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Preservation Protocols for Human Adipose Tissue-Derived Adult Stem Cells

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Adipose-Derived Stem Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 702))

Abstract

The development of simple but effective storage protocols for adult stem cells will greatly enhance their use and utility in tissue-engineering applications. There are three primary storage techniques, freezing (cryopreservation), drying (anhydrobiosis), and freeze drying (lyophilization), each with its own advantages and disadvantages. Cryopreservation has shown the most promise but is a fairly complex process, necessitating the use of chemicals called cryoprotective agents (CPAs), freezing equipment, and obviously, storage in liquid nitrogen. Preservation by desiccation is an alternative that attempts to reproduce a naturally occurring preservative technique, namely, the phenomenon of anhydrobiosis and requires the use of high (and possibly, toxic) concentration of CPAs as well as disaccharides (sugars). Lyophilization works by first cryopreserving (freezing) the material and then desiccating (drying) it by the process of sublimation or the conversion of ice (solid) to water vapor (gas phase). The purpose of this chapter is to present a general overview of these storage techniques and the optimal protocols/results obtained in our laboratory for long-term storage of adult stem cells using freezing storage and drying storage.

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References

  1. De Ugarte, D. A., Morizono, K., Elbarbary, A., Alfonso, Z., Zuk, P. A., Zhu, M., Dragoo, J. L., Ashjian, P., Thomas, B., Benhaim, P., Chen, I., Fraser, J., and Hedrick, M. H. (2003) Comparison of multi-lineage cells from human adipose tissue and bone marrow, Cells Tissues Organs 174, 101–109.

    Article  PubMed  Google Scholar 

  2. Kang, S. K., Putnam, L., Dufour, J., Ylostalo, J., Jung, J. S., and Bunnell, B. A. (2004) Expression of telomerase extends the lifespan and enhances osteogenic differentiation of adipose tissue-derived stromal cells, Stem Cells 22, 1356–1372.

    Article  PubMed  CAS  Google Scholar 

  3. Mitchell, J. B., McIntosh, K., Zvonic, S., Garrett, S., Floyd, Z. E., Kloster, A., Di Halvorsen, Y., Storms, R. W., Goh, B., Kilroy, G., Wu, X., and Gimble, J. M. (2006) Immunophenotype of human adipose-derived cells: temporal changes in stromal-associated and stem cell-associated markers, Stem Cells 24, 376–385.

    Article  PubMed  Google Scholar 

  4. Rubio, D., Garcia-Castro, J., Martin, M. C., de la Fuente, R., Cigudosa, J. C., Lloyd, A. C., and Bernad, A. (2005) Spontaneous human adult stem cell transformation, Cancer Res 65, 3035–3039.

    PubMed  CAS  Google Scholar 

  5. Wall, M. E., Bernacki, S. H., and Loboa, E. G. (2007) Effects of serial passaging on the adipogenic and osteogenic differentiation potential of adipose-derived human mesenchymal stem cells, Tissue Eng 13, 1291–1298.

    Article  PubMed  CAS  Google Scholar 

  6. Gimble, J. M., Katz, A. J., and Bunnell, B. A. (2007) Adipose-derived stem cells for regenerative medicine, Circ Res 100, 1249–1260.

    Article  PubMed  CAS  Google Scholar 

  7. Zhang, F. B., Li, L., Fang, B., Zhu, D. L., Yang, H. T., and Gao, P. J. (2005) Passage-restricted differentiation potential of mesenchymal stem cells into cardiomyocyte-like cells, Biochem Biophys Res Commun 336, 784–792.

    Article  PubMed  CAS  Google Scholar 

  8. Tsutsumi, S., Shimazu, A., Miyazaki, K., Pan, H., Koike, C., Yoshida, E., Takagishi, K., and Kato, Y. (2001) Retention of multilineage differentiation potential of mesenchymal cells during proliferation in response to FGF, Biochem Biophys Res Commun 288, 413–419.

    Article  PubMed  CAS  Google Scholar 

  9. Zeng, X., and Rao, M. S. (2007) Human embryonic stem cells: long term stability, absence of senescence and a potential cell source for neural replacement, Neuroscience 145, 1348–1358.

    Article  PubMed  CAS  Google Scholar 

  10. Zimmermann, S., and Martens, U. M. (2008) Telomeres, senescence, and hematopoietic stem cells, Cell Tissue Res 331, 79–90.

