Tandem Mass Spectrometry Multiplex Analysis of Glucosylceramide and Galactosylceramide Isoforms in Brain Tissues at Different Stages of Parkinson Disease
Tandem Mass Spectrometry Multiplex Analysis of Glucosylceramide and Galactosylceramide Isoforms in Brain Tissues at Different Stages of Parkinson Disease
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DOI：
10.1021/acs.analchem.5b04227
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发表时间：
2016-02-02
影响因子：
7.4
通讯作者：
Auray-Blais, Christiane
中科院分区：
文献类型：
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作者：
Boutin, Michel;Sun, Ying;Auray-Blais, Christiane
先前的研究表明，帕金森病（PD）与脑组织中葡萄糖脑苷酶（GCase）酶活性降低有关。本研究的目的是确定GCase缺乏是否与PD脑组织中葡萄糖神经酰胺（GluCer）底物的积累有关。建立了超高效液相色谱串联质谱（UPLC-MS/MS）方法，对C18:0、C20:0、C22:0、C24:1和C24:0的多重分析方法进行了优化和验证。这些分子用正相色谱法从它们的等压半乳糖神经酰胺（GalCer）分子中分离出来。采用多反应监测（MRM）采集模式，采用串联质谱法进行分析。所分析的不同葡聚糖同工异构体的脑组织检出限为0.4 ~ 1.1 nmol/g。根据路易体疾病统一分期系统，我们首次分析了26例PD患者的颞皮质脑组织样本中葡萄糖醛酸盐的水平，这些患者被分为三个PD疾病阶段（IIa， III和IV）。将这些脑组织标本与12例对照组和6例偶发性路易体病患者的脑组织标本进行比较。5个样品组间葡萄糖浓度无显著差异。葡萄糖异构体水平也与其匹配的GalCer异构体归一化。归一化结果显示，胰高血糖水平随PD严重程度的增加而增加。然而，由于测量到的高标准偏差，两组之间观察到的差异并不显著。
Previous studies demonstrated that Parkinson disease (PD) is associated with a decreased activity of the glucocerebrosidase (GCase) enzyme in brain tissues. The objective of this study was to determine if GCase deficiency is associated with the accumulation of its glucosylceramide (GluCer) substrate in PD brain tissues. An ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed, optimized, and validated for the multiplex analysis of GluCer isoforms (C18:0, C20:0, C22:0, C24:1, and C24:0) in brain tissue samples. These molecules were chromatographically separated from their isobaric galactosylceramide (GalCer) counterparts using normal phase chromatography. The analysis was performed by tandem mass spectrometry in the multiple reaction monitoring (MRM) acquisition mode. Limits of detection ranging from 0.4 to 1.1 nmol/g brain tissue were established for the different GluCer isoforms analyzed. For the first time, GluCer isoform levels were analyzed in temporal cortex brain tissue samples from 26 PD patients who were divided into three PD disease stages (IIa, III, and IV) according to the Unified Staging System for Lewy Body Disorders. These specimens were compared with brain tissue samples from 12 controls and 6 patients with Incidental Lewy Body Disease. No significant GluCer concentration differences were observed between the 5 sample groups. The GluCer isoform levels were also normalized with their matching GalCer isoforms. The normalized results showed a trend for GluCer levels which increased with PD severity. However, the differences observed between the groups were not significant, owing likely to the high standard deviations measured.