title: L-Citrulline Protects Skeletal Muscle Cells from Cachectic Stimuli through an iNOS-Dependent Mechanism.
abstract: Dietary L-citrulline is thought to modulate muscle protein turnover by increasing L-arginine availability. To date, the direct effects of increased L-citrulline concentrations in muscle have been completely neglected. Therefore, we determined the role of L-citrulline in regulating cell size during catabolic conditions by depriving mature C2C12 myotubes of growth factors (serum free; SF) or growth factors and nutrients (HEPES buffered saline; HBS). Cells were treated with L-citrulline or equimolar concentrations of L-arginine (positive control) or L-alanine (negative control) and changes in cell size and protein turnover were assessed. In myotubes incubated in HBS or SF media, L-citrulline improved rates of protein synthesis (HBS: +63%, SF: +37%) and myotube diameter (HBS: +18%, SF: +29%). L-citrulline treatment substantially increased iNOS mRNA expression (SF: 350%, HBS: 750%). The general NOS inhibitor L-NAME and the iNOS specific inhibitor aminoguanidine prevented these effects in both models. Depriving myotubes in SF media of L-arginine or L-leucine, exacerbated wasting which was not attenuated by L-citrulline. The increased iNOS mRNA expression was temporally associated with increases in mRNA of the endogenous antioxidants SOD1, SOD3 and catalase. Furthermore, L-citrulline prevented inflammation (LPS) and oxidative stress (H2O2) induced muscle cell wasting. In conclusion, we demonstrate a novel direct protective effect of L-citrulline on skeletal muscle cell size independent of L-arginine that is mediated through induction of the inducible NOS (iNOS) isoform. This discovery of a nutritional modulator of iNOS mRNA expression in skeletal muscle cells could have substantial implications for the treatment of muscle wasting conditions.
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Ham
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Benjamin G
authors.initials: BG
authors.lastname: Gleeson
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Annabel
authors.initials: A
authors.lastname: Chee
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Dale M
authors.initials: DM
authors.lastname: Baum
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Marissa K
authors.initials: MK
authors.lastname: Caldow
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: Gordon S
authors.initials: GS
authors.lastname: Lynch
authors.affiliation: Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Victoria, Australia.
authors.forename: René
authors.initials: R
authors.lastname: Koopman
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D002374:Catalase / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002956:Citrulline / Q000494:pharmacology*; D051379:Mice; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism; D000072105:Superoxide Dismutase-1
chemical_list: D000975:Antioxidants; D012333:RNA, Messenger; D002956:Citrulline; D002374:Catalase; D052247:Nitric Oxide Synthase Type II; C000606291:Sod1 protein, mouse; C499090:Sod3 protein, mouse; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1