    Article  PubMed  Google Scholar 

  11. Piacibello, W., Gammaitoni, L., and Pignochino, Y. (2005) Proliferative senescence in hematopoietic stem cells during ex-vivo expansion, Folia Histochem Cytobiol 43, 197–202.

    PubMed  CAS  Google Scholar 

  12. Pegg, D. E. (1987) Mechanisms of freezing damage, Symp Soc Exp Biol 41, 363–378.

    PubMed  CAS  Google Scholar 

  13. Day, J. G. (2007) Cryopreservation of microalgae and cyanobacteria, Methods Mol Biol 368, 141–151.

    Article  PubMed  CAS  Google Scholar 

  14. Pegg, D. E. (2007) Principles of cryopreservation, Methods Mol Biol 368, 39–57.

    Article  PubMed  CAS  Google Scholar 

  15. Bischof, J. C., and Rubinsky, B. (1993) Large ice crystals in the nucleus of rapidly frozen liver cells, Cryobiology 30, 597–603.

    Article  PubMed  CAS  Google Scholar 

  16. Toner, M., Cravalho, E. G., Stachecki, J., Fitzgerald, T., Tompkins, R. G., Yarmush, M. L., and Armant, D. R. (1993) Nonequilibrium freezing of one-cell mouse embryos. Memb­rane integrity and developmental potential, Biophys J 64, 1908–1921.

    Article  PubMed  CAS  Google Scholar 

  17. Venkatasubramanian, R. T., Grassl, E. D., Barocas, V. H., Lafontaine, D., and Bischof, J. C. (2006) Effects of freezing and cryopreservation on the mechanical properties of arteries, Ann Biomed Eng 34, 823–832.

    Article  PubMed  Google Scholar 

  18. Schill, R. O., Mali, B., Dandekar, T., Schnolzer, M., Reuter, D., and Frohme, M. (2009) Molecular mechanisms of tolerance in tardigrades: new perspectives for preservation and stabilization of biological material, Biotechnol Adv 27, 348–352.

    Article  PubMed  CAS  Google Scholar 

  19. Franca, M. B., Panek, A. D., and Eleutherio, E. C. (2007) Oxidative stress and its effects during dehydration, Comp Biochem Physiol A Mol Integr Physiol 146, 621–631.

    Article  PubMed  CAS  Google Scholar 

  20. Liang, Y., and Sun, W. Q. (2002) Rate of dehydration and cumulative desiccation stress interacted to modulate desiccation tolerance of recalcitrant cocoa and ginkgo embryonic tissues, Plant Physiol 128, 1323–1331.

    Article  PubMed  CAS  Google Scholar 

  21. Hoekstra, F. A., Golovina, E. A., and Buitink, J. (2001) Mechanisms of plant desiccation tolerance, Trends Plant Sci 6, 431–438.

    Article  PubMed  CAS  Google Scholar 

  22. Allison, S. D., Randolph, T. W., Manning, M. C., Middleton, K., Davis, A., and Carpenter, J. F. (1998) Effects of drying methods and additives on structure and function of actin: mechanisms of dehydration-induced damage and its inhibition, Arch Biochem Biophys 358, 171–181.

    Article  PubMed  CAS  Google Scholar 

  23. Setlow, P. (1995) Mechanisms for the prevention of damage to DNA in spores of Bacillus species, Annu Rev Microbiol 49, 29–54.

    Article  PubMed  CAS  Google Scholar 

  24. Crowe, L. M., and Crowe, J. H. (1992) Anhydrobiosis: a strategy for survival, Adv Space Res 12, 239–247.

    Article  PubMed  CAS  Google Scholar 

  25. Clegg, J. S., Seitz, P., Seitz, W., and Hazlewood, C. F. (1982) Cellular responses to extreme water loss: the water-replacement hypothesis, Cryobiology 19, 306–316.

    Article  PubMed  CAS  Google Scholar 

  26. Yu, J., and Anchordoquy, T. J. (2009) Synergistic effects of surfactants and sugars on lipoplex stability during freeze-drying and rehydration, J Pharm Sci 98, 3319–3328.

    Article  PubMed  CAS  Google Scholar 

  27. Jiang, W., and Schwendeman, S. P. (2008) Stabilization of tetanus toxoid encapsulated in PLGA microspheres, Mol Pharm 5, 808–817.

    Article  PubMed  CAS  Google Scholar 

  28. Santivarangkna, C., Kulozik, U., and Foerst, P. (2008) Inactivation mechanisms of lactic acid starter cultures preserved by drying processes, J Appl Microbiol 105, 1–13.

