Red photobiomodulation promotes skeletal myoblast differentiation and counteracts anti myogenic action of TGF-b1 in vitro: morpho-functional evidences

Background: adult skeletal muscle regenerates lost damaged tissue mainly thanks to the activity of a small population of resident stem cells, namely satellite cells (SCs). In the case of chronic or severe damage, SCs’ functionality may be compromised by the occurrence of an aberrant fibrotic reparative response. Strategies aimed to improve the muscle intrinsic regenerative capacity while limiting the excessive deposition of fibrotic tissue may be promising. In this perspective, photobiomodulation (PBM) (i. e. application of light with 400-1100 nm wavelength using different laser or LED devices, power density less than 100 mW/cm2 and energy density less than 10 J/ cm2 at target) may represent a valid option based on its well-known pro-regenerative effects and increasing evidence of its antifibrotic potential. However, PBM's effects on skeletal muscle are controversial, and there are no univocal guidelines for its use. Aim: to evaluate the effects of different treatments of red PBM (laser diode 635±5 nm, energy density: 0.4, 4 and 8 J/cm2, single exposure) on murine C2C12 myoblasts undergoing differentiation in the presence or absence of TGF-β1 (2ng/ml) and on differentiated myotubes. Methods and Results: MTS cell viability assay, morphological analyses (myotube formation, confocal immunofluorescence analysis of the expression of myogenic markers and of mitochondrial PGC1-ɑ, transmission electron microscopy) and electrophysiological recordings (cell membrane passive properties and ion currents) revealed that red PBM with 4 J/cm2 energy density improved myoblast differentiation and did not alter differentiated myotube viability and features. Red PBM with 4 J/cm2 energy density was able to promote cell myogenic differentiation in the presence of TGF-β1 thus counteracting the TGF-β1 anti-myogenic action. Conclusions: this study provides experimental compelling evidence supporting red PBM’ pro-myogenic effects offering new cues for further investigations. Future Investigations: exploring red PBM’s effects on myoblasts cultured in the presence of corticosteroids (Triamcinolone acetonide, Dexamethasone) usually clinically used to treat some inflammatory states of musculoskeletal tissues. Despite their anti-inflammatory properties, the effectiveness of corticosteroid injections on healing is debated. Evaluating red PBM on human skeletal myoblasts and on both murine and human myoblasts induced to differentiate on a liquid crystalline network used as a cell instructive scaffold to support a correct myogenic differentiation.