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Updated: Jul 28, 2025
In Situ Immunofluorescent Staining of Autophagy in Muscle Stem Cells
Published on: June 12, 2017
iPSCs ameliorate hypoxia-induced autophagy and atrophy in C2C12 myotubes via the AMPK/ULK1 pathway
Haimei Cen1, Pin Fan1, Yuting Ding1
1Department of Neurology, The First Affiliated Hospital, Jinan University, Guangzhou, 510630, Guangdong, China.
Background:
Duchenne muscular dystrophy (DMD) is an X-linked lethal genetic disorder for which there is no effective treatment. Previous studies have shown that stem cell transplantation into mdx mice can promote muscle regeneration and improve muscle function, however, the specific molecular mechanisms remain unclear. DMD suffers varying degrees of hypoxic damage during disease progression. This study aimed to investigate whether induced pluripotent stem cells (iPSCs) have protective effects against hypoxia-induced skeletal muscle injury.
Results:
In this study, we co-cultured iPSCs with C2C12 myoblasts using a Transwell nested system and placed them in a DG250 anaerobic workstation for oxygen deprivation for 24 h. We found that iPSCs reduced the levels of lactate dehydrogenase and reactive oxygen species and downregulated the mRNA and protein levels of BAX/BCL2 and LC3II/LC3I in hypoxia-induced C2C12 myoblasts. Meanwhile, iPSCs decreased the mRNA and protein levels of atrogin-1 and MuRF-1 and increased myotube width. Furthermore, iPSCs downregulated the phosphorylation of AMPKα and ULK1 in C2C12 myotubes exposed to hypoxic damage.
Conclusions:
Our study showed that iPSCs enhanced the resistance of C2C12 myoblasts to hypoxia and inhibited apoptosis and autophagy in the presence of oxidative stress. Further, iPSCs improved hypoxia-induced autophagy and atrophy of C2C12 myotubes through the AMPK/ULK1 pathway. This study may provide a new theoretical basis for the treatment of muscular dystrophy in stem cells.
Insights
Induced pluripotent stem cells (iPSCs) protect skeletal muscle cells from hypoxia-induced damage. iPSCs reduce cell death and atrophy by regulating key molecular pathways, offering potential for muscular dystrophy treatment.
Area of Science:
- Biomedical Science
- Regenerative Medicine
- Molecular Biology
Background:
- Duchenne muscular dystrophy (DMD) is a severe genetic disorder with no cure.
- Hypoxic damage is a significant factor in DMD progression.
- Stem cell therapy shows promise, but mechanisms require clarification.
Purpose of the Study:
- To investigate the protective effects of induced pluripotent stem cells (iPSCs) against hypoxia-induced skeletal muscle injury.
- To elucidate the molecular mechanisms underlying iPSC-mediated protection.
Main Methods:
- Co-culture of iPSCs with C2C12 myoblasts under hypoxic conditions using a Transwell system.
- Assessment of cell viability markers (lactate dehydrogenase, reactive oxygen species).
- Analysis of apoptosis (BAX/BCL2), autophagy (LC3II/LC3I), and muscle atrophy (atrogin-1, MuRF-1) related gene and protein expression.
- Evaluation of AMPK/ULK1 pathway activation.
Main Results:
- iPSCs reduced hypoxia-induced cell death and reactive oxygen species.
- iPSCs downregulated pro-apoptotic and autophagic markers.
- iPSCs inhibited muscle atrophy markers and increased myotube width.
- iPSCs suppressed AMPKα and ULK1 phosphorylation.
Conclusions:
- iPSCs enhance skeletal muscle cell resistance to hypoxia and oxidative stress.
- iPSCs mitigate hypoxia-induced apoptosis and autophagy via the AMPK/ULK1 pathway.
- This study provides a foundation for stem cell-based therapeutic strategies for muscular dystrophy.
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