Understanding Ice Recrystallization Inhibitors - Design, Synthesis and Mechanism(s) of Action

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Cell-based therapeutics emerged as a critical aspect of modern healthcare in regenerative medicine and transfusion. Unfortunately, current cryopreservation protocols remain suboptimal and limit the effectiveness of such therapies. Loss of cell viability during cryopreservation has been shown to dramatically reduce potency of progenitor cells such as those used in hematopoietic stem cell (HSC) transplantation and red blood cell (RBC) transfusion resulting in poor engraftment or a need for additional transfusions. Recent studies have indicated that the most significant cause of decreased post-thaw cell viability and loss of function is cellular damage from ice recrystallization during freezing and thawing. A detailed understanding of this process is fundamental to the natural sciences since practical strategies to inhibit it are lacking. This multidisciplinary proposal will address this issue using in vitro approaches relying on advanced spectroscopic (confocal Raman) and modern cellular/biophyscial techniques (nanoparticle tracking, Ektacytometric analysis, cryomicroscopy etc.). Specific objectives are; 1) study and correlate the ability of our novel ice recrystallization inhibitors (IRIs) to their physical parameters (hydrophobicity, self-assembly and hydration properties) and 2) study the in vitro cellular effects of these molecules on cellular membranes, microparticle (MP) formation, nucleation of ice and the recrystallization of intracellular ice. The knowledge gained from understanding how these inhibitors function will significantly advance our understanding of ice recrystallization (both in the presence and absence of cells). This knowledge will be directly relevant to other processes such as biomineralization, and calcification and lead to the generation of novel methods to control these physiologically important processes.***Significance/Impact:***Our integrated approach of using in vitro strategies relying on advanced spectroscopic (confocal Raman) and modern cellular/biophyscial techniques (nanoparticle tracking, Ektacytometric analysis, cryomicroscopy etc.) to study small molecule IRIs will expedite an understanding of how these inhibitors function. In addition, the design of new and improved cryoprotectants is timely and will address many of the storage problems associated with technologies in the areas of regenerative medicine, cellular therapy and tissue engineering.