Evaluating flash freezing for preservation of rat abdominal aorta for delayed biomechanical characterization

Most studies investigating arterial stiffening use animal rather than human arteries. This is because human tissue becomes available in small amounts and at irregular times, which complicates planning of experimental work. Suitable tissue preservation methods for delayed biomechanical testing prevents the need for testing fresh tissue and alleviates some of the logistical challenges of human ex vivo studies. Therefore, the present study aimed to investigate whether the existing method of flash freezing and subsequent cryostorage provides is suitable for delaying the characterization of arterial biomechanics. Fresh and flash frozen abdominal aortas (n=16 and 14, respectively) were quasi- statically and dynamically tested using a biaxial testing set-up with dynamic pressurization capabilities. The acquired biomechanical data was modeled using a constituent-based quasi-linear viscoelastic modeling framework, deriving directional stiffness parameters, individual constituent biomechanical contributions, and viscoelastic stiffening under dynamic pressurization conditions. Flash freezing reduced arterial wall thickness, increased circumferential stiffness, as well as reduced viscoelastic stiffening at higher pressures. These findings reflected those in the modeled contribution of collagen to arterial biomechanics, showing increased collagen load bearing at higher pressures. However, despite the above mentioned detectable changes, flash freezing did not alter the mechanical relation between elastin and collagen, maintaining a non-linear response to pressurization and stretch. Flash freezing may thus be suitable for studies requiring delayed characterization of passive arterial biomechanics, assuming care is taken to ascert that the impact of flash freezing on study groups can be approached as a systematic error.