Citrulline does not prevent skeletal muscle wasting or weakness in limb-casted mice.
BACKGROUND
Increasing arginine (Arg) availability reduces atrophy in cultured skeletal muscle cells. Supplementation with its metabolic precursor citrulline (Cit) is more effective at improving skeletal muscle Arg concentrations.


OBJECTIVE
We tested the hypothesis that Cit supplementation would attenuate skeletal muscle atrophy and loss of function during hindlimb immobilization in mice.


METHODS
Male C57BL/6JArc mice underwent 14 d of unilateral hindlimb immobilization/plaster casting and were supplemented with ~0.81 g Cit · kg⁻¹ · d⁻¹ (CIT group) or Ala (ALA group) mixed into their food. The uncasted contralateral limb (internal control) and an uncasted group (CON) served as controls. Muscle atrophy was evaluated with mass, fiber area, and in situ muscle function.


RESULTS
Tibialis anterior (TA) muscle mass [ALA: 37.6 ± 0.92 mg; CIT: 38.3 ± 1.25 mg] and peak tetanic force (ALA: 1150 ± 38.5 mN; CIT: 1150 ± 52.0 mN) were lower (P < 0.001) in the ALA (53.9 ± 0.42 mg) and CIT (1760 ± 28.5 mN) groups than in the CON group. No difference was found between ALA and CIT groups for TA mass, fiber area, or peak force. The mRNA expression of the nitric oxide synthase 2, inducible (Nos2; ~15-fold) and B-cell chronic lymphoid leukemia/lymphoma 2/adenovirus E1B 19 kDa interacting protein 3 (Bnip3; ~17-fold) genes and the ratio of microtubule-associated protein light chain 3BII to 3BI (LC3BII:LC3BI) (50.5% ± 17.7%) were higher (P < 0.05) in the ALA group than in the CON group, suggesting increased autophagy. In the CIT group, Bnip3 mRNA was lower (-70%; P < 0.05) and Nos2 mRNA tended to be lower (-45%; P = 0.05) than in the ALA group, whereas LC3BII:LC3BI was not different from the CON group.


CONCLUSIONS
Cit treatment of male mice did not affect therapeutically relevant outcome measures such as skeletal muscle mass and peak muscle force after 14 d of hindlimb immobilization.
Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Australia.
Daniel J
DJ
Ham
Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Australia.
Tahnee L
TL
Kennedy
Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Australia.
Marissa K
MK
Caldow
Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Australia.
Annabel
A
Chee
Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Australia.
Gordon S
GS
Lynch
Basic and Clinical Myology Laboratory, Department of Physiology, The University of Melbourne, Parkville, Australia rkoopman@unimelb.edu.au.
René
R
Koopman
D000818:Animals; D001343:Autophagy; D015415:Biomarkers / Q000378:metabolism; D002956:Citrulline / Q000378:metabolism / Q000627:therapeutic use*; D019587:Dietary Supplements; D004195:Disease Models, Animal; D005592:Fracture Fixation / Q000009:adverse effects; D005786:Gene Expression Regulation; D006614:Hindlimb; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008810:Mice, Inbred C57BL; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D018908:Muscle Weakness / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology / Q000517:prevention & control*; D018482:Muscle, Skeletal / Q000378:metabolism / Q000473:pathology* / Q000503:physiopathology; D009133:Muscular Atrophy / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology / Q000517:prevention & control*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012149:Restraint, Physical / Q000009:adverse effects
C508686:BNip3 protein, mouse; D015415:Biomarkers; C480860:Map1lc3b protein, mouse; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D024101:Mitochondrial Proteins; D009124:Muscle Proteins; D020033:Protein Isoforms; D002956:Citrulline; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse
amino acids; autophagy; disuse; immobilization; muscle atrophy; muscle breakdown; muscle force