sphingosine-1-phosphate and Leukodystrophy--Globoid-Cell

Globoid cell leukodystrophy (Krabbe disease) is a rare infantile neurodegenerative disorder. Krabbe disease is caused by deficiency in the lysosomal enzyme galactocerebrosidase (GALC) resulting in accumulation, in the micromolar range, of the toxic metabolite galactosylsphingosine (psychosine) in the brain. Here we find that psychosine induces human astrocyte cell death probably via an apoptotic process in a concentration- and time-dependent manner (EC50 ∼ 15 μM at 4 h). We show these effects of psychosine are attenuated by pre-treatment with the sphingosine 1-phosphate receptor agonist pFTY720 (fingolimod) (IC50 ∼ 100 nM). Psychosine (1 μM, 10 μM) also enhances LPS-induced (EC50 ∼ 100 ng/ml) production of pro-inflammatory cytokines in mouse astrocytes, which is also attenuated by pFTY720 (1 μM). Most notably, for the first time, we show that psychosine, at a concentration found in the brains of patients with Krabbe disease (EC50 ∼ 100 nM), directly induces demyelination in mouse organotypic cerebellar slices in a manner that is independent of pro-inflammatory cytokine response and that pFTY720 (0.1 nM) significantly inhibits. These results support the idea that psychosine is a pathogenic agent in Krabbe disease and suggest that sphingosine 1-phosphate signalling could be a potential drug target for this disorder.

Topics: Animals; Apoptosis; Astrocytes; Demyelinating Diseases; Fingolimod Hydrochloride; Humans; Leukodystrophy, Globoid Cell; Lipopolysaccharides; Lysophospholipids; Mice; Microscopy, Fluorescence; Psychosine; Rats; Signal Transduction; Sphingosine

Globoid cell leukodystrophy (Krabbe disease) is caused by mutations in galactosylceramidase, a lysosomal enzyme that acts to digest galactosylceramide, a glycolipid concentrated in myelin, and psychosine (galactosylsphingosine). Globoid cell leukodystrophy has been identified in many species including humans and twitcher mice. Several studies on human tissue have examined the lipid profile in this disease by gas, liquid or thin layer chromatography. Electrospray ionization tandem mass spectrometry combined with reverse phase HPLC has become a powerful alternative strategy, used here to compare the sphingolipid profile of pons/medulla tissue from twitcher mice with control tissue. In this lipidomics LC-MS approach, we scanned for precursors of m/z 264 to obtain a semi-quantitative profile of ceramides and galactosylceramides. Sphingosine-1-phosphate, C18:0 ceramide, C22:0 ceramide and C24:0 ceramide levels were reduced in the pons/medulla of twitcher mice compared to levels in control mice at 31 and 35-37 days of age. The levels of C22:0 and C24:0 galactosylceramide were similar between twitcher and control specimens and there was a trend toward reduced levels of C24:1 galactosylceramide and C24:1 hydroxy-galactosylceramide in twitcher specimens. Psychosine, C 16:0 ceramide and C 18:0 galactosylceramide levels were increased in the CNS of twitcher mice compared to levels in control mice. These data indicate that there is a trend toward decreased levels of long chain fatty acids and increased levels of shorter chain fatty acids in galactosylceramides and ceramides from twitcher mice compared with control mice, and such changes may be due to demyelination characteristic of acute pathology.

Topics: Animals; Central Nervous System; Ceramides; Chromatography, High Pressure Liquid; Chromatography, Liquid; Disease Models, Animal; Fatty Acids; Galactosylceramides; Leukodystrophy, Globoid Cell; Lysophospholipids; Mass Spectrometry; Medulla Oblongata; Mice; Mice, Inbred C57BL; Mice, Neurologic Mutants; Pons; Psychosine; Sphingolipids; Sphingosine