Polyampholytes as cryoprotective agents for mammalian cell cryopreservation

Area of Science:

• Biomaterials Science
• Cell Biology
• Cryobiology

Background:

• Conventional cryoprotective agents (CPAs) like dimethyl sulfoxide (DMSO) are effective but exhibit significant cytotoxicity and can impact cell differentiation.
• There is a need for safer and more effective cryopreservation methods to maintain cell viability and function after thawing.

Purpose of the Study:

• To evaluate the efficacy of novel ɛ-poly-L-lysine (PLL) derivatives as cryoprotective agents.
• To compare the cryopreservation efficiency of PLL derivatives with conventional CPAs, including DMSO.
• To assess the impact of PLL-based CPAs on cell viability, proliferation, and differentiation.

Main Methods:

• Synthesis of PLL derivatives with modified amino groups (converted to carboxyl groups using succinic anhydride).
• Cryopreservation of L929 cells and rat mesenchymal stem cells using PLL solutions (7.5 w/w%) and conventional CPAs.
• Assessment of post-thaw cell survival, proliferation, and differentiation potential.

Main Results:

• PLL derivatives demonstrated higher post-thaw survival efficiency for L929 cells compared to current CPAs, even without protein additives.
• Rat mesenchymal stem cells were cryopreserved more effectively with PLL than with DMSO, retaining full proliferation and differentiation potential.
• PLL with an appropriate ratio of carboxyl groups enabled cryopreservation of diverse cell types, including adherent and suspension cells, primary cells, and cell lines.

Conclusions:

• Polymeric extracellular CPAs based on PLL derivatives show promise as replacements for conventional CPAs.
• The high viability after thawing and lack of serum requirement suggest broad applicability of these novel CPAs in various preservation fields.
• The antifreeze protein-like properties of PLL may contribute to its enhanced cryoprotective capabilities.