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Background Tenocytes as specialised fibroblasts and inherent cells of tendons require mechanical load for their homeostasis. However, how mechanical overload compared to physiological load impacts on the tenogenic differentiation potential of fibroblasts is largely unknown. Methods Three-dimensional bioartificial tendons (BATs) seeded with murine fibroblasts (cell line C3H10T1/2) were subjected to uniaxial sinusoidal elongation at either overload conditions (0–16%, Ø 8%) or physiological load (0–8%, Ø 4%). This regime was applied for 2 h a day at 0.1 Hz for 7 days. Controls were unloaded, but under static tension. Results Cell survival did not differ among overload, physiological load and control BATs. However, gene expression of tenogenic and extra-cellular matrix markers ( Scx ,  Mkx ,  Tnmd ,  Col1a1  and  Col3a1)  was significantly decreased in overload  versus  physiological load and controls, respectively. In contrast,  Mmp3  was significantly increased at overload compared to physiological load, and significantly decreased under physiological load compared to controls.  Mkx  and  Tnmd  were significantly increased in BATs subjected to physiological load compared to controls. Proinflammatory interleukin-6 showed increased protein levels comparing load (both over and physiological)  versus  unloaded controls. Alignment of the cytoskeleton in strain direction was decreased in overload compared to physiological load, while other parameters such as nuclear area, roundness or cell density were less affected. Conclusions Mechanical overload decreases tenogenic differentiation and increases ECM remodelling/inflammation in 3D-stimulated fibroblasts, whereas physiological load may induce opposite effects.

Mechanical overload decreases tenogenic differentiation compared to physiological load in bioartificial tendons