Leucine supplementation improves skeletal muscle regeneration after cryolesion in rats

This study was undertaken in order to provide further insight into the role of leucine supplementation in the skeletal muscle regeneration process, focusing on myofiber size and strength recovery. Young (2-month-old) rats were subjected or not to leucine supplementation (1.35 g/kg per day) started 3 days prior to cryolesion. Then, soleus muscles were cryolesioned and continued receiving leucine supplementation until 1, 3 and 10 days later. Soleus muscles from leucine-supplemented animals displayed an increase in myofiber size and a reduction in collagen type III expression on post-cryolesion day 10. Leucine was also effective in reducing FOXO3a activation and ubiquitinated protein accumulation in muscles at post-cryolesion days 3 and 10. In addition, leucine supplementation minimized the cryolesion-induced decrease in tetanic strength and increase in fatigue in regenerating muscles at post-cryolesion day 10. These beneficial effects of leucine were not accompanied by activation of any elements of the phosphoinositide 3-kinase/Akt/mechanistic target of rapamycin signalling pathway in the regenerating muscles. Our results show that leucine improves myofiber size gain and strength recovery in regenerating soleus muscles through attenuation of protein ubiquitination. In addition, leucine might have therapeutic effects for muscle recovery following injury and in some muscle diseases.FAPESP, 2010/52520-0FAPESP, 2012/15276-

Leucine and HMB differentially modulate proteasome system in skeletal muscle under different sarcopenic conditions

In the present study we have compared the effects of leucine supplementation and its metabolite β-hydroxy-β-methyl butyrate (HMB) on the ubiquitin-proteasome system and the PI3K/Akt pathway during two distinct atrophic conditions, hindlimb immobilization and dexamethasone treatment. Leucine supplementation was able to minimize the reduction in rat soleus mass driven by immobilization. On the other hand, leucine supplementation was unable to provide protection against soleus mass loss in dexamethasone treated rats. Interestingly, HMB supplementation was unable to provide protection against mass loss in all treatments. While solely fiber type I cross sectional area (CSA) was protected in immobilized soleus of leucine-supplemented rats, none of the fiber types were protected by leucine supplementation in rats under dexamethasone treatment. In addition and in line with muscle mass results, HMB treatment did not attenuate CSA decrease in all fiber types against either immobilization or dexamethasone treatment. While leucine supplementation was able to minimize increased expression of both Mafbx/Atrogin and MuRF1 in immobilized rats, leucine was only able to minimize Mafbx/Atrogin in dexamethasone treated rats. In contrast, HMB was unable to restrain the increase in those atrogenes in immobilized rats, but in dexamethasone treated rats, HMB minimized increased expression of Mafbx/Atrogin. The amount of ubiquitinated proteins, as expected, was increased in immobilized and dexamethasone treated rats and only leucine was able to block this increase in immobilized rats but not in dexamethasone treated rats. Leucine supplementation maintained soleus tetanic peak force in immobilized rats at normal level. On the other hand, HMB treatment failed to maintain tetanic peak force regardless of treatment. The present data suggested that the anti-atrophic effects of leucine are not mediated by its metabolite HMB.State of São Paulo Research Foundation, FAPESP - 07/57613-3State of São Paulo Research Foundation, FAPESP - 12/13071-0State of São Paulo Research Foundation, FAPESP - 06/61523-7CNP

Role of E3 ligases on skeletal muscle plasticity.

Neste estudo analisamos o envolvimento de E3 ligases sob três aspectos da plasticidade muscular esquelética: a perda de massa decorrente do desuso, a manutenção de fibras tipo I e II e a regeneração do tecido muscular. O primeiro objetivo foi determinar se leucina atenuaria a perda de massa provocada pela atrofia. Nossos dados mostraram que este aminoácido evitou a perda de massa, sobretudo através da inibição da expressão de E3 ligases. A seguir, analisamos se a ausência de duas E3 ligases, MuRF1 e MuRF2, poderia alterar a proporção de miofibras tipo I e tipo II. Detectamos que na ausência destas E3 ligases, a identidade de fibras tipo II era perdida, além destas fibras estarem protegidas da atrofia sem MuRF1. Por fim, analisamos o papel de MuRF1 e MuRF2 durante a regeneração. Os resultados mostraram que estas E3 ligases em conjunto são cruciais para a fisiologia das células-satélites e portanto para a regeneração do tecido. Esta tese mostrou que determinadas E3 ligases exercem um papel crucial para a plasticidade muscular.In the present thesis we analyzed the involvement of E3 ligases under three aspects of skeletal muscle plasticity: mass loss resulting from disuse, maintenance of fiber type I and II and regeneration of muscle tissue. Our first aim was to determine whether leucine, was able to attenuate the mass loss caused by wasting. Our results showed that this amino acid prevented the mass loss, mainly by inhibiting the expression of E3 ligases. The second aim was to determine whether the absence of two E3 ligases, MuRF1 and MuRF2, could alter the proportion of type I and type II fibers. We found that in the absence of these E3 ligases, the identity of type II fibers was lost, and these fibers were protected against atrophy in the absence of MuRF1. Lastly, we analyze the role of MuRF1 and MuRF2 in muscle tissue regeneration. The results showed that these E3 ligases together are crucial to satellite-cells physiology and consequently an adequate tissue regeneration. This thesis show that certain E3 ligases can play a crucial role in muscle plasticity