Objective: Severe skeletal muscle contusions result in significant inflammation, fibrotic scarring, and incomplete functional recovery. Betulin, a natural triterpenoid, has demonstrated anti-inflammatory and regenerative properties. This study aimed to investigate the efficacy of betulin in promoting molecular and functional recovery in a murine model of muscle contusion.

Methods: A standardized contusion injury was induced in the tibialis anterior muscle of C57BL/6 mice. Mice were treated daily with either betulin (50 mg/kg, IP) or a vehicle control. Functional recovery was assessed at days 3, 7, 14, and 28 post-injury using in vivo grip strength tests. Muscle tissues were harvested at the same time points for analysis. Histological evaluation was performed using Hematoxylin & Eosin (H&E) for general morphology and inflammation, and Masson’s Trichrome for fibrosis. Myogenic regeneration and key molecular markers of inflammation and fibrosis were quantified using immunohistochemistry and quantitative real-time PCR (qRT-PCR).

Results: Betulin treatment was associated with a significant acceleration in the recovery of muscle strength compared to vehicle-treated controls. Histological analysis revealed that betulin markedly reduced inflammatory cell infiltration at early time points and decreased the total area of fibrotic tissue by day 28. This was correlated with a significant downregulation of pro-inflammatory (Tnf-α, Il-6) and pro-fibrotic (Tgf-β1, Col1a1) gene expression. Furthermore, betulin-treated muscles exhibited an increased number of newly formed, centrally nucleated myofibers. This enhanced myogenesis was supported by the upregulation of myogenic regulatory factors, including Myod1 and Myog, indicating an augmented satellite cell-mediated repair process.

Conclusion: Betulin appears to promote comprehensive recovery following muscle contusion by targeting multiple pathological processes. It is associated with mitigated acute inflammation and downstream fibrosis while simultaneously enhancing the intrinsic myogenic regenerative capacity of the tissue. These findings establish betulin as a potent therapeutic candidate for improving outcomes after severe muscle injuries.