34 | Hyaluronan enhances proliferation and myogenic differentiation of C2C12 murine myoblasts under inflammatory stress

Fabio Ferrini, G. Annibalini, M. Battistelli, F. Fanelli, S. Moosavi, R. Osman, I. Capparucci, P. Sestili, E. Barbieri | Dept. of biomolecular sciences, University of Urbino, Italy.

Hyaluronan (HA) is a non-sulfated glycosaminoglycan, widely used for medical and pharmaceutical applications including tissue muscle repair. A recent study demonstrates that HA activates muscle stem cells to repair damaged muscle. When muscle damage occurs, stem cells start producing and coating themselves with hyaluronic acid, driven by demethylase JMJD3, which allows muscle stem cell adaptation to inflammation and the initiation of muscle repair (Nakka et al., 2022). The aim of this work is to characterize the effect of Multifractionated - HA on myoblast rescue under inflammatory conditions, using C2C12 murine muscle cell proliferation and differentiation.  In this study, we investigated the potential of a HA mixture (2-1000 KDa, 1 mg/mL, Regenflex TeM, Regenyal Laboratories SRL) in increasing the proliferative and myogenic capacity of myoblasts in the presence or absence of pro-inflammatory agents (IL-1β, LPS), known to impair proliferation. Results revealed that TeM significantly improved reparative mechanisms and exerts a strong pro-proliferative effect, enhancing wound healing within 24 hours post-injury, even in the presence of inflammatory conditions. Additionally, we evaluated the myogenic potential of C2C12 cells treated with TeM by analyzing the gene and protein expression of key myogenic markers, including IGF-1, MyoD, Myogenin, Mrf4, myogenin, and MHC-1, even under inflammatory conditions. Moreover, TeM treatment upregulated myogenic biomarkers, suggesting a positive impact on differentiation pathways. Regenflex T&M has been shown to increase the repair process in injured myoblast monolayers, to restore healing capacity under otherwise limiting conditions and to stimulate the muscle differentation process. This data streghtens the rationale for using infiltrative intramuscular T&M to promote the repair of injured muscles in clinical settings.