The effect of multiple freeze-thaw cycles on the viscoelastic properties and microstructure of bovine superficial digital flexor tendon

The most common injuries of the human musculoskeletal system are related to soft tissue structures such as tendons or ligaments. To repair torn structures, surgical intervention and application of a biological or synthetic graft may be required. A typical procedure for the processing, storage, and distribution of soft tissue grafts involves at least two freezing/thawing (F/T) cycles. Even though repeated F/T cycles decrease the mechanical performance and change the structure of tendons, it is unclear whether there exists a maximum number of F/T cycles above which tendons should not be approved for use as a tissue allograft. To fill this research gap, we present an ex vivo study on the effects of repetitive F/T cycles on the biomechanical stability of bovine superficial digital flexor tendon tissue. Using mechanical testing supported with scanning electron microscopy imaging, we show that multiple F/T cycles affect the viscoelastic and structural properties of tissue by significantly reducing its tensile modulus after the 3rd or 4th F/T cycle (depending on the strain range), stress drop during relaxation after the 8th F/T cycle (regardless the strain values), mechanical hysteresis after the 10th F/T cycle, and by causing a significant decrease in collagen fibril diameter. Our results provide a deeper insight into understanding the mechanisms responsible for tissue damage during multiple F/T cycles, and thus, may be useful for the future optimization of tissue storage protocols.