Abstract
Structural and functional changes in cytoplasmic membranes and cell organelles play a crucial role in cell damage at low temperatures. These changes are reversible if adequate measures are taken to protect biological membranes against cold-induced injuries. In this study, the possibility of cryopreservation of membrane integrity by long-term storage of samples at low temperatures (−80°C) is demonstrated using differentiated nuclear blood cells as an example. In addition to classical methods, freezing of human leukocyte suspensions was carried out in a novel nontoxic cryoprotecting solution under a newly developed exponential freezing program based on the use of cryoprotectors and repair additives. This program ensures the same cryopreservation effect as the linear program, but is economically more efficient and less labor-consuming. After exponential freezing in a cryoprotecting solution and storage at −80°C, blood leukocytes retain their eosin resistance (91 ± 5% of thawed leukocytes) and phagocytic activity (76.7 ± 14.7% of thawed neutrophils) for 180 days. The novel technology of cell cryopreservation employing a nontoxic cryoprotecting solution, exponential freezing program, and fast thawing is economically efficient, easy to perform and applicable to storage of any animal cells.
Similar content being viewed by others
References
Belous, A.M. and Bondarenko, V.A., Strukturnye izmeneniya biologicheskikh membran pri okhlazhdenii (Cold-Induced Structural Changes in Biological Membranes), Kiev: Naukova Dumka, 1982.
Farrant, J. and Morris, G.J., Thermal Shock and Dilution Shock as the Causes of Freezing Injury, Cryobiology, 1973, vol. 10, pp. 134–140.
Belous, A.M. and Grishchenko, V.I., Kriobiologiya (Cryobiology), Kiev: Naukova Dumka, 1994.
Levin, S.V., Strukturnye izmeneniya kletochnykh membran (Structural Changes in Cellular Membranes), Leningrad: Nauka, 1976.
Svedentsov, E.P., Poluchenie i konservirovanie kostnogo mozga dlya klinicheskogo primeneniya (Preparation and Cryopreservation of Bone Marrow for Clinical Purposes), Extended Abstract of Doct. Sci. Dissertation, Kirov Research Institute of Hematology and Blood Transfusion, Leningrad, 1987.
Yalensky, A.Y.u., Funktsional'noe sostoyanie trombotsitov, vyshedshikh iz kanabioza pri umerenno nizkoi temperature (Functional State of Platelets after Cryoanabiosis at Moderately Low Temperatures), Extended Abstract of Cand. Sci. Dissertation, Kirov Research Institute of Hematology and Blood Transfusion, Kirov: GOU VPO “KGMA,” 2007.
Gordienko, E.A. and Pushkar', N.S., Fizicheskie osnovy nizkotemperaturnogo konservirovaniya kletochnykh suspenzii (Physical Principles of Cryopreservation of Cell Suspensions), Kiev: Naukova Dumka, 1994.
Kostyaev, A.A., Nizkotemperaturnoe konservirovanie gemopoeticheskikh stvolovykh kletok pri -80°C v rezhime bystrogo dvukhstupenchatogo zamorazhivaniya (Cryopreservation of Hemopoietic Stem Cells at -80°C by Fast Two-Phase Freezing), Extended Abstract of Doct. Sci. Dissertation, Russian Research Institute of Hematology and Transfusiology, Saint-Petersburg, 2003.
Svedentsov, E.P., Tumanova, T.V., Shcheglova, O.O., Devetyarova, O.N., and Stepanova, E.S., Freezing Leukocytes to -40°C and -20°C under an Exponential Programme with Original Cryopreservatives, Internat. J. Refrigeration, 2006, vol. 29, pp. 369–373.
Krovezameniteli, konservanty krovi i kostnogo mozga (Blood Substitutes and Preservatives of Blood and Bone Marrow), Khlyabich, G.N., Ed., Moscow: Meditsina, 1997, pp. 160–161.
Crowley, J.P., Rene, A., and Valeri, C.R., The Recovery, Structure, and Function of Human Blood Leukocytes after Freeze-Preservation, Cryobiology, 1974, vol. 11, pp. 395–409.
Lionetti, F.J., Hunt, S.M., Gore, J.M., and Curby, W.A., Cryopreservation of Human Granulocytes, Cryobiology, 1975, vol. 12, pp. 181–191.
Boonlayangoor, P., Telischi, M., Boonlayangoor, S., Sinclair, T.F., and Millhouse, E.W., Cryopreservation of Human Granulocytes: Study of Granulocyte Function and Ultrastructure, Blood, 1980, vol. 56, pp. 237–245.
Utemov, S.V., Nizkotemperaturnoe (80°C) konservirovanie leikotsitnykh kontsentratov (Low-Temperature (80°C) Preservation of Leukocyte Concentrates), Extended Abstract of Cand. Sci. (Med.) Dissertation, Kirov Research Institute of Hematology and Blood Transfusion, Saint-Petersburg, 2005.
