Abstract
The morphological changes of small (~100 µm) alginate microcapsules and the biophysical alterations of water in the microcapsules during cryopreservation were studied using cryomicroscopy and scanning calorimetry. It was found that water in the small microcapsules can be preferentially vitrified over water in the bulk solution in the presence of 10% (v/v) or more dimethylsulfoxide (DMSO, a cryoprotectant), which resulted in an intact morphology of the microcapsules post cryopreservation with a cooling rate of 100°C/min. A small amount of Ca2+ (up to 0.15 M) was also found to help maintain the microcapsule integrity during cryopreservation, which is attributed to the enhancement of the alginate matrix strength by Ca2+ rather than promoting vitrification of water in the microcapsules. The preferential vitrification of water in small microcapsules was further found to significantly augment cell cryopreservation by vitrification at a low concentration of cryoprotectants (i.e., 10% (v/v)) using a small quartz microcapillary (400 µm in diameter). Therefore, the small alginate microcapsule could be a great system for protecting living cells that are highly sensitive to stresses due to freezing (i.e., ice formation) and high concentration of cryoprotectants from injury during cryopreservation.
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References
V. Berejnov, N.S. Husseini, O.A. Alsaied, R.E. Thorne, Effects of cryoprotectant concentration and cooling rate on vitrification of aqueous solutions. J. Appl. Crystallogr. 39, 244–251 (2006)
N. Blow, Biobanking: freezer burn. Nature Methods 6(2), 173–177 (2009)
T.M.S. Chang, Semipermeable Microcapsules. Science 146(364), 524–526 (1964)
R. Coger, M. Toner, Preservation Techniques for Biomaterials, in The Biomedical Engineering Handbook:, 2nd edn., ed. by J.D. Bronzino (CRC Press LLC, Boca Raton, 2000)
P. de Vos, M.M. Faas, B. Strand, R. Calafiore, Alginate-based microcapsules for immunoisolation of pancreatic islets. Biomaterials 27(32), 5603–5617 (2006)
K.R. Diller, Bioheat and mass transfer as viewed through a microscope. J Biomech Eng 127(1), 67–84 (2005)
V. Dixit, G. Gitnick, Transplantation of microencapsulated hepatocytes for liver function replacement. J Biomater Sci Polym Ed 7(4), 343–357 (1995)
G.M. Fahy, B. Wowk, J. Wu, S. Paynter, Improved vitrification solutions based on the predictability of vitrification solution toxicity. Cryobiology 48(1), 22–35 (2004)
J. Gearhart, New potential for human embryonic stem cells. Science 282(5391), 1061–1062 (1998)
B. Han, J.C. Bischof, Thermodynamic nonequilibrium phase change behavior and thermal properties of biological solutions for cryobiology applications. J Biomech Eng 126(2), 196–203 (2004)
B. Han, J.H. Choi, J.A. Dantzig, J.C. Bischof, A quantitative analysis on latent heat of an aqueous binary mixture. Cryobiology 52(1), 146–151 (2006)
X. He, E.Y. Park, A. Fowler, M.L. Yarmush, M. Toner, Vitrification by ultra-fast cooling at a low concentration of cryoprotectants in a quartz micro-capillary: a study using murine embryonic stem cells. Cryobiology 56(3), 223–232 (2008)
B.C. Heng, H. Yu, S.C. Ng, Strategies for the cryopreservation of microencapsulated cells. Biotech Bioeng 85(2), 202–213 (2004)
J.O.M. Karlsson, M. Toner, Long-term storage of tissues by cryopreservation: Critical issues. Biomaterials 17(3), 243–256 (1996)
R. Langer, J.P. Vacanti, Tissue engineering. Science 260(5110), 920–926 (1993)
F. Lim, A.M. Sun, Microencapsulated Islets as Bioartificial Endocrine Pancreas. Science 210(4472), 908–910 (1980)
A. Murua, A. Portero, G. Orive, R.M. Hernandez, M. de Castro, J.L. Pedraz, Cell microencapsulation technology: towards clinical application. J Control Release 132(2), 76–83 (2008)
G. Orive, R.M. Hernandez, A. Rodriguez Gascon, R. Calafiore, T.M. Chang, P. de Vos, G. Hortelano, D. Hunkeler, I. Lacik, J.L. Pedraz, History, challenges and perspectives of cell microencapsulation. Trends Biotechnol 22(2), 87–92 (2004)
D.E. Pegg, The role of vitrification techniques of cryopreservation in reproductive medicine. Hum Fertil (Camb) 8(4), 231–239 (2005)
W.F. Rall, G.M. Fahy, Ice-Free Cryopreservation of Mouse Embryos at -196-Degrees-C by Vitrification. Nature 313(6003), 573–575 (1985)
V. Stensvaag, T. Furmanek, K. Lonning, A.J. Terzis, R. Bjerkvig, T. Visted, Cryopreservation of alginate-encapsulated recombinant cells for antiangiogenic therapy. Cell Transplant 13(1), 35–44 (2004)
M. Toner, J. Kocsis, Storage and translational issues in reparative medicine. Ann. N. Y. Acad. Sci. 961, 258–262 (2002)
Y. Wu, H. Yu, S. Chang, R. Magalhaes, L.L. Kuleshova, Vitreous cryopreservation of cell-biomaterial constructs involving encapsulated hepatocytes. Tissue Eng 13(3), 649–658 (2007)
G. Yang, A. Zhang, LX. Xu, X. He, Modeling the cell-type dependence of diffusion-limited intracellular ice nucleation and growth during both vitrification and slow freezing. J. Appl. Physi. 105(11):114701 (11pp) (2009).
W. Zhang, X. He, Encapsulation of Living Cells in Small (~ 100 mu m) Alginate Microcapsules by Electrostatic Spraying: A Parametric Study. J. Biomech. Eng. 131(7):074515 (6pp) (2009).
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This work was partially supported by a startup fund from the USC Research Foundation (provided through NSF grant # EPS-0447660) and by the Chinese Ministry of Education for a joint doctoral training program.
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Wujie Zhang and Geer Yang have contributed equally to this work.
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Zhang, W., Yang, G., Zhang, A. et al. Preferential vitrification of water in small alginate microcapsules significantly augments cell cryopreservation by vitrification. Biomed Microdevices 12, 89–96 (2010). https://doi.org/10.1007/s10544-009-9363-z
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DOI: https://doi.org/10.1007/s10544-009-9363-z