Cyclic mechanical stimulation of TDCs improves tendon tissue maturation in vitro by enhancing collagen formation

Tendinopathy represent a growing clinical problem and unmet therapeutic need, mainly due to the limited intrinsic regenerative capacity of tendons and our poor understanding of the underlying disease mechanisms. A major obstacle to progress in treatment is the lack of physiologically relevant models that accurately reflect the structure, mechanics and biology of tendons. While animal models provide valuable insights into tendon pathology and repair, in vitro 3D models offer a controlled and ethical alternative for studying the pathogenesis and progression of tendinopathy, especially investigating specific mechanobiological mechanisms. However, the lack of a standardized in vitro model makes it difficult to compare results from different studies, particularly with respect to the mechanical stimulation regimes applied. Existing approaches use either static or cyclic mechanical loading, and there is a lack of scientific consensus over which approach is advantageous. In this study, we addressed this question using a custom-made displacement-controlled bioreactor to compare the effect of static vs. cyclic loading on the development of 3D tissue-engineered tendons. In our experiments, cyclic loading application resulted in increased expression of tendon maturation markers, namely Col1, Tnmd, and Sparc, and cell elongation and alignment along the strain axis compared to static loading. Additionally, we report that cyclic loading promotes cell survival, proliferation and matrix remodeling, with enhanced collagen deposition. Structural and mechanical characterization confirmed fibril formation upon cyclic loading and higher stiffness of cyclically stimulated constructs. Together, these findings indicate that cyclic mechanical stimulation promotes key biological, structural, and mechanical features of early-stage tendon tissue maturation and provides a more favorable microenvironment for the development of in vitro tendon models compared to static loading.