Heart valve cryopreservation was employed as a model for the development of complex tissue preservation methods based upon vitrification and nanowarming. Porcine heart valves were loaded with cryoprotectant formulations step wise and vitrified in 1–30 mL cryoprotectant formulations ± Fe nanoparticles ± 0.6 M disaccharides, cooled to −100 °C, and stored at −135 °C. Nanowarming was performed in a single ~100 s step by inductive heating within a magnetic field. Controls consisted of fresh and convection-warmed vitrified heart valves without nanoparticles. After washing, cell viability was assessed by metabolic assay. The nanowarmed leaflets were well preserved, with a viability similar to untreated fresh leaflets over several days post warming. The convection-warmed leaflet viability was not significantly different than that of the nanowarmed leaflets immediately after rewarming; however, a significantly higher nanowarmed leaflet viability (p < 0.05) was observed over time in vitro. In contrast, the associated artery and fibrous cardiac muscle were at best 75% viable, and viability decreased over time in vitro. Supplementation of lower concentration cryoprotectant formulations with disaccharides promoted viability. Thicker tissues benefited from longer-duration cryoprotectant loading steps. The best outcomes included a post-warming incubation step with α-tocopherol and an apoptosis inhibitor, Q-VD-OPH. This work demonstrates progress in the control of ice formation and cytotoxicity hurdles for the preservation of complex tissues.

Static cold storage is the cheapest and easiest method and current gold standard to store and preserve donor organs. This study aimed to compare the preservative capacity of gluconate-lactobionate-dextran (Unisol) solutions to histidine-tryptophan-ketoglutarate (HTK) solution. Murine syngeneic heterotopic heart transplantations (Balb/c-Balb/c) were carried out after 18 h of static cold storage. Cardiac grafts were either flushed and stored with Unisol-based solutions with high-(UHK) and low-potassium (ULK) ± glutathione, or HTK. Cardiac grafts were assessed for rebeating and functionality, histomorphologic alterations, and cytokine expression. Unisol-based solutions demonstrated a faster rebeating time (UHK 56 s, UHK + Glut 44 s, ULK 45 s, ULK + Glut 47 s) compared to HTK (119.5 s) along with a better contractility early after reperfusion and at the endpoint on POD 3. Ischemic injury led to a significantly increased leukocyte recruitment, with similar degrees of tissue damage and inflammatory infiltrate in all groups, yet the number of apoptotic cells tended to be lower in ULK compared to HTK. In UHK- and ULK-treated animals, a trend toward decreased expression of proinflammatory markers was seen when compared to HTK. Unisol-based solutions showed an improved preservative capacity compared with the gold standard HTK early after cardiac transplantation. Supplemented glutathione did not further improve tissue-protective properties.