35 research outputs found
Update on LIPID MAPS classification, nomenclature, and shorthand notation for MS-derived lipid structures
A comprehensive and standardized system to report lipid structures analyzed by mass spectrometry is essentialfor the communication and storage of lipidomics data. Herein, an update on both the LIPID MAPSclassification system and shorthand notation of lipid structures is presented for lipid categories Fatty Acyls(FA), Glycerolipids (GL), Glycerophospholipids (GP), Sphingolipids (SP), and Sterols (ST). With its majorchanges, i.e. annotation of ring double bond equivalents and number of oxygens, the updated shorthandnotation facilitates reporting of newly delineated oxygenated lipid species as well. For standardized reportingin lipidomics, the hierarchical architecture of shorthand notation reflects the diverse structural resolutionpowers provided by mass spectrometric assays. Moreover, shorthand notation is expanded beyond mammalianphyla to lipids from plant and yeast phyla. Finally, annotation of atoms is included for the use of stableisotope-labelled compounds in metabolic labelling experiments or as internal standards
Oxidized phospholipids induce ceramide accumulation in RAW 264.7 macrophages: role of ceramide synthases.
Oxidized phospholipids (OxPLs), including 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphocholine (PGPC) and 1-palmitoyl-2-oxovaleroyl-sn-glycero-3-phosphocholine (POVPC) are among several biologically active derivatives that are generated during oxidation of low-density lipoproteins (LDLs). These OxPLs are factors contributing to pro-atherogenic effects of oxidized LDLs (OxLDLs), including inflammation, proliferation and death of vascular cells. OxLDL also elicits formation of the lipid messenger ceramide (Cer) which plays a pivotal role in apoptotic signaling pathways. Here we report that both PGPC and POVPC are cytotoxic to cultured macrophages and induce apoptosis in these cells which is associated with increased cellular ceramide levels after several hours. In addition, exposure of RAW 264.7 cells to POVPC and PGPC under the same conditions resulted in a significant increase in ceramide synthase activity, whereas, acid or neutral sphingomyelinase activities were not affected. PGPC is not only more toxic than POVPC, but also a more potent inducer of ceramide formation by activating a limited subset of CerS isoforms. The stimulated CerS activities are in line with the C16-, C22-, and C24:0-Cer species that are generated under the influence of the OxPL. Fumonisin B1, a specific inhibitor of CerS, suppressed OxPL-induced ceramide generation, demonstrating that OxPL-induced CerS activity in macrophages is responsible for the accumulation of ceramide. OxLDL elicits the same cellular ceramide and CerS effects. Thus, it is concluded that PGPC and POVPC are active components that contribute to the capacity of this lipoprotein to elevate ceramide levels in macrophages
Effect of lipid extracts from LDL and OxLDL on CerS activity.
<p>(A) Oxidation of LDL was performed as described under “Experimental Procedure”. RAW 264.7 cells were stimulated with lipid extracts from native LDL and OxLDL (50 µg protein/mL respectively) for 24 h. CerS activity in cell homogenates was measured using C<sub>16</sub>-CoA and C<sub>22</sub>-CoA substrates. Results are means ± S.D., *p < 0.05, of a typical experiment repeated four times with similar results. (B) Cells were treated with intact LDL and OxLDL (50 µg protein/mL respectively) as described above for 24 h. CerS activity in cell homogenates was measured using C<sub>16</sub>-CoA and C<sub>22</sub>-CoA substrates. Results are means ± S.D. *p < 0.05, of a typical experiment repeated four times with similar results. (C) Cells were treated as described above and lipids were extracted and analyzed for ceramide content as described under “Experimental Procedure”. No probability values are given for total ceramide levels because these levels are the sum of ceramide species with different acyl chain lengths. (D) Ceramide species were analyzed after treatment with lipid extracts from native LDL and OxLDL as described earlier. The data are means ± S.E., *p < 0.05, n = 4.</p
Influence of Fenofibrate treatment on triacylglycerides diacylglycerides and fatty acids in fructose fed rats
