Lipidomic characterization of exosomes isolated from human plasma using various mass spectrometry techniques.

Ultrahigh-performance supercritical fluid chromatography - mass spectrometry (UHPSFC/MS), ultrahigh-performance liquid chromatography - mass spectrometry (UHPLC/MS), and matrix-assisted laser desorption/ionization (MALDI) - MS techniques were used for the lipidomic characterization of exosomes isolated from human plasma. The high-throughput methods UHPSFC/MS and UHPLC/MS using a silica-based column containing sub-2 µm particles enabled the lipid class separation and the quantitation based on exogenous class internal standards in <7 minute run time. MALDI provided the complementary information on anionic lipid classes, such as sulfatides. The nontargeted analysis of 12 healthy volunteers was performed, and absolute molar concentration of 244 lipids in exosomes and 191 lipids in plasma belonging to 10 lipid classes were quantified. The statistical evaluation of data included principal component analysis, orthogonal partial least square discriminant analysis, S-plots, p-values, T-values, fold changes, false discovery rate, box plots, and correlation plots, which resulted in the information on lipid changes in exosomes in comparison to plasma. The major changes were detected in the composition of triacylglycerols, diacylglycerols, phosphatidylcholines, and lysophosphatidylcholines, whereby sphingomyelins, phosphatidylinositols, and sulfatides showed rather similar profiles in both biological matrices.

N-butyldeoxynojirimycin delays motor deficits, cerebellar microgliosis, and Purkinje cell loss in a mouse model of mucolipidosis type IV.

Mucolipidosis type IV (MLIV) is a lysosomal storage disease exhibiting progressive intellectual disability, motor impairment, and premature death. There is currently no cure or corrective treatment. The disease results from mutations in the gene encoding mucolipin-1, a transient receptor potential channel believed to play a key role in lysosomal calcium egress. Loss of mucolipin-1 and subsequent defects lead to a host of cellular aberrations, including accumulation of glycosphingolipids (GSLs) in neurons and other cell types, microgliosis and, as reported here, cerebellar Purkinje cell loss. Several studies have demonstrated that N-butyldeoxynojirimycin (NB-DNJ, also known as miglustat), an inhibitor of the enzyme glucosylceramide synthase (GCS), successfully delays the onset of motor deficits, improves longevity, and rescues some of the cerebellar abnormalities (e.g., Purkinje cell death) seen in another lysosomal disease known as Niemann-Pick type C (NPC). Given the similarities in pathology between MLIV and NPC, we examined whether miglustat would be efficacious in ameliorating disease progression in MLIV. Using a full mucolipin-1 knockout mouse (Mcoln1-/-), we found that early miglustat treatment delays the onset and progression of motor deficits, delays cerebellar Purkinje cell loss, and reduces cerebellar microgliosis characteristic of MLIV disease. Quantitative mass spectrometry analyses provided new data on the GSL profiles of murine MLIV brain tissue and showed that miglustat partially restored the wild type profile of white matter enriched lipids. Collectively, our findings indicate that early miglustat treatment delays the progression of clinically relevant pathology in an MLIV mouse model, and therefore supports consideration of miglustat as a therapeutic agent for MLIV disease in humans.