Author(s): KATO, Y., TOKINOYA, K., AOKI, K., TAKEKOSHI, K., Institution: UNIVERSITY OF TSUKUBA, Country: JAPAN, Abstract-ID: 1748

INTRODUCTION:
Skeletal muscle regeneration is mediated by skeletal muscle stem cells, known as satellite cells. Satellite cell proliferation is regulated by various secreted factors, including exercise-induced cytokines. Renalase is a secreted protein that activates intracellular signaling pathways, including p38 MAPK, ERK1/2, and Akt, via its receptor, plasma membrane Ca²⁺-ATPase 4b (PMCA4b). In tumor cells, increased renalase expression promotes cell proliferation through activation of these pathways. Exercise also increases circulating renalase levels as well as its expression in skeletal muscle. However, the role of renalase in skeletal muscle and satellite cells remains unclear. This study aimed to elucidate the role of renalase in satellite cell proliferation.
METHODS:
Satellite cells were isolated from the extensor digitorum longus (EDL) muscles of 8–12-week-old male C57BL/6J mice and cultured for 1 week. For renalase knockdown, the satellite cells were seeded in 12- or 24-well plates and infected with adenoviruses expressing short hairpin RNA (shRNA) at a multiplicity of infection (MOI) of 50. After 12 h, the medium was replaced with fresh growth medium (designated as day 0), and the cells were cultured 3 or 4 days. Cell numbers were quantified by DAPI staining from day 0 to day 4. On day 3, immunostaining, quantitative PCR (qPCR), and Western blotting were performed to assess proliferation-related markers.
RESULTS:
Renalase mRNA expression was reduced by approximately 70% on day 3. Renalase knockdown significantly suppressed cell proliferation, particularly reducing the Pax7⁺/MyoD⁺ cell population. Consistent with these findings, the numbers of Ki67-positive and EdU-positive cells were significantly decreased, accompanied by reduced expression of genes involved in cell-cycle progression. Furthermore, phosphorylation of ERK1/2 and Akt, key regulators of cell proliferation, was significantly reduced following renalase knockdown. In addition, phosphorylation of Akt downstream targets, including mTOR, p70 S6 kinase (p70S6K), ribosomal protein S6, and eukaryotic translation initiation factor 4E–binding protein 1 (4E-BP1), was significantly reduced.
CONCLUSION:
These results indicate that renalase is a critical regulator of satellite cell proliferation through activation of the Akt–mTOR and ERK1/2 signaling pathways. Considering the central role of satellite cells in skeletal muscle regeneration, reduced renalase expression may impair satellite cell proliferation, ultimately leading to defective muscle regeneration.