## 4 Discussion
FOXO03a is a transcription factor that induces the expression of Atrogin-1 and Murf1, ubiquitin ligases important in muscle atrophy, and is inactivated by phosphorylation [27]. Therefore, it would be necessary to examine activity of PGC-1\(\alpha\) in order to clarify the molecular mechanism of metabolic protein expression changes induced by glucose concentration. Skeletal muscle fibers are composed of slow muscle fibers, which are rich in mitochondria and predominantly use an oxidative metabolism, and fast muscle fibers, which predominantly use glycolysis. Under conditions of reduced mechanical load, such as when bedridden or in space, it is known that slow muscle fibers are dominantly degraded accompanied with impaired oxidative metabolism [28]. Therefore, it is suggested that intracellular energy metabolism and myofiber degradation are tightly related. The metabolic reprogramming by glucose concentration could counteract unloading-induced muscle atrophy. Actually, in this study, we observed increased _Myth_ mRNA expression and proteins. It would be interesting to examine protective effect of Low glucose condition on muscle unloading stress in future study. Glucose induces nonenzymatic production of advanced glycation end products (AGEs) from proteins. AGEs are involved with senescence, diabetes, cardiovascular diseases, neurological disorders and cancers [29]. AGE's action is mainly mediated by the binding to receptor of AGE (RAGES), which activates various signaling pathways as the mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinases 1 and 2 (ERK), p21\({}^{\text{ns}}\), p38, and Janus kinase [29]. In skeletal muscles, AGEs accumulate on skeletal muscles' extracellular matrix (ECM), which increases stiffness and activates AGE-RAGE signaling in satellite cells and muscle fibers [30]. Recently, histone glycation has been reported as an epigenetic modification, which might affect the transcriptional regulation [31; 32]. The underlying mechanism of glucose condition in this study might be related with AGE-RAGE signaling and/or epigenetic regulation by glycation of histone proteins. In the present study, we found that the aerobic metabolism of C2C12 myocytes was activated by decreasing the glucose concentration in the culture media. We also identified the molecules responsible for these phenomena by proteomic analysis and found HK2, PFKP, FABP3, and NDUFA11 as targets. Now, we need to investigate whether these molecules play a key role in intracellular metabolism in cell/animal models in which the expression of individual molecules is regulated or overexpressed. The following supporting information can be downloaded at: [[https://www.mdpi.com/article/10.3390/app12031553/s1](https://www.mdpi.com/article/10.3390/app12031553/s1)]([https://www.mdpi.com/article/10.3390/app12031553/s1](https://www.mdpi.com/article/10.3390/app12031553/s1)), Table S1: The C2C12 myotubes at different glucose concentrations in the culture media after 5 days post-differentiation. Conceptualization, T.U. ; formal analysis, A.K. and T.F. ([PERSON]); investigation, A.K. ; data curation, I.S. ; writing--original draft preparation, A.K. ; writing--review and editing, I.S., T.U., K.S. and T.F ([PERSON]); supervision, T.U. ; project administration, T.U. ; funding acquisition, T.U. and T.N. All authors have read and agreed to the published version of the manuscript. This research was funded by the 3 rd Joint research to explore new research themes, Nutteruciticals Division, Otsuka Pharmaceutical Co, Ltd., Saga, Japan, and Japan Society for the Promotion of Science (JSPS) KAKENHI Grants (20K19639 and 19 KK0253). **Acknowledgments:** IPA analysis was supported by Support Center for Advanced Medical Sciences, Tokushima University Graduate School of Biomedical Sciences. This research was also supported by Saga Nutraceuticalulicals Research Institute, Otsuka Pharmaceutical Co, Ltd., Saga, Japan. **Conflicts of Interest:** The authors declare no conflict of interest.