Chemistry and Metabolism of Sphingolipids. 3-Oxo Derivatives of N-Acetylsphingosine and N-Acetyldihydrosphingosine

The ceramide (Cer) and sphingomyelin (SM) species of cultured differentiated rat cerebellar granule cells and human fibroblasts were characterized by electrospray ionization-mass spectrometry. We identified 35 different species of Cer and 18 species of SM in human fibroblasts, and 35 different species of Cer and 9 species of SM were characterized in rat neurons. The main Cer species of rat cerebellar granule cells contained d18:1 sphingosine linked with palmitic, stearic, or nervonic fatty acid, and the two main SM species were d18:1,16:0 and d18:1,18:0. Both sphingolipids were enriched in detergent-resistant membranes (DRMs; or lipid rafts), and significant differences were found in the sphingolipid patterns of DRMs and of detergentsoluble fractions (DSF) from these cells. In human fibroblasts, the main Cer species were d18:1,16:0, d18:2,16:0, d18:1,24:0, d18:2,24:0, d18:1,24:1, and d18:2,24:1; the most represented species of SM were d18:1,16:0, d18:1,24:0, and d18:1,24:1. In these cells, SM was highly enriched in DRMs and Cer was mainly associated with DSF, and the species found in DRMs were markedly different from those found in DSF.-Valsecchi, M., L. Mauri, R. Casellato, S. Prioni, N. Loberto, A. Prinetti, V. Chigorno, and S. Sonnino. Ceramide and sphingomyelin species of fibroblasts and neurons in culture. J. Lipid Res. 2007. 48: 417-424.

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“…Lyso-SM was from Avanti Polar Lipids. C17-Cer was prepared by acylation of sphingosine and C17-SM of lyso-SM using heptadecanoic anhydride (18).…”

Section: Methodsmentioning

confidence: 99%

Ceramide and sphingomyelin species of fibroblasts and neurons in culture

Valsecchi

Mauri

Casellato

et al. 2007

Journal of Lipid Research

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“…[1-3 H]sphingosine was prepared by specific chemical oxidation of the primary hydroxyl group of sphingosine followed by reduction with sodium boro[ 3 H]hydride (44) (radiochemical purity over 98%; specific radioactivity 2.2 Ci/mmol). N-Acetylsphingosine (C2Cer) and N-palmitoylsphingosine (C16Cer) were prepared by N-acylation of sphingosine using acetic anhydride and hexadecanoic anhydride, respectively (45). The same procedure applied to [1-3 H]sphingosine was used to synthesize tritium-labeled N-palmitoylsphingosine (radiochemical purity over 99%; specific radioactivity 2.2 Ci/mmol).…”

Section: Methodsmentioning

confidence: 99%

Altered Sphingolipid Metabolism inN-(4-Hydroxyphenyl)- retinamide-resistant A2780 Human Ovarian Carcinoma Cells

Prinetti

Basso

Appierto

et al. 2003

Journal of Biological Chemistry

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In the present work, we studied the effects of fenretinide (N-(4-hydroxyphenyl)retinamide (HPR)), a hydroxyphenyl derivative of all-trans-retinoic acid, on sphingolipid metabolism and expression in human ovarian carcinoma A2780 cells. A2780 cells, which are sensitive to a pharmacologically achievable HPR concentration, become 10-fold more resistant after exposure to increasing HPR concentrations. Our results showed that HPR was able to induce a dose-and time-dependent increase in cellular ceramide levels in sensitive but not in resistant cells. This form of resistance in A2780 cells was not accompanied by the overexpression of multidrug resistance-specific proteins MDR1 P-glycoprotein and multidrug resistance-associated protein, whose mRNA levels did not differ in sensitive and resistant A2780 cells. HPR-resistant cells were characterized by an overall altered sphingolipid metabolism. The overall content in glycosphingolipids was similar in both cell types, but the expression of specific glycosphingolipids was different. Specifically, our findings indicated that glucosylceramide levels were similar in sensitive and resistant cells, but resistant cells were characterized by a 6-fold lower expression of lactosylceramide levels and by a 6-fold higher expression of ganglioside levels than sensitive cells. The main gangliosides from resistant A2780 cells were identified as GM3 and GM2. The possible metabolic mechanisms leading to this difference were investigated. Interestingly, the mRNA levels of glucosylceramide and lactosylceramide synthases were similar in sensitive and resistant cells, whereas GM3 synthase mRNA level and GM3 synthase activity were remarkably higher in resistant cells.

