title: Effects of salmeterol on skeletal muscle cells:  metabolic and proapoptotic features.
abstract: OBJECTIVE
  Salmeterol is a β2-adrenergic receptor agonist widely used for the treatment of asthma and chronic obstructive pulmonary disease. It has been shown that salmeterol is also used at supratherapeutic doses as performance-enhancing substance in sport practice. Although the abuse of β-agonists might determine some adverse effects, the molecular effects of salmeterol on skeletal muscle cells remain unclear.
  
  
  METHODS
  We evaluated the effects of salmeterol (0.1-10 μM) on both proliferative and differentiated rat L6C5 and mouse C2C12 skeletal muscle cell lines. The metabolic effects were evaluated by glyceraldehyde phosphate dehydrogenase, lactate dehydrogenase, citrate synthase, 3-OH acyl-CoA dehydrogenase, and alanine transglutaminase activities. Cytotoxic and apoptotic effects were analyzed by 3-(4,5-dimethylthiazol-1)-5-(3-carboxymeth-oxyphenyl)-2H-tetrazolium, trypan blue exclusion assay, terminal deoxynucleotidyl transferase dUTP nick end labeling assay, Western blot analysis, and immunofluorescence staining.
  
  
  RESULTS
  We showed that salmeterol reduced the growth rate of proliferating cells in a dose- and time-dependent manner (6-48 h). An increase in oxidative metabolism was found after 6 h in C2C12 and L6C5 myoblasts and in C2C12 myotubes with respect to control cells, while in L6C5 myotubes, anaerobic metabolism prevailed. Exposure of myoblasts and myotubes for 48 and 72 h at high salmeterol concentrations induced apoptosis by the activation of the intrinsic apoptotic pathway, as confirmed by the modulation of the apoptotic proteins Bcl-xL, caspase-9, and poly (ADP-ribose) polymerase and by the cytoplasmic release of Smac/DIABLO.
  
  
  CONCLUSIONS
  Altogether, our results demonstrate that short-term supratherapeutic salmeterol exposure increased oxidative metabolic pathways on skeletal muscle cells, whereas prolonged treatment inhibits cell growth and exerts either a cytostatic or a proapoptotic effect in a time- and dose-dependent way.
authors.affiliation: Department of Health Sciences, University of Rome Foro Italico, Rome, Italy. guglielmo.duranti@uniroma4.it
authors.forename: Guglielmo
authors.initials: G
authors.lastname: Duranti
authors.forename: Piergiorgio
authors.initials: P
authors.lastname: La Rosa
authors.forename: Ivan
authors.initials: I
authors.lastname: Dimauro
authors.forename: Francesca
authors.initials: F
authors.lastname: Wannenes
authors.forename: Sergio
authors.initials: S
authors.lastname: Bonini
authors.forename: Stefania
authors.initials: S
authors.lastname: Sabatini
authors.forename: Paolo
authors.initials: P
authors.lastname: Parisi
authors.forename: Daniela
authors.initials: D
authors.lastname: Caporossi
mesh_terms: D015094:3-Hydroxyacyl CoA Dehydrogenases / Q000378:metabolism; D058666:Adrenergic beta-2 Receptor Agonists / Q000494:pharmacology*; D000420:Albuterol / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D053453:Caspase 9 / Q000378:metabolism; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002950:Citrate (si)-Synthase / Q000378:metabolism; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D051379:Mice; D018482:Muscle, Skeletal / Q000187:drug effects* / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051381:Rats; D000068299:Salmeterol Xinafoate; D011503:Transglutaminases / Q000378:metabolism; D051020:bcl-X Protein / Q000378:metabolism
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D051020:bcl-X Protein; D000068299:Salmeterol Xinafoate; D015094:3-Hydroxyacyl CoA Dehydrogenases; D007770:L-Lactate Dehydrogenase; D011503:Transglutaminases; D002950:Citrate (si)-Synthase; D011065:Poly(ADP-ribose) Polymerases; C055686:glyceraldehyde 3-phosphate phosphatase; D010744:Phosphoric Monoester Hydrolases; D053453:Caspase 9; D000420:Albuterol; D010100:Oxygen