Characterizing modes of action and interaction for multicomponent osmolyte solutions on Jurkat cells

Abstract

This study examined the post‐thaw recovery of Jurkat cells cryopreserved in three combinations of five osmolytes including trehalose, sucrose, glycerol, mannitol, and creatine. Cellular response was characterized using low‐temperature Raman spectroscopy, and variation of post‐thaw recovery was analyzed using statistical modeling. Combinations of osmolytes displayed distinct trends of post‐thaw recovery, and a nonlinear relationship between compositions and post‐thaw recovery was observed, suggesting interactions not only between different solutes but also between solutes and cells. The post‐thaw recovery for optimized cryoprotectants in different combinations of osmolytes at a cooling rate of 1°C/min was comparable to that measured with 10% dimethyl sulfoxide. Statistical modeling was used to understand the importance of individual osmolytes as well as interactions between osmolytes on post‐thaw recovery. Both higher concentrations of glycerol and certain interactions between sugars and glycerol were found to typically increase the post‐thaw recovery. Raman images showed the influence of osmolytes and combinations of osmolytes on ice crystal shape, which reflected the interactions between osmolytes and water. Differences in the composition also influenced the presence or absence of intracellular ice formation, which could also be detected by Raman. These studies help us understand the modes of action for cryoprotective agents in these osmolyte solutions.

Cryopreservation of Human iPS Cell Aggregates in a DMSO-Free Solution—An Optimization and Comparative Study

“…The combination of previous studies (Pollock et al, 2016;Pi et al, 2019b) and new findings from this investigation provides insights into how the DMSO-free CPAs act to protect the hiPSCs from freezing damage. Mechanisms of cryoprotection for DMSO-free CPAs differ from that of DMSO, which is an organic solvent.…”

“…Both DOE and Taguchi methods assume that the topology of the parameter space that is being studied is unimodal and that there are no interactions between parameters. Both of these assumptions are not true, as this study has demonstrated that the topology is not unimodal, and our statistical model (Pi et al, 2019b) has shown that that there are synergistic effects between the CPA molecules tested, making these alternative approaches unsuitable. Emergent population staying constant for two generations was defined as the criterion of convergence for the DE algorithm.…”

“…We have demonstrated in previous studies that combinations of sugar, sugar alcohols and amino acids can be used to preserve human mesenchymal stem cells and T lymphocytes (Pollock et al, 2016(Pollock et al, , 2017Pi et al, 2018Pi et al, , 2019b. When used to cryopreserve cells in vitro, they have been shown to act in concert with each other and amino acids to protect and stabilize cells (Pollock et al, 2016;Pi et al, 2019b). This study uses a cocktail of DMSO-free CPAs including sucrose, glycerol, isoleucine, human serum albumin, and poloxamer 188 (P188), which are non-toxic and FDA-approved for infusion.…”

Preservation of cell-based immunotherapies for clinical trials

“…These might account for loss of functional cells in cryopreserved T-cell product for immunotherapy. In comparison, DMSO-free freezing media composed of sugars, sugar alcohols and amino acids have been demonstrated to successfully preserve Jurkat cells as a CD3 + CD4 + T-lymphocyte model with high postthaw viability [8,64] and can potentially be used to cryopreserve other types of T cells. Similar to T cells, although NK cells cryopreserved in DMSO have shown significant functional loss in terms of their cytotoxicity [60], a DMSO-free freezing medium composed of poly-l-lysine, dextran and ectoine has demonstrated improved post-thaw cytotoxicity in cryopreserved NK cells [65].…”

Natural deep eutectic systems for nature‐inspired cryopreservation of cells

“…The MCO solution was prepared by adding trehalose, glycerol, and isoleucine directly to Normosol‐R and mixed using a vortex mixer until a clear liquid was formed. The concentration of each component of the TGI MCO has been published previously 14 . A 10% vol/vol.…”