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“…Long-chain bases were purified from the hydrolysate extract by TLC using the conditions described above. Long-chain bases were converted to N-acetyl derivates by reaction with 0.1 mL of methanol:acetic anhydride, 4:1 (v/v), for 16 to 18 h at room temperature (9). Long-chain base derivatives were dried under nitrogen, dissolved in chloroform:methanol:water, 1:1:0.8 (v/v/v), and recovered in the Figure 1 b (i) = trans isomer (tentative identification).…”

Section: Gc/ms and Gc Analyses Of Glucocerebroside Hydrolysatesmentioning

confidence: 99%

Analysis of Glucocerebrosides of Rye (Secale cereale L. cv Puma) Leaf and Plasma Membrane

Cahoon¹,

Lynch²

1991

Plant Physiol.

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Glucocerebrosides of whole rye (Secale cerale L. cv Puma) leaf and plasma membrane were analyzed using gas chromatography-mass spectrometry and gas chromatography following hydrolysis or as intact molecules purified by reverse-phase high performance liquid chromatography. Fatty acids of acid-hydrolyzed leaf and plasma membrane glucocerebrosides consisted of >98 weight percent saturated and monounsaturated 2-hydroxy fatty acids which contained 16 to 26 carbon atoms. The major fatty acids detected were 2-hydroxynervonic acid (24:1h), 2-hydroxylignoceric acid (24:0h), 2-hydroxyerucic acid (22:1h), and 2-hydroxybehenic acid (22:0h). Long-chain bases of alkalinehydrolyzed glucocerebrosides consisted primarily of cis-trans isomers of the trihydroxy base 4-hydroxysphingenine (t18:1) and the dihydroxy base sphingadienine (dl8:2) with lesser amounts of 4-hydroxysphinganine (t18:0) and isomers of sphingenine (d18: 1). Intact, underivatized glucocerebroside molecular species of rye leaf and plasma membrane were separated into more than 30 molecular species using reverse-phase HPLC. The molecular species composition of leaf and plasma membrane were quantitatively and qualitatively similar. The major molecular species was 24:1h-t18:1 which constituted nearly 40 weight percent of leaf and plasma membrane extracts. Several other species including 22:1h-t18:1, 24:1h-t18:1 (isomer), 22:0h-t18:1, 24:1h-d18:2, and 24:0h-t18:1 each comprised 4 to 8% of the total. It is anticipated that the high performance liquid chromatography procedure developed in this study to separate intact, underivatized lipid molecular species will be useful in future studies of the physical properties and biosynthesis of plant glucocerebrosides.Glucocerebrosides are a class of glycosphingolipids that have been reported to constitute less than 1% of the total lipid extract of plant tissues including runner bean leaves (29) and rice bran (8). Recently, however, glucocerebrosides have been identified as a major lipid component of the plasma membrane and tonoplast of cells of several plant species. Plasma membrane isolated from rye (18) and barley (27) leaves, oat root (28), oat coleoptile (28), and mung bean hypocotyl (33)

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Chemistry and Metabolism of Sphingolipids. 3-Oxo Derivatives of N-Acetylsphingosine and N-Acetyldihydrosphingosine

Cited by 182 publications

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Ceramide and sphingomyelin species of fibroblasts and neurons in culture

Ceramide and sphingomyelin species of fibroblasts and neurons in culture

Altered Sphingolipid Metabolism inN-(4-Hydroxyphenyl)- retinamide-resistant A2780 Human Ovarian Carcinoma Cells

Analysis of Glucocerebrosides of Rye (Secale cereale L. cv Puma) Leaf and Plasma Membrane

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