Abstract
Thawing in the water bath is often considered as a standard procedure. The thermal history of samples thawed in this way is poorly controlled, but cryopreservation and banking of cell-based products require standardization, automation and safety of all the technological stages including thawing. The programmable freezers allow implementation of the controlled cooling as well as the controlled thawing. As the cell damage occurs during the phase transformation that takes place in the cryoprotectant medium in the process of freezing–thawing, the choice of warming rates within the temperature intervals of transformations is very important. The goal of the study was to investigate the influence of warming rates within the intervals of the phase transformations in the DMSO-based cryoprotectant medium on the cell recovery and to develop a cryopreservation protocol with controlled cooling and warming rates. The temperature intervals of phase transformations such as melting of the eutectic mixture of the cryoprotectant solution (MEMCS), melting of the eutectic salt solution (MESS), melting of the main ice mass (MMIM), recrystallization before MEMCS, recrystallization before MESS and recrystallization before MMIM were determined by thermo-mechanical analysis. The biological experiments were performed on the rat testicular interstitial cells (TIC). The highest levels of the cell recovery and metabolic activity after cryopreservation were obtained using the protocol with the high (20 °C/min) warming rate in the temperature intervals of crystallization of the eutectics as well as recrystallizations and the low (1 °C/min) warming rate in the temperature intervals of melting of the eutectics as well as MMIM. The total cell recovery was 65.3 ± 2.1 %, the recovery of the 3-beta-HSD-positive (Leydig) cells was 82.9 ± 1.8 %, the MTT staining was 32.5 ± 0.9 % versus 42.1 ± 1.7 %; 57.4 ± 2.1 % and 24.0 ± 1.1 % respectively, when compared to the thawing in the water bath.
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References
Akhtar T, Pegg DE, Foreman J (1979) The effect of cooling and warming rates on the survival of cryopreserved L-cells. Cryobiology 16:424–429
Bank H (1973) Visualization of freezing damage. II. Structural alterations during warming. Cryobiology 10:157–170
Barth AD, Bowman PA (1988) Determination of the best practical method of thawing bovine semen. Can Vet J 29:366–369
Berridge MV, Herst PM, Tan AS (2005) Tetrazolium dyes as tools in cell biology: new insights into their cellular reduction. Biotechnol Annu Rev 11:127–152
Coco-Martin JM, Oberink JW, van der Velden-de Groot TA, Beuvery EC (1992) Viability measurements of hybridoma cells in suspension cultures. Cytotechnology 8:57–64
Dowell LG, Rinfert AP (1960) Low temperature forms of ice as studied by X-ray diffraction. Nature 188:1144–1149
Dumont F, Marechal P, Gervais P (2003) Influens of cooling rate on Saccharomyces cerevisiae destruction during freezing: unexpected viability at ultra-rapid cooling rates. Cryobiology 46:33–42
El-Naggar MM, Al-Mashat FM, Elayat AA, Sibiany AR, Ardawi MS, Badawoud MH (2006) Effect of thawing rate and post-thaw culture on the cryopreserved fetal rat islets: functional and morphological correlation. Life Sci 78:1925–1932
Etheridge ML, Xu Y, Rott L, Choi J, Glasmacher B, Bischof JC (2014) RF heating of magnetic nanoparticles improves the thawing of cryopreserved biomaterials. Technology 2:229–242
Fuller B, Green C, Grischenko V (2003) Cryopreservation for cell banking: current concepts at the turn of the 21st century. Probl Cryobiol 10:63–82
Gao D, Critser JK (2000) Mechanisms of cryoinjury in living cells. ILAR J 41:187–196
Gurina TM, Kyryliuk AL (2012) Temperature ranges of phase transformations in the cryoprotective media components determined by thermoplastic deformation method. Probl Cryobiol 22:410–422
Gurina TM, Pakhomov AV, Kyryliuk AL, Bozok GA (2011) Development of a cryopreservation protocol for testicular interstitial cells with the account of temperature intervals for controlled cooling below −60 °C. Cryobiology 62:107–114
