Title

 

Authors

J.-F. Desaphy¹, S. Pierno¹, A. Liantonio¹, M. De Bellis¹, G.M. Camerino¹, M.M. Dinardo¹, G. Gramegna¹, A. Scaramuzzi¹, D. Conte Camerino^1
 
1Section of Pharmacology, Dept. of Pharmacobiology, University of Bari Aldo Moro, Bari, Italy

 

Abstract

In the hindlimb-unloaded (HU) mouse, muscle disuse induces a severe atrophy and a phenotypic shift of antigravity muscles, such as the soleus muscle, which may impede postural maintenance and movement upon return to normal condition (Desaphy et al., 2010). This model was first developed to mimic the effects of microgravity exposure during spaceflight, but is now considered as well as a model of muscle disuse associated with aging or bed rest. No countermeasure is available so far. In the present study, we wondered whether a treatment with the anabolic androgenic steroid, nandrolone decanoate (ND), may prevent muscle wasting induced by HU. Mice (C57BL, male, 6-month-old) were divided in four experimental groups: CTR (control mice), CHU (mice hindlimb-unloaded for 14 days), VHU (hindlimb-unloaded mice receiving vehicle), and NHU (hindlimb-unloaded mice receiving 5 mg/kg/day ND s.c. for 4 weeks and hindlimb-unloaded during the last 14 days). After HU, soleus muscle weight was significantly reduced by about 25 % in CHU and VHU with respect to CTR mice, whereas the reduction was only 10 % in NHU mice. The protein content of whole soleus muscle, measured with a commercial kit,  followed the same direction, suggesting that ND was able to counteract muscle atrophy. In accord with the slow-to-fast transition of soleus muscle phenotype, the resting chloride conductance of sarcolemma (gCl), measured in current-clamp mode with endocellular microlectrodes, was increased by 45 % in CHU compared to CTR mice. No effect of vehicle or ND on the gCl was observed in VHU and NHU groups, which showed a gCl similar to CHU mice. The resting cytosolic calcium ion concentration (restCa) in soleus muscle fibers, measured with FURA-2 cytofluorometric technique, was reduced by 7 % in CHU compared to CTR mice, as expected from the phenotypic shift. The reduction of restCa was about 20 % in VHU and NHU mice compared to CTR group. The level of expression of various genes involved in muscle atrophy was assayed by realtime PCR. Expression of MuRF-1 and atrogin-1, two muscle-specific ubiquitin ligases involved in proteasome activation, was increased in CHU soleus muscle compared to CTR mice (significant only for MurF-1). Neither vehicle nor ND had any effect on these atrogenes in VHU and NHU mice. Expression of cathepsin-L and LC3-β, two markers of autophagy, was unchanged in CHU, VHU, and NHU soleus muscles compared to CTR mice. Expression of PGC1α , a master gene for mitochondria biogenesis and function that is phenotypically more expressed in slow, oxidative muscles compared to fast, glycolitic muscles, was significantly reduced in CHU compared to CTR mice, and remained unchanged in VHU and NHU compared to CHU mice. Overall, the results suggest that ND treatment was not able to counteract the disuse-associated phenotypic change of soleus muscle, since the gCl, restCa, and PGC1α  expression were not affected by the drug. Interestingly, ND treatment was however able to contrast the drop of muscle mass and protein content, probably through the stimulation of protein synthesis, since atrogene expression remained unchanged. A treatment combining ND with a drug able to counteract phenotypic change may represent a valuable option for treatment of muscle wasting in various ill conditions. This study was supported by the Italian Space Agency and Galapagos SASU (France)