Higman, M.A., Port, J.D., Beauchamp, N.J., and Chen, A.R., Reversible Leukoencephalopathy Associated with Re-Infusion of DMSO Preserved Stem Cells, Bone Marrow Transplant., 2000, vol. 26, pp. 797–800.
Windrum, P., Morris, T.C., Drake, M.B., et al., EBMT Chronic Leukaemia Working Party Complications Subcommittee. Variation in Dimethyl Sulfoxide Use in Stem Cell Transplantation: A Survey of EBMT Centres, Bone Marrow Transplant., 2005, vol. 36, pp. 601–603.
Svedentsov, E.P., Arkhieerev, V.P., Utemov, S.V., et al., Fiziko-khimicheskie i toksiko farmakologicheskie kharakteristiki zhidkoi formy krioprotektora A-378 (Phys ico-Chemical and Toxico-Pharmacological Characteristics of the Liquid Form of Cryoprotector A-378), in Sbornik nauchnykh trudov: aktual'nye voprosy transfuzionnoi I klinicheskoi meditsiny (Topical Problems of Transfusion And Clinical Medicine: Collection of Articles), Kirov, 1997, p. 34.
Svedentsov, E.P., Tumanova, T.V., Devetyarova, O.N., Shcheglova, O.O., Utemov, S.V., and Kostyaev, A.A., Krioprotektornyi rastvor dlya zamorazhivania leikotsi tov pri nizkoi temperature (A Cryoprotector Solution for Freezing Leukocytes at Low Temperatures), RF Patent no. 2290808, Byull. Izobret., 2007, no. 1, p. 8.
Goldberg, E.D., Dygai, A.M., and Shakhov, V.P., Metody kul'tury tkani v gematologii (Methods of Tissue Culture in Hematology), Tomsk: Izdatel'stvo Tomskogo Gosudarstvennogo Universiteta, 1992, pp. 224–226.
Saran, M., To What End Does Nature Produce Superoxide? NADPH Oxidase as an Autocrine Modifier of Membrane Phospholipids Generating Paracrine Lipid Messengers, Free Radic. Res., 2003, vol. 37, pp. 1045–1059.
Potapova, S.G., Khrustikov, V.S., Demidova, N.V., et al., Izuchenie poglotitel'noi sposobnosti neitrofilov krovi s ispol'zovaniem inertnykh chastits lateksa (Studies into Absorptive Capacity of Blood Neutrophils Using Inert Latex Particles), Problemy Gamatologii i Perelivaniya Krovi (Rus.), 1977, no. 9, pp. 58–59.
Slavinsky, A.A. and Nikitina, G.V., Cytochemical Detection of Cationic Proteins in Blood Granulocytes with Amido Black 10B for Visualization and Computer Imag ing, Klinicheskaya Laboratornaya Diagnostika (Rus.), 1999, no. 2, pp. 35–37.
Glants, S., Medico-biologicheskaya statistika (Medicobiological Statistics), Moscow: Praktika, 1998.
Lukyanova, L.D. and Romanova, V.E., Some Peculiarities of Anti-hypoxic Action of Mexidol Linked to Its Specific Effect on Energy Metabolism, Khem.-Pharmakol. Zhumal (Rus.), 1990, no. 38, pp. 9–11.
Devyatkina, T.A., Lutsenko, R.V., Vazhnichaya, E.M., and Smirnov, L.D., Effect of Mexidol and Its Structural Components on Carbohydrate Content and Lipid Peroxidation in Acute Stress, Voprosy Meditsinskoi Khimii (Rus.), 1999, vol. 45, no. 3, pp. 246–249.
Mashkovsky, M.D., Lekarstvennye sredstva (Medicinal Drugs), Moscow: Novaya Volna, 2002, pp. 186–187.
Men'shchikova, E.B., Lankin, V.Z., Bondar', I.A., Krugovykh, N.F., and Trufakin, V.A., Okislitel'nyi stress. Prooksidanty i antioksidanty (Oxidative Stress: Prooxidants and Antioxidants), Moscow: Slovo, 2006, pp. 22–72.
Alexeev, N.A., Klinicheskie aspekty leikopenii, neitropenii i funktsional'nykh narushenii neitrofilov (Clinical Aspects of Leukopenias, Neutropenias, and Functional Disturbances in Neutrophils), St. Petersburg: Foliant, 2002, pp. 147–150.
Author information
Authors and Affiliations
Corresponding author
Additional information
Original Russian Text © E.P. Svedentsov, T.V. Tumanova, A.N. Khudyakov, O.O. Zaytseva, O.N. Solomina, S.V. Utemov, F.S. Sherstnev, 2008, published in Biologicheskie Membrany, 2008, Vol. 25, No. 1, pp. 18–24.
Rights and permissions
About this article
Cite this article
Svedentsov, E.P., Tumanova, T.V., Khudyakov, A.N. et al. Cryopreservation of functionally active blood nuclear cell membranes at −80°C. Biochem. Moscow Suppl. Ser. A 2, 19–25 (2008). https://doi.org/10.1007/s11827-008-1004-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11827-008-1004-9