UC Davis

Cell-based therapies have been a growing topic of interest within biomedical research. Despite many advances, the challenges with clinical translation twenty years ago persist today: problems associated with scaling up production and cell death associated with implantation. For clinical success, a holistic approach incorporating the current challenges in cell-based therapies would increase the probability of success. Two prospective methods to improve cell delivery outcomes include biomaterials and spheroids. Nevertheless, it is unclear how innovations with these methods will integrate with the need to preserve cell-based therapies via cryopreservation, the storage of cells in frozen stasis. Notably, the freezing and thawing process has negative consequences for cell replication, unintended differentiation, poor recovery, and morphological and functional changes. These issues are worsened when freezing larger tissues or constructs, such as spheroids and cells encapsulated in hydrogels. To address this challenge, we developed a novel gelation procedure for alginate hydrogels capable of encapsulating cells as they thaw from a cryopreserved state, which we call thaw-induced gelation. This method is a distinct alternative to cryopreserving cell-based therapies whole. We developed three new alginate hydrogels compatible with cryopreservation: homo-IPN, alginate cryogels, and hydrogel capsules. Using each method with cryopreserved cells could instruct cell aggregation to form spheroids. Additionally, these hydrogels showed promise for protecting cargo from freeze/thaw damage and could enhance electrostatic-based release from the hydrogels. Lastly, we interrogated the optimal procedure to cryopreserve cells to obtain spheroids using HUVECs, MSCs, and a combination of both cell types. Our results suggested that best practices are likely cell-dependent, indicating cryopreservation will not have a one-size-fits-all approach for potential cell therapies. Overall, thaw-induced gelation represents one of the first efforts to design hydrogel-based systems around the mechanisms of cryopreservation, minimizing the conflicts with cryopreserving tissue-engineered constructs whole. Together, the system represents an alternative approach to incorporating cryopreservation with prospective additional uses in tissue or disease modeling, scale-up production of 3D cultures, and studying the effects of cryopreservation.