetalis, lysohexosylceramide measurement in amniotic fluid offers a rapid screening of fetal Gaucher disease without waiting for glucocerebrosidase activity measurement in cultured amniocytes.
鞘脂类疾病的生物学诊断目前依赖于特定酶活性的测定和/或基因研究。溶血鞘脂最近已成为血浆中鞘脂类疾病和尼曼-皮克C型病的潜在生物标志物。
我们开发了一种灵敏且特异的方法，可通过液相色谱-串联质谱法（LC-MS/MS）同时定量溶血鞘脂：用于法布里病的溶血球三糖神经酰胺，用于戈谢病和克拉伯病的溶血己糖神经酰胺（即溶血葡萄糖神经酰胺和/或溶血半乳糖神经酰胺），用于尼曼-皮克A型或B型以及C型病的溶血鞘磷脂及其羧化类似物溶血鞘磷脂-509，用于GM1神经节苷脂贮积症的溶血GM1神经节苷脂和用于GM2神经节苷脂贮积症的溶血GM2神经节苷脂。
在分析大量患有鞘脂类疾病和尼曼-皮克C型病（n = 98）、其他先天性代谢缺陷（n = 23）以及对照（n = 228）的患者的血浆样本中验证了诊断性能。溶血鞘脂的多重测量能够以高灵敏度和特异性筛查法布里病（包括女性患者和迟发型变体）、戈谢病、婴儿型克拉伯病、尼曼-皮克A型/B型和C型病。溶血GM1和溶血GM2分别在大多数患有GM1和GM2神经节苷脂贮积症的患者中升高。在出现非免疫性胎儿水肿的妊娠羊水上清液（n = 77，包括先前诊断为戈谢病（n = 5）、GM1神经节苷脂贮积症（n = 4）和唾液酸贮积症（n = 4）的胎儿）以及正常妊娠（n = 15）中，仅在戈谢病羊水样本中观察到溶血己糖神经酰胺有特异性且显著的升高。
这种能够同时测量血浆中溶血鞘脂的多重检测方法改变了鞘脂类疾病和尼曼-皮克C型病的诊断策略。此外，在出现非免疫性胎儿水肿的妊娠中，羊水溶血己糖神经酰胺的测量无需等待培养羊水中葡萄糖脑苷脂酶活性的测定，即可快速筛查胎儿戈谢病。