Fenofibrate (FF) lowers plasma triglycerides via PPARα activation. Here, we analyzed lipidomic changes upon FF treatment of fructose fed rats. Three groups with 6 animals each were defined as control, fructose-fed and fructose-fed/FF treated. Male Wistar Unilever Rats were subjected to 10% fructose-feeding for 20 days. On day 14, fenofibrate treatment (100 mg/kg p.o.) was initiated and maintained for 7 days. Lipid species in serum were analyzed using mass spectrometry (ESI-MS/MS; LC-FT-MS, GC-MS) on days 0, 14 and 20 in all three groups. In addition, lipid levels in liver and intestine were determined. Short-chain TAGs increased in serum and liver upon fructose-feeding, while almost all TAG-species decreased under FF treatment. Long-chain unsaturated DAG-levels (36:1, 36:2, 36:4, 38:3, 38:4, 38:5) increased upon FF treatment in rat liver and decreased in rat serum. FAs, especially short-chain FAs (12:0, 14:0, 16:0) increased during fructose-challenge. VLDL secretion increased upon fructose-feeding and together with FA-levels decreased to control levels during FF treatment. Fructose challenge of de novo fatty acid synthesis through fatty acid synthase (FAS) may enhance the release of FAs ≤16:0 chain length, a process reversed by FF-mediated PPARα-activation
Additional file 1: Figure S1. of Methyl group donors abrogate adaptive responses to dietary restriction in C. elegans
Similar changes of the triglyceride-to-phospholipid ratio and the fatty acid composition of phosphatidylcholine (PC) in dietary restricted L4 larvae (P0 generation) compared to embryos (F1 generation) obtained from dietary restricted worms. (PPTX 106 kb
Update on guidelines for lipidomics analysis and reporting
Ekroos K, Ejsing C, Liebisch G, et al. Update on guidelines for lipidomics analysis and reporting. Journal of the American Oil Chemists' Society. 2022;99(Suppl. 1):13-14
Influence of POVPC and PGPC on ceramide levels and CerS activation in RAW 264.7 cells.
<p>(A) Both POVPC and PGPC elevate ceramide generation in RAW 264.7 cells. Cells were stimulated for 24 h with respective OxPLs (50 µM) in parallel to ethanol treated control cells. Lipids were extracted and analyzed for ceramide levels by LC/MS-MS as described under “Experimental Procedure”. No probability. values are given for total ceramide levels because these levels are the sum of ceramide species with different acyl chain lengths. (B) Ceramide speciation was performed after OxPL treatment as above. The data are means ± S.E., *p<u><</u>0.05, **p < 0.01 compared with control, n = 4. (C) After OxPL treatment cells were harvested and CerS activity in cell homogenates was measured as described earlier. Results are means ± S.E., *p < 0.01, **p < 0.05, of a typical experiment repeated four times with similar results. (D) CerS mRNA levels were measured by RT-qPCR after 24 h incubation with 50 µM POVPC or PGPC as described under “Experimental Procedures”. The data are normalized to GAPDH mRNA expression and data are means ± S.E. for four independent experiments performed in triplicate. (E) nSMase activity was measured in cell homogenates after exposure to OxPL as described under “Experimental Procedures”. The data are represented as means ± S.E., n = 4.</p
Effect of FB1 on OxPL induced CerS activation and ceramide levels.
<p>(A) Raw 264.7 cells were pre-incubated with FB1 (20 µM) for 2 h prior to the 24 h OxPL treatment. Lipids were extracted and ceramide levels were analyzed as described under “Experimental Procedure”. No probability values are given for total ceramide levels because these levels are the sum of ceramide species with different acyl chain lengths. (B) Ceramide species were analyzed as earlier. The data are means ± S.E., *p < 0.05, **p < 0.01 compared with control, n = 4.</p
The oxidized phospholipids POVPC and PGPC are cytotoxic and induce apoptosis.
<p>(A) Chemical structures of POVPC and PGPC. Dotted circle represents the functional group at <i>sn-2</i> position. (B) RAW 264.7 cells were incubated with 50 µM POVPC and PGPC for indicated periods. Control cells were treated with 1% ethanol. Cell viability was determined by Vybrant® MTT assay kit. Results are expressed as a percentage of viable cell number in treated cells compared with that of untreated control cells. Data are means ± S.D., n = 8 in each group. (C) Cells were incubated with the stated concentrations of POVPC and PGPC for 4 h. The cells were analyzed for Alexa Fluor<sub>488</sub>-annexin V and propidium iodide fluorescence staining by Flow cytometry as described under “Experimental Procedures”. Results are represented as means ± S.D. Probabilities compared to control were determined by Student’s t-test (two-tailed, unpaired); ***p < 0.001,(n = 8 in each group).</p
TAG-species in rat liver.
<p>Values given are means±s.d. of the sums of the different chain lengths (C46–C60) on the left and degree of desaturation (sat – deca) on the right. Serum data is shown for day 14 and day 20; the control group is shown as black bar, the fructose-fed group is shown as red bar and the FF treated group is shown as a blue bar; significant changes are indicated using *: P<0.05; **: P<0.01; ***: P<0,001.</p