H-silicene nanosheets as a novel therapeutic approach for disuse muscle atrophy by modulating macrophage polarization.

UNLABELLED: Disuse muscle atrophy (DMA) is characterized by progressive loss of muscle mass and strength, often accompanied by inflammation and macrophage imbalance. Here, we introduce hydrogenated silicene nanosheets (H-silicene) as a novel nanotherapeutic strategy to mitigate DMA through modulating macrophage polarization. H-silicene exhibited good biocompatibility and sustained hydrogen release. In vitro, H-silicene suppressed LPS-induced M1 macrophage activation while promoting M2 polarization, and alleviated myotube atrophy in co-culture assays. In a murine immobilization model, intramuscular H-silicene administration significantly mitigated muscle wasting, reduced fibrosis, and improved functional outcomes. Immunofluorescence staining confirmed a decrease in iNOS+/TNF-α+ cells and an elevated CD206+/IL-10+ populations in treated muscle. Integrated transcriptomic and proteomic analyses revealed H-silicene-mediated modulation of pathways related to inflammation, oxidative stress, and myogenesis. These findings highlight H-silicene as a promising immunoregulatory nanomaterial for the treatment of disuse-induced muscle atrophy.

METHODS

H-silicene nanosheets were synthesized from CaSi2 under argon-protected acid treatment and probe-sonicated to obtain dispersible nanosheets. Cytocompatibility was assessed in RAW264.7 and C2C12 cells. Macrophage polarization was analyzed by flow cytometry, ELISA, and immunofluorescence after LPS stimulation ± H-silicene. Conditioned media were applied to differentiated C2C12 myotubes to assess paracrine effects. In vivo, a mouse hindlimb immobilization model was used to induce DMA. Mice received intramuscular injections of H-silicene (250 ppm). Muscle tissue was analyzed by histology, immunofluorescence, behavioral assays, and RNA-seq and proteomics.

RESULTS

H-silicene exhibited low cytotoxicity and dose-dependently suppressed LPS-induced M1 polarization while enhancing M2 polarization. It reduced proinflammatory cytokines and preserved C2C12 myotube morphology in co-culture models. In vivo, H-silicene improved muscle fiber area, reduced collagen deposition, restored grip strength , and improved rotarod performance in DMA mice. Immunostaining confirmed reduced iNOS+/TNF-α+ and increased CD206+/IL-10+ macrophages. Multi-omics analysis revealed regulation of inflammatory and regenerative signaling, including NF-κB, IL-17, and myoblast differentiation pathways.

CONCLUSION

This study demonstrates that H-silicene alleviates disuse-induced muscle atrophy by promoting the transition from pro-inflammatory M1 macrophages to anti-inflammatory M2 macrophages and remodeling the local immune microenvironment, making it a promising nanotherapeutic for muscle-wasting disorders.