Dextran polyampholyte hydrogels for cell cryopreservation

One of the goals of tissue engineers is to manufacture “spare body parts” made of cells and biomaterials to repair damaged or diseased tissues. A current challenge for commercializing cell-biomaterial constructs is maintaining their viability during shipping and/or storage. The storage of cells in liquid suspension is relatively simple with the use of common cryoprotectants (such as dimethyl sulfoxide), but current preservation methods for large cell-biomaterial constructs yield low cell viability in comparison.  More effective preservation methods for cell-biomaterial constructs are needed to produce “off the shelf” tissue engineered products and accelerate the translation of tissue engineered products to the clinic.

In a current study conducted at the Japan Advanced Institute of Science and Technology (JAIST), researchers developed a dextran-based polyampholyte (charged polymer) hydrogel with cryoprotective properties. First, dextran polymers were functionalized with azide groups (azide-Dex) to provide sites for crosslinking. Then, positively charged amine groups were introduced on the dextran using poly-L-lysine to form azide-amino-Dex. To add negatively charged carboxyl groups, succinic anhydride was added at various concentrations to azide-amino-Dex to convert amine groups to carboxyl groups. This newly formed dextran polyampholyte (azide-Dex-PA) polymer was utilized for cryopreservation studies. After encapsulating and freezing cells with various azide-Dex-PA solutions, cell viability post-cryopreservation was shown to increase with higher concentrations of azide-Dex-PA. Viability was also dependent on the ratio of amine and carboxyl groups on the polymer, indicating that the cryoprotective properties of the polyampholyte are likely due to polymer charge.

The azide groups on the azide-Dex-PA were utilized to crosslink the dextran polyampholytes to form hydrogels. After synthesizing alkyne-functionalized dextran with dibenzylcyclooctyne (DBCO-Dex), the two components were mixed with cells to form hydrogels with azide-alkyne click chemistry. Encapsulated cells in the hydrogel were cryopreserved, thawed, and evaluated for viability. Compared to the 0% viability observed with collagen hydrogel and non-polyampholyte dextran hydrogel controls, the dextran polyampholyte hydrogels substantially increased cell viability to 93-94%. The increased viability post-cryopreservation may be due to the polyampholyte polymer adsorbing onto the cell membrane, providing a polymer “shell” of protection during thawing. Images of cells coated with FITC labeled azide-Dex-PA provide evidence to support this hypothesis.

All in all, dextran polyampholyte hydrogels have cryoprotective properties, and are able to preserve cell viability during the cryopreservation process. The dramatic increase in cell viability post-cryopreservation justifies future studies for using these hydrogels for preserving larger cell-biomaterial constructs for long term storage.

Hydrogelation of dextran-based polyampholytes with cryoprotective properties via click chemistry
Minkle Jain, Robin Rajan, Suong-Hyu Hyon, and Kazuaki Matsumura
Biomaterials Science, 2014, Advance Article DOI: 10.1039/C3BM60261C

Brian Aguado is currently a Ph.D. Candidate and NSF Fellow in the Biomedical Engineering department at Northwestern University. He holds a B.S. degree in Biomechanical Engineering from Stanford University and a M.S. degree in Biomedical Engineering from Northwestern University. Read more about Brian’s research publications here.

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