Quantification of sulfatides and lysosulfatides in tissues and body fluids by liquid chromatography-tandem mass spectrometry

Abstract

This article is available online at http://www.jlr.org high concentrations are found in selected organs, including kidney, colon, pancreas, and gall bladder. Increased concentrations of sulfatides have been reported in renal cell carcinoma, adenocarcinoma of colon and lung, and ovarian cancer ( 2, 4 ). Sulfatides have been proposed to play a role as ion barriers to osmotic stress and as counterions of ammonium in the renal adaptation to chronic metabolic acidosis ( 5,6 ) Sulfatides are synthesized from their precursor galactosylceramide in the Golgi apparatus by 3-O-sulfation of the galactose residue by the enzyme 3 ′ -phosphoadenosine-5 ′ -phosphosulfate: cerebroside sulfotransferase (CST) (EC2.8.2.11). Galactosylceramide is synthesized in the endoplasmic reticulum from ceramides and UDP-galactose by the enzyme UDP-galactose:ceramide galactosyltransferase (CGT) (EC 2.4.1.45) and is then transported to the Golgi apparatus prior to sulfation to sulfatides. Sulfatides consist of many molecular species, with structures differing in acyl chain length and hydroxylation and sphingoid base. These sulfatide molecular species are cell and tissue specifi c, which may be explained by the cell-and tissuespecifi c expression of ceramide synthases and fatty acid 2-hydroxylase ( 1, 7 ). It is diffi cult to assign specifi c functions to the different molecular species of sulfatides, but sulfatides with C16:0 fatty acids were reported to be involved in the regulation of insulin secretion in rat pancreatic ␤ -cells ( 8 ), and elevated levels of C18:0 sulfatides have been implicated as a cause of audiogenic seizers in mice overexpressing CGT ( 9 ). There is also heterogeneity of the sphingoid base composition of sulfatides. Most of the Abstract Sulfatides are found in brain as components of myelin, oligodendrocytes, and neurons but are also present in various visceral tissues. Metachromatic leukodystrophy (MLD) is an inherited lysosomal storage disorder caused by a defi ciency of arylsulfatase A, leading to severe white matter disease due to the accumulation of sulfatides and lysosulfatides. To study the physiological role of sulfatides, accessible and sensitive quantitative methods are required. We developed a sensitive LC/MS/MS method to quantify total sulfatide and lysosulfatide content as well as individual molecular species in urine and plasma from MLD patients and plasma and tissues from an MLD mouse model. Our results demonstrate that the method can quantify a wide range of sulfatide concentrations and can be used to quantify total sulfatide content and levels of individual molecular species of sulfatides in tissues, cells, and body fl uids. Even though plasma sulfatides and lysosulfatides would seem attractive candidate biomarkers that could possibly correlate with the severity of MLD and be of use to monitor the effects of therapeutic intervention, our results indicate that it is unlikely that the determination of these storage products in plasma will be useful in this respect. Sulfatides are anionic sulfoglycolipids main...

A Sensitive and Wide Coverage Ambient Mass Spectrometry Imaging Method for Functional Metabolites Based Molecular Histology

“…As shown in Figure 3 , ST almost just located in renal medulla with a high signal‐to‐noise ratio (Figure 3A). The ion intensity ratio of C24:0‐OH‐ST (Figure 3A2) to C24:1‐OH‐ST (Figure 3A3) was consistent with previous quantitative LC‐MS/MS study 11. As for low‐abundance C20:0‐OH‐ST (Figure 3A4), though the ion intensity is 4.3E 2 , a renal medulla‐specific distribution was still clearly presented.…”

“…In rat brain, STs were primarily found in corpus callosum and internal capsule (Figure 3B). What is more meaningful is that the low‐abundance C20:0‐OH‐ST (Figure 3B4), which has been identified as being unique to kidney and undetected in brain by LC‐MS/MS,11 still demonstrated a region‐specific distribution. To our knowledge, this is the first report on the detection of low‐abundance C20:0‐OH‐ST by MSI.…”

High-throughput analysis of sulfatides in cerebrospinal fluid using automated extraction and UPLC-MS/MS

“…Han and coworkers have analyzed STs in CSF, but have not reported the concentration of individual STs (14,39). However, for reliable measurement, 0.5-1.0 ml of CSF was needed, indicating a lower LOQ, which can in part be explained by the use of direct infusion MS. For plasma LOQ, values between 5 and 50 nmol/l have been reported (23,26,28,38). In this report, we chose to only focus on CSF as matrix, but we believe that the method could also be applied to other matrices.…”

“…Similar to many other groups, we used reversed-phase chromatography for the separation of ST species prior to MS detection (23,24,26,28,38). Using this approach, the separation of individual species was mainly based on the difference in fatty acid moieties attached to the sphingosine backbone.…”

“…In the last 15 years several methods have been described using TOF-secondary ion MS (19), MALDI-TOF MS (20,21), HPLC-MS (22)(23)(24)(25)(26), and direct infusion MS (15,27) for measuring ST species in different biological matrices. Quantitative and qualitative analyses of STs by LC-MS/MS have emerged as a powerful tool in research (28) and also in clinical diagnostics (29,30).…”