Low-toxicity vitrification of small-diameter arterial grafts by achieving a Cryo-Stasis state

Small-diameter arterial grafts are crucial for vascular reconstruction yet lack preservation methods that maintain viability and function, as current approaches face a trade-off between ice damage from slow freezing and cytotoxicity from high-concentration cryoprotectants. We discovered that low-dose combinatorial ice inhibitors enhance cellular tolerance to cryoprotectant toxicity via lamin B1-mediated nuclear stabilization, which inhibits synergistic chemical and osmotic shock injury during unloading. Based on these findings, we developed CTX-RYST (Cryo & Toxico-Regulated Yoke-Steadying Technology), integrating a low-dose ice inhibitor (CTX) with a one-step unloading method that reduces processing time by 75% (to 15 min) and achieves superior viability and attachment versus conventional multi-step unloading. This strategy improved graft quality: Human Umbilical Vein Endothelial Cells adhesion increased from 8% to 89%, ice formation was nearly eliminated (melting enthalpy 0.28 J/g), and viscoelasticity and migration reached native levels. In rat carotid artery transplants, stenosis decreased from 24% to 4%. Thus, CTX-RYST achieves "Cryo-Stasis" (simultaneous suppression of ice and solution damage), establishing a low-toxicity cryopreservation paradigm that restores functional integrity for small-diameter vascular transplantation.