Dynamic tensile loading improves neotendon formation at moderate daily loading cycles, but impairs neotendon formation at high daily loading cycles

Synthetic polymer scaffolds with aligned fibers permit engineered tendon development and the formation of an aligned, collagen rich matrix. However, many tissue engineered constructs cannot withstand physiologically relevant loads. Both tendon development and normal homeostasis require loading, and tendons adapt to loading. In vitro loading of tissue engineered constructs further promotes engineered tendon development, but results vary by cell type, strain applied, and frequency of the loading. However, both under- and overloading of tendon is detrimental and disrupts stiffness and collagen organization in vivo, and levels of loading comparable to that which occurs during fetal development are generally unexplored for engineered tendon development. The objective of this study was to determine the optimal number of daily cycles of dynamic tensile loading for in vitro human adipose stem cell (hASC) cultured on poly(ε-caprolactone) 3D meltblown scaffolds. hASC-seeded scaffolds were loaded for 0 (control), 1,000 (late development fetal kicks/day), 5,000 (moderate exercise), 10,000 (used previously, analogous to target steps/day) cycles 3 times/week to 6% strain at 1 Hz. Loading at 5,000-cycles/session led to increased dsDNA, collagen, and collagen/dsDNA compared to unloaded control, and 1,000-cycles/session was intermediate in response. Loading up to 10,000-cycles/session increased dsDNA compared to unloaded control and the 5,000-cycle group but did not increase collagen content or collagen/dsDNA. Dynamic loading had no effect on glycosaminoglycan expression or collagen alignment. Loading at 5,000-cycles/session increased linear region modulus, yield stress, phase shift, and hysteresis and secant stiffness at high strains compared to the unloaded control but did not affect yield stretch or stress relaxation. However, the 10,000-cycle group was detrimental to mechanical properties, suggesting an overload phenotype.