Immunolabelling and Micro-Computed Tomography Revealed Age-Related Alterations in 3D Microvasculature of Tendons

Tendon degeneration is common, and its risk increases with age both in humans and horses. Tendon regeneration and healing is limited due to inherent low cell density and vascularisation, and current treatments are insufficient as indicated by scar tissue formation and a high re-injury rate. The tendon vasculature plays a crucial role in tendon homeostasis, regeneration and healing, making it a potential therapeutic target. However, the effect of ageing on the tendon microvasculature is poorly understood. Here, we provide the first comprehensive characterisation of the tendon microvasculature. We employed high-resolution 3D imaging techniques, using micro-computed tomography (μCT) and confocal microscopy, to investigate age-related alterations in the vasculature within the equine superficial digital flexor tendon (SDFT), a functional equivalent of the human Achilles tendon. μCT analysis revealed a well-developed vascular network within the interfascicular matrix (IFM) and demonstrated significant age-associated reductions in vascular volume (70%), vessel diameter (30%) and density (74%). 3D immunolabelling showed significant reductions in MYH11- (96%) and desmin-positive (78%) volumes; however, there was a pronounced age-associated increase in von Willebrand factor (VWF)-positive volume (220%), which was accompanied by a significantly higher (249%) pericyte density. Taken together, these results indicate a loss of larger blood vessels in the IFM but an increase in small vessel formation, suggesting that neo-angiogenesis is induced in aged tendon alongside a loss of vascular homeostasis. These insights enhance our understanding of tendon ageing and may contribute to developing new therapeutic approaches for improving tendon health and repair in older individuals.