    Article  PubMed  CAS  Google Scholar 

  29. Stoner, G. D., Chen, T., Kresty, L. A., Aziz, R. M., Reinemann, T., and Nines, R. (2006) Protection against esophageal cancer in rodents with lyophilized berries: potential mechanisms, Nutr Cancer 54, 33–46.

    Article  PubMed  CAS  Google Scholar 

  30. Hansen, R. K., Zhai, S., Skepper, J. N., Johnston, M. D., Alpar, H. O., and Slater, N. K. (2005) Mechanisms of inactivation of HSV-2 during storage in frozen and lyophilized forms, Biotechnol Prog 21, 911–917.

    Article  PubMed  CAS  Google Scholar 

  31. Arakawa, T., Prestrelski, S. J., Kenney, W. C., and Carpenter, J. F. (2001) Factors affecting short-term and long-term stabilities of proteins, Adv Drug Deliv Rev 46, 307–326.

    Article  PubMed  CAS  Google Scholar 

  32. Milano, M. T., and Bernhard, W. A. (1999) The influence of packing on free radical yields in solid-state DNA: film compared to lyophilized frozen solution, Radiat Res 152, 196–201.

    Article  PubMed  CAS  Google Scholar 

  33. Kreilgaard, L., Frokjaer, S., Flink, J. M., Randolph, T. W., and Carpenter, J. F. (1998) Effects of additives on the stability of recombinant human factor XIII during freeze-drying and storage in the dried solid, Arch Biochem Biophys 360, 121–134.

    Article  PubMed  CAS  Google Scholar 

  34. Fuller, R., and Devireddy, R. V. (2008) The effect of two different freezing methods on the immediate post-thaw membrane integrity of adipose tissue derived stem, Int J Heat Mass Transfer 51, 5650–5654.

    Article  CAS  Google Scholar 

  35. Goh, B. C., Thirumala, S., Kilroy, G., Devireddy, R. V., and Gimble, J. M. (2007) Cryopreservation characteristics of adipose-derived stem cells: maintenance of differentiation potential and viability, J Tissue Eng Regen Med 1, 322–324.

    Article  PubMed  CAS  Google Scholar 

  36. Thirumala, S., Gimble, J., and Devireddy, R. V. (2010) Evaluation of methylcellulose and dimethylsulfoxide as the cryoprotectants in a serum free freezing media for cryopreservation of adipose derived adult stem cells, Stem Cells Dev 19, 513–522.

    Article  PubMed  CAS  Google Scholar 

  37. Thirumala, S., Gimble, J., and Devireddy, R. V. (2010) Cryopreservation of stromal vascular fraction of adipose tissue in a serum free freezing media, J Tissue Eng Regen Med 4, 224–232.

    Article  PubMed  CAS  Google Scholar 

  38. Thirumala, S., Wu, X., Gimble, J., and Devireddy, R. V. (2010) Evaluation of polyvinylpyrollidone (PVP) as a cryoprotectant for adipose derived adult stem cells (ASCs), Tissue Eng Part C Methods 16, 783–792.

    Article  PubMed  CAS  Google Scholar 

  39. Mittal, S., and Devireddy, R. V. (2008) Desiccation tolerance of adult stem cells in the presence of trehalose and glycerol, Open Biotechnol J 2, 211–218.

    Article  CAS  Google Scholar 

  40. Devireddy, R. V., Thirumala, S., and Gimble, J. M. (2005) Cellular response of adipose derived passage-4 adult stem cells to freezing stress, J Biomech Eng 127, 1081–1086.

    Article  PubMed  Google Scholar 

  41. Thirumala, S., Gimble, J. M., and Devireddy, R. V. (2005) Transport phenomena during freezing of adipose tissue derived adult stem cells, Biotechnol Bioeng 92, 372–383.

    Article  PubMed  CAS  Google Scholar 

  42. Thirumala, S., Zvonic, S., Floyd, E., Gimble, J. M., and Devireddy, R. V. (2005) Effect of various freezing parameters on the immediate post-thaw membrane integrity of adipose tissue derived adult stem cells, Biotechnol Prog 21, 1511–1524.

    Article  PubMed  CAS  Google Scholar 

  43. Mazur, P. (1984) Freezing of living cells: mechanisms and implications, Am J Physiol 247, C125–C142.

    PubMed  CAS  Google Scholar 

  44. Polge, C., Smith, A. U., and Parkes, A. S. (1949) Revival of spermatozoa after vitrification and dehydration at low temperatures, Nature 164, 666.

    Article  PubMed  CAