Potential roles of neuronal nitric oxide synthase and the PTEN-induced kinase 1 (PINK1)/Parkin pathway for mitochondrial protein degradation in disuse-induced soleus muscle atrophy in adult rats

Abstract

Excessive nitric oxide (NO) production and mitochondrial dysfunction can activate protein degradation in disuse-induced skeletal muscle atrophy. However, the increase in NO production in atrophied muscles remains controversial. In addition, although several studies have investigated the PTEN-induced kinase 1 (PINK1)/Parkin pathway, a mitophagy pathway, in atrophied muscle, the involvement of this pathway in soleus muscle atrophy is unclear. In this study, we investigated the involvement of neuronal nitric oxide synthase (nNOS) and the PINK1/Parkin pathway in soleus muscle atrophy induced by 14 days of hindlimb unloading (HU) in adult rats. HU lowered the weight of the soleus muscles. nNOS expression showed an increase in atrophied soleus muscles. Although HU increased malondialdehyde as oxidative modification of the protein, it decreased 6-nitrotryptophan, a marker of protein nitration. Additionally, the nitrosocysteine content and S-nitrosylated Parkin were not altered, suggesting the absence of excessive nitrosative stress after HU. The expression of PINK1 and Parkin was also unchanged, whereas the expression of heat shock protein 70 (HSP70), which is required for Parkin activity, was reduced in atrophied soleus muscles. Moreover, we observed accumulation and reduced ubiquitination of high molecular weight mitofusin 2, which is a target of Parkin, in atrophied soleus muscles. These results indicate that excessive NO is not produced in atrophied soleus muscles despite nNOS accumulation, suggesting that excessive NO dose not mediate in soleus muscle atrophy at least after 14 days of HU. Furthermore, the PINK1/Parkin pathway may not play a role in mitophagy at this time point. In contrast, the activity of Parkin may be downregulated because of reduced HSP70 expression, which may contribute to attenuated degradation of target proteins in the atrophied soleus muscles after 14 days of HU. The present study provides new insights into the roles of nNOS and a protein degradation pathway in soleus muscle atrophy.