20. The complexity of tissue and organ cryopreservation: A call for interdisciplinary research

Abstract

The development of cryopreservation protocols for cell-based therapeutics have been based on established strategies to balance the “two-factors” of cryoinjury that influence cell recovery. By managing the competing effects of ice formation and solute composition changes on the cell through the use of carefully choreographed cryoprotectant addition/removal and cooling/thawing processes, successful protocols have been developed for many cell systems. Knowledge of cryoprotectant toxicity, ice formation and cell volume excursions, has allowed different cryopreservation strategies to be custom developed for the same cell system to meet the needs of the biobank or clinical program. The unique physical properties of tissues and organs have been shown to significantly affect the biological response to freezing and thawing. The diversity of cell types and cell densities as well as the morphological differences between constituent cells significantly affects the osmotic and thermal state of tissues and organs. This has dramatic implications on the cooling and thawing rates that can be attained and hence the response of the tissue to freezing and thawing. In addition, the requisite cell–cell and cell–matrix interactions in a tissue have been implicated in the poor survival of tissues following freezing. It is clear that the freezing response of tissues and organs, and the mechanism by which damage occurs are affected by the physical structures of these complex systems. In complex systems, ice crystallization becomes much more complicated and difficult to balance through the use of conventional cryoprotectants and cooling/thawing rates due to thermal and mass transfer limitations creating damaged cellular zones within the tissue or organ. Conventional biophysical approaches to modulate cell response to freezing and thawing may need to be supplemented with molecular strategies to directly modulate cell stress response. Vitrification, or the avoidance of ice crystallization, may address the direct effects of ice crystallization and solute concentrations, but challenges with cryoprotectant permeation and toxicity create new obstacles. Minimization of interstitial and extracellular ice in specific locations within tissues and organs may be more important and achievable than complete elimination. Developing tools to selectively control ice crystallization will be required. To advance the science of tissue and organ cryopreservation will require a concerted effort by a broad interdisciplinary field of scientists, engineers, and clinicians working to develop new strategies and tools. The ideas presented at this conference will hopefully stimulate not only the search for improved techniques for the cryopreservation of tissues and organs, but more importantly, a desire to clearly understand the response of biological systems to low temperatures.