title: Remodeling lipid metabolism and improving insulin responsiveness in human primary myotubes.
abstract: OBJECTIVE
  Disturbances in lipid metabolism are strongly associated with insulin resistance and type 2 diabetes (T2D). We hypothesized that activation of cAMP/PKA and calcium signaling pathways in cultured human myotubes would provide further insight into regulation of lipid storage, lipolysis, lipid oxidation and insulin responsiveness.
  
  
  METHODS
  Human myoblasts were isolated from vastus lateralis, purified, cultured and differentiated into myotubes. All cells were incubated with palmitate during differentiation. Treatment cells were pulsed 1 hour each day with forskolin and ionomycin (PFI) during the final 3 days of differentiation to activate the cAMP/PKA and calcium signaling pathways. Control cells were not pulsed (control). Mitochondrial content, (14)C lipid oxidation and storage were measured, as well as lipolysis and insulin-stimulated glycogen storage. Myotubes were stained for lipids and gene expression measured.
  
  
  RESULTS
  PFI increased oxidation of oleate and palmitate to CO(2) (p<0.001), isoproterenol-stimulated lipolysis (p = 0.01), triacylglycerol (TAG) storage (p<0.05) and mitochondrial DNA copy number (p = 0.01) and related enzyme activities. Candidate gene and microarray analysis revealed increased expression of genes involved in lipolysis, TAG synthesis and mitochondrial biogenesis. PFI increased the organization of lipid droplets along the myofibrillar apparatus. These changes in lipid metabolism were associated with an increase in insulin-mediated glycogen storage (p<0.001).
  
  
  CONCLUSIONS
  Activation of cAMP/PKA and calcium signaling pathways in myotubes induces a remodeling of lipid droplets and functional changes in lipid metabolism. These results provide a novel pharmacological approach to promote lipid metabolism and improve insulin responsiveness in myotubes, which may be of therapeutic importance for obesity and type 2 diabetes.
authors.affiliation: Molecular and Experimental Endocrinology Laboratory, Pennington Biomedical Research Center, Baton Rouge, Louisiana, United States of America.
authors.forename: Lauren M
authors.initials: LM
authors.lastname: Sparks
authors.forename: Cedric
authors.initials: C
authors.lastname: Moro
authors.forename: Barbara
authors.initials: B
authors.lastname: Ukropcova
authors.forename: Sudip
authors.initials: S
authors.lastname: Bajpeyi
authors.forename: Anthony E
authors.initials: AE
authors.lastname: Civitarese
authors.forename: Matthew W
authors.initials: MW
authors.lastname: Hulver
authors.forename: G Hege
authors.initials: GH
authors.lastname: Thoresen
authors.forename: Arild C
authors.initials: AC
authors.lastname: Rustan
authors.forename: Steven R
authors.initials: SR
authors.lastname: Smith
mesh_terms: D000328:Adult; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D005227:Fatty Acids / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D015759:Ionomycin / Q000494:pharmacology; D050356:Lipid Metabolism / Q000187:drug effects* / Q000235:genetics; D008066:Lipolysis / Q000187:drug effects / Q000235:genetics; D008297:Male; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology / Q000235:genetics; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D010084:Oxidation-Reduction / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects
chemical_list: D005227:Fatty Acids; D007328:Insulin; D005576:Colforsin; D015759:Ionomycin; D005947:Glucose