Guttman FM, Bosisio RG, Bolongo D, Segal N, Borzone J (1980) Microwave illumination for thawing frozen canine kidneys: (A) assessment of two ovens by direct measurement and thermography. (B) The use of effective dielectric temperature to monitor change during microwave thawing. Cryobiology 17:465–472
Harris LW, Griffiths JB (1977) Relative effects of cooling and warming rates on mammalian cells during the freezing–thaw cycle. Cryobiology 14:662–669
Henry MA, Noiles EE, Gao D, Mazur P, Critser JK (1993) Cryopreservation of human spermatozoa. IV. The effects of cooling rate and warming rate on the maintenance of motility, plasma membrane integrity, and mitochondrial function. Fertil Steril 60:911–918
Hopkins JB, Badeau R, Warkentin M, Thorne RE (2012) Effect of common cryoprotectants on critical warming rates and ice formation in aqueous solutions. Cryobiology 65:169–178
Hunt CJ (2011) Cryopreservation of human stem cells for clinical application: a review. Transfus Med Hemother 38:107–123
Jin B, Mazur P (2015) High survival of mouse oocytes/embryos after vitrification without permeating cryoprotectants followed by ultra-rapid warming with an IR laser pulse. Sci Rep 5:9271
Jin B, Kusanagi K, Ueda M, Seki S, Valdez DM Jr, Edashige K, Kasai M (2008) Formation of extracellular and intracellular ice during warming of vitrified mouse morulae and its effect on embryo survival. Cryobiology 56:233–240
Karlsson JOM (2001) A theoretical model of intracellular devitrification. Cryobiology 42:154–169
Karlsson JOM, Cravalho EG, Toner M (1993) Intracellular ice formation: causes and consequences. CryoLetters 14:323–336
Klinefelter GR, Hall PF, Ewing LL (1987) Effect of luteinizing hormone deprivation in situ on steroidogenesis of rat Leydig cells purified by a multistep procedure. Biol Reprod 36:769–783
Koshimoto C, Mazur P (2002) Effects of cooling and warming rate to and from −70 degrees C, and effect of further cooling from −70 to −196 degrees C on the motility of mouse spermatozoa. Biol Reprod 66:1477–1484
Leibo SP, Farrant J, Mazur P, Hanna MG Jr, Smith LH (1970) Effect of freezing on marrow stem cell suspensions: interaction of cooling and warming rates in present of PVP, sucrose or glycerol. Cryobiology 6:315–332
Leibo SP, McGrath JJ, Gravalho EG (1978) Microscopic observation of intracellular ice formation in unfertilized mouse ova as a function of cooling rates. Cryobiology 15:257–271
Li Y, Ma T (2012) Bioprocessing of cryopreservation for large-scale banking of human pluripotent stem cells. Biores Open Access 1:205–214
Lopez E, Cipri K, Naso V (2012) Technologies for cryopreservation: overview and innovation. In: Katkov I (ed) Current frontiers in cryobiology. InTech, Available from: http://www.intechopen.com/books/current-frontiers-in-cryobiology/technologies-for-cryopreservation-overview-and-innovation
Maeno N (1988) Science about Ice (translat. from Jap.), Moscow, Mir, p. 231
Maffei S, Brevini TAL, Gandolfi F (2014) Freezing and freeze-drying: the future perspective of organ and cell preservation. In: Brevini TAL (ed) Stem cells in animal species from pre-clinic to biodiversity. Humana Press, pp 167–184. doi:10.1007/978-3-319-03572-7
May SR, Wainwright JF (1985) Optimum warming rates to maintain glucose metabolism in porcine skin cryopreserved by slow cooling. Cryobiology 22:196–202
Mazur P (1966) Theoretical and experimental effects of cooling and warming velocity on the survival of frozen and thawed cells. Cryobiology 2:181–192
Mazur P (1967) Physical and chemical basis of injury in single-celled microorganisms subjected to freezing and thawing. In: Meryman HT (ed) Cryobiology. Academic Press, New York, pp 214–316
Mazur P (1984) Freezing of living cells: mechanisms and implications. Am J Physiol Cell Physiol 247:125–142
Mazur P, Seki S (2011) Survival of mouse oocytes after being cooled in a vitrification solution to −196 °C at 95° to 70,000 °C/min and warmed at 610° to 118,000 °C/min: a new paradigm for cryopreservation by vitrification. Cryobiology 62:1–7
Mazur P, Pinn IL, Kleinhans FW (2007) Intracellular ice formation in mouse oocytes subjected to interrupted rapid cooling. Cryobiology 55:158–166
Osetsky AI (2009) Peculiarities of state diagrams of aqueous solutions of cryoprotective agents. Cryobiology 59:141–149
Parmegiani L, Tatone C, Cognigni GE, Bernardi S, Troilo E, Arnone A, Maccarini AM, Di Emidio G, Vitti M, Filicori M (2014) Rapid warming increases survival of slow-frozen sibling oocytes: a step towards a single warming procedure irrespective of the freezing protocol? Reprod Biomed Online 28:614–623
Pegg DE, Diaper MP, Skaer HL, Hunt CJ (1984) The effect of cooling rate and warming rate on the packing effect in human erythrocytes frozen and thawed in presence of 2 M glycerol. Cryobiology 21:491–502
Perseghin P, Balduzzi A, Bonanomi S, Dassi M, Buscemi F, Longoni D, Rovelli A, Uderzo C (2000) Infusion-related side-effects in children undergoing autologous hematopoietic stem cell transplantation for acute leukemia. Bone Marrow Transplant 26:116–118
Polchow B, Kebbel K, Schmiedeknecht G, Reichardt A, Henrich W, Hetzer R, Lueders C (2012) Cryopreservation of human vascular umbilical cord cells under good manufacturing practice conditions for future cell banks. J Transl Med 10:98
Rajotte RV, Mazur P (1981) Survival of frozen-thawed fetal rat pancreases as a function of the permeation of dimethylsulfoxide and glycerol, warming rate, and fetal age. Cryobiology 18:17–31
Rall W, Raid D, Polge C (1984) Analysis of slow-freezing injury of mouse embryos by cryomicroscopical and physicochemical methods. Cryobiology 21:106–121
Röllig C, Babatz J, Wagner I, Maiwald A, Schwarze V, Ehninger G, Bornhäuser M (2002) Thawing of cryopreserved mobilized peripheral blood–comparison between waterbath and dry warming device. Cytotherapy 4:551–555
Routledge C, Armitage WJ (2003) Cryopreservation of cornea: a low cooling rate improves functional survival of endothelium after freezing and thawing. Cryobiology 46:277–283
Seki S, Mazur P (2008) Kinetics and activation energy of recrystallization of intracellular ice in mouse oocytes subjected to interrupted rapid cooling. Cryobiology 56:171–180
Seki S, Mazur P (2009) The dominance of warming rate over cooling rate in the survival of mouse oocytes subjected to a vitrification procedure. Cryobiology 59:75–82
Soler AJ, García AJ, Fernández-Santos MR, Esteso MC, Garde JJ (2003) Effects of thawing procedure on postthawed in vitro viability and in vivo fertility of red deer epididymal spermatozoa cryopreserved at −196 °C. J Androl 24:746–756
Souzu H, Mazur P (1978) Temperature dependence of the survival of human erythrocytes frozen slowly in various concentrations of glycerol. Biophys J 23:89–100
Tao J, Du J, Kleinhans FW, Critser ES, Mazur P, Critser JK (1995) The effect of collection temperature, cooling rate and warming rate on chilling injury and cryopreservation of mouse spermatozoa. J Reprod Fertil 104:231–236
Teerds KJ, Rijntjes E, Veldhuizen-Tsoerkan MB, Rommerts FF, de Boer-Brouwer M (2007) The development of rat Leydig cell progenitors in vitro: how essential is luteinising hormones. J Endocrinol 3:579–593
Triana E, Ortega S, Azqueta C, Pomares H, Valdivia E, Duarte R, Massuet L, Martín-Henao GA (2013) Thawing of cryopreserved hematopoietic progenitor cells from apheresis with a new dry-warming device. Transfusion 53:85–90
Vysekantsev IP, Gurina TM, Martsenyuk VF, Petrenko TF, Kudokotseva EV, Koshchiy SV, Groshevoy MI (2005) Probability of lethal damages of cryopreserved biological objects during storage. CryoLetters 26:401–408
Yu I, Songsasen N, Godke RA, Leibo SP (2002) Differences among dogs in response of their spermatozoa to cryopreservation using various cooling and warming rates. Cryobiology 44:62–78
Acknowledgments
The authors would like to thank UNESCO Chair in Cryobiology in the persons of Professor Colin J. Green and Professor Barry J. Fuller for the support of the work and the critical discussion of the results. This work was supported with a special funding by The National Academy of Science, Research topics 0111U001196, 0104U003919; 0110U000404.
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Gurina, T.M., Pakhomov, A.V., Polyakova, A.L. et al. The development of the cell cryopreservation protocol with controlled rate thawing. Cell Tissue Bank 17, 303–316 (2016). https://doi.org/10.1007/s10561-015-9533-6
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DOI: https://doi.org/10.1007/s10561-015-9533-6