Impact of cold storage protocols on passive arterial wall mechanical behavior

Mechanical characterization of arterial tissue is essential for understanding vascular function, pathologies, and for developing computational models and vascular devices. However, tissue storage prior to ex vivo testing can alter mechanical behavior, complicating data interpretation and comparability. This study systematically evaluated how commonly used cold storage protocols affect the passive viscoelastic behavior of the arterial wall. Specimens (n=177) from porcine descending aortas (n=12) underwent biaxial mechanical testing either fresh or after cold storage. Storage protocols (n=6) varied in temperature, freezing rate, and cryoprotectant use. Stored strain energy density during loading and strain energy density difference between loading and unloading cycles (loss factor) quantified elastic and viscous behavior, respectively. Linear mixed-effects models accounted for variability among animals and anatomical locations. Viscous behavior was largely preserved, whereas elastic behavior was significantly affected by cold storage. Refrigeration decreased strain energy density, while snap freezing and cryoprotectant use increased it, with more pronounced effects under circumferential loading. Slow freezing in phosphate-buffered saline to -80°C best preserved the passive mechanical behavior. Variability among animals and anatomical locations dominated variance in elastic (55%-65%) and viscous (∼65%) behavior, exceeding storage effects (15%-20% and <5%, respectively). Although limited to healthy porcine tissue and without direct assessment of the underlying mechanisms, these findings demonstrate that cold storage alters passive arterial mechanics, particularly elasticity, while variability among animals and anatomical locations is a critical source of variation. This study provides guidance for selecting appropriate storage protocols pre-mechanical testing and highlights the importance of variability among test subjects and anatomical locations.