title: Fatty acid-induced defects in insulin signalling, in myotubes derived from children, are related to ceramide production from palmitate rather than the accumulation of intramyocellular lipid.
abstract: The elevation of free fatty acids (FFAs), observed in childhood obesity results in intramyocellular lipid (IMCL) accumulation with consequent insulin resistance. Using in vitro differentiated myotubes from normal weight pre-pubertal children (n = 8), we examined the effects of saturated (palmitate) and unsaturated (oleate) FFAs on insulin-stimulated AKT phosphorylation (pAKT) and IMCL accumulation. Palmitate decreased pAKT (Mean [SEM] % change pAKT with palmitate 750 microM vs. control; pThr308 site -50.5% [28.7] and pSer473 site -38.7% [11.7]; P < 0.001) with no effect on IMCL formation. Equimolar bromopalmitate did not effect pAKT and blocking ceramide production abolished the palmitate-induced reduction in signalling, suggesting that ceramide synthesis is critical for palmitate's actions. Oleate did not effect pAKT (1,000 microM oleate; pSer473 site -3.4% [11.4]; P = NS) but increased IMCL accumulation (+32.3% [7.1%]; P < 0.001). Co-administration of oleate diminished the reduction in pAKT seen with palmitate (+36.4% [23.6] vs. -13.3% [13.6]; P = 0.28), with similar IMCL levels to oleate alone. Co-administration also caused a significant reduction in 14C-ceramide synthesis from 14C-palmitate (101.6 [21.6] vs. 371.5 [122.4] DPM/mg protein; P < 0.001). In summary, palmitate appears to cause insulin resistance in children's myotubes via its metabolism to ceramide, and this process appears unrelated to IMCL formation and is ameliorated by oleate.
authors.affiliation: Institute of Child Life and Health and Clinical Science at North Bristol, The University of Bristol and Royal Hospital for Children, Bristol, United Kingdom.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Sabin
authors.forename: Claire E H
authors.initials: CE
authors.lastname: Stewart
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Crowne
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Turner
authors.forename: Linda P
authors.initials: LP
authors.lastname: Hunt
authors.forename: Gavin I
authors.initials: GI
authors.lastname: Welsh
authors.forename: Malcolm J
authors.initials: MJ
authors.lastname: Grohmann
authors.forename: Jeff M P
authors.initials: JM
authors.lastname: Holly
authors.forename: Julian P H
authors.initials: JP
authors.lastname: Shield
mesh_terms: D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D002518:Ceramides / Q000096:biosynthesis*; D002648:Child; D002675:Child, Preschool; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D007223:Infant; D007328:Insulin / Q000378:metabolism* / Q000494:pharmacology; D050356:Lipid Metabolism / Q000187:drug effects; D008297:Male; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000378:metabolism*; D019301:Oleic Acid / Q000494:pharmacology; D010078:Oxazines; D010168:Palmitates / Q000378:metabolism* / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D010768:Phosphoserine / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
chemical_list: D002518:Ceramides; D007328:Insulin; D010078:Oxazines; D010168:Palmitates; D010750:Phosphoproteins; D010768:Phosphoserine; D019301:Oleic Acid; D051057:Proto-Oncogene Proteins c-akt; C044808:nile red