Purpose: It is known that macroautophagy plays a pivotal role in cell differentia- tion and function. We recently showed that MR antagonism reduced autophagy rate and induced browning of adipose tissue. Interestingly, a critical role of autophagy has been recently shown also in skeletal muscle development and it is known that MR antagonism is able to improve insulin-stimulated glucose transport in rat skele- tal muscle. Therefore we investigated a possible involvement of MR in regulating skeletal muscle differentiation and function in vitro and in vivo. Methods: We investigated the involvement of MR in regulating macroautophagy in murine skeletal muscle cells (C2C12) and we started to characterize the impact of adipose MR activation on skeletal muscle metabolic profile and development in a novel adipose-specific MR Knockout (MRKO) mouse model. Results: We first characterised the ontogenesis of MR in a murine myoblast cell line (C2C12) by RT-PCR analysis in order to evaluate the expression of MR during myotube differentiation in vitro. We observed a 2,5 fold-increase in basal MR mRNA expression in skeletal myoblasts compared with myotubes after 72h of differentiation. To evaluate the effects of MR activity on C2C12 cells we treated C2C12 myoblasts and myotubes with aldosterone (Aldo, 10-8 mol/L) for 24h and observed a marked increase in LC3 mRNA expression suggesting an increased autophagic rate. This effect was reverted by treatment with MR antagonist spironolactone (Spiro, 10-5 mol/L). However, protein levels of LC3 were not affected by any treatment, suggesting that longer treatments could be necessary to reveal an effect. Moreover we observed that Aldo significantly reduced in Heavy Chain Myosin (MHC) mRNA levels, suggesting that MR activity represses muscle cell differentiation. Such effect was also MR dependent, given that Spiro was able to revert such effect. We also analyzed skeletal muscle insulin sensitivity in vivo in MRKO mice evalu- ating soleus muscle insulin receptor substrate-1, tyrosine phosphorylated IRS-1, GLUT4 levels mRNA expression and Akt phosphorylation. Interestingly, we observed improved soleus muscle insulin signalling parameters and systemic insulin sensitivity in MRKO mice compared to controls. Further studies are necessary to explore the effect of MR activation on skeletal muscle development in vivo. Conclusions: These data reveal a potential role of MR in modulating skeletal mus- cle differentiation, autophagic rate and insulin sensitivity, which suggest a potential application of MR antagonists to improve skeletal muscle function.
Role of mineralocorticoid receptor in the regulation of skeletal muscle differentiation and function
Ferraro E;
2015-01-01
Abstract
Purpose: It is known that macroautophagy plays a pivotal role in cell differentia- tion and function. We recently showed that MR antagonism reduced autophagy rate and induced browning of adipose tissue. Interestingly, a critical role of autophagy has been recently shown also in skeletal muscle development and it is known that MR antagonism is able to improve insulin-stimulated glucose transport in rat skele- tal muscle. Therefore we investigated a possible involvement of MR in regulating skeletal muscle differentiation and function in vitro and in vivo. Methods: We investigated the involvement of MR in regulating macroautophagy in murine skeletal muscle cells (C2C12) and we started to characterize the impact of adipose MR activation on skeletal muscle metabolic profile and development in a novel adipose-specific MR Knockout (MRKO) mouse model. Results: We first characterised the ontogenesis of MR in a murine myoblast cell line (C2C12) by RT-PCR analysis in order to evaluate the expression of MR during myotube differentiation in vitro. We observed a 2,5 fold-increase in basal MR mRNA expression in skeletal myoblasts compared with myotubes after 72h of differentiation. To evaluate the effects of MR activity on C2C12 cells we treated C2C12 myoblasts and myotubes with aldosterone (Aldo, 10-8 mol/L) for 24h and observed a marked increase in LC3 mRNA expression suggesting an increased autophagic rate. This effect was reverted by treatment with MR antagonist spironolactone (Spiro, 10-5 mol/L). However, protein levels of LC3 were not affected by any treatment, suggesting that longer treatments could be necessary to reveal an effect. Moreover we observed that Aldo significantly reduced in Heavy Chain Myosin (MHC) mRNA levels, suggesting that MR activity represses muscle cell differentiation. Such effect was also MR dependent, given that Spiro was able to revert such effect. We also analyzed skeletal muscle insulin sensitivity in vivo in MRKO mice evalu- ating soleus muscle insulin receptor substrate-1, tyrosine phosphorylated IRS-1, GLUT4 levels mRNA expression and Akt phosphorylation. Interestingly, we observed improved soleus muscle insulin signalling parameters and systemic insulin sensitivity in MRKO mice compared to controls. Further studies are necessary to explore the effect of MR activation on skeletal muscle development in vivo. Conclusions: These data reveal a potential role of MR in modulating skeletal mus- cle differentiation, autophagic rate and insulin sensitivity, which suggest a potential application of MR antagonists to improve skeletal muscle function.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
