Ageing-Related Structural and Cellular Alterations in the Mouse Muscle-Tendon Junction

The muscle tendon junction (MTJ) is a specialised interface between muscle and tendon and transmits muscle-generated force to the tendon. The MTJ is particularly vulnerable to injuries compared to muscle and tendon and becomes more injury prone with age. Further, current treatments for MTJ injuries are insufficient as indicated by scar tissue formation and a high re-injury rate. Despite its clinical importance, the mechanisms driving MTJ ageing and age-related functional deterioration remain poorly understood. In this study, the first comprehensive three-dimensional characterisation of age-related structural and cellular changes at the mouse MTJ was performed using the high-resolution imaging techniques, micro-computed tomography (µCT) and confocal microscopy. µCT analysis revealed a 27% reduction in muscle fibre diameter with age, accompanied by a trend toward increased MTJ surface area and a 19% reduction in pennation angle, indicating diminished force generation capacity. Confocal imaging showed a 49% reduction in endothelial cell volume (VWF-labelled) in the old mouse muscle-tendon unit, suggesting a loss of vascularity. In situ hybridisation and immunofluorescence demonstrated increased expression of senescence markers p16 and p21 in endothelial and MTJ-specific cells, with MTJ-specific cells showing the greatest accumulation of p16 and p21 (270% and 310% increases, respectively) with age. These findings suggest that vascular and MTJ-specific cells are particularly susceptible to ageing and may collectively contribute to the age-related functional decline of the MTJ. Understanding these mechanisms may help to develop targeted therapeutic strategies to preserve or restore MTJ integrity and function in ageing populations.