Translating cryoprotectant agents from zebrafish to mammals: a two-stage approach to advance cardiac cryopreservation

Cryopreservation of organs is an emerging technique to address shortage of donated organs for transplantation. Cryoprotectant agents (CPAs) serve to halt unwanted ice formation, prevent cell damage, and ultimately protect organs from harm during cryopreservation. The challenges in cryopreserving mammalian hearts have highlighted the need for a comprehensive screening of promising CPAs in high throughput and translationally relevant model organisms.

To meet this need, a large-scale CPA screening was first conducted in zebrafish larvae with the goal of determining optimal cocktails of CPAs. With identified cocktails, survival rates and morphological changes were assessed in larvae following cryopreservation. To translate the findings from zebrafish to mammals, the two most successful combinations of CPAs were chosen based on efficacy and survival for further investigation in rodent hearts. To accommodate the complexity of mammalian hearts, the parameters of the preservation protocol have been optimized for rodent hearts, anticipating further optimization for larger mammalian hearts.

To that end, this study focused on optimizing the protocol of loading CPAs into the rodent cardiac tissue - an ongoing challenge in achieving successful cryopreservation of organs. To identify the best way of minimizing cardiac damage caused by sudden changes in osmotic gradient and CPA toxicity while also achieving accurate preservation outcomes, this study investigated several CPA loading protocols based on the status of cardiac graft health. Using a Langendorff perfusion system, functional and injury markers were collected during a four-hour recovery period, providing information to evaluate the health of the cardiac graft after CPA loading and unloading - all prior to sub-zero preservation.

The findings have important implications for the advancement of cryopreservation techniques for cardiac allografts, which may be an important step in tackling the organ shortage crisis. Moving forward, further research should focus on optimizing cryopreservation protocols, exploring species-specific variations, and evaluating CPA performance in both animal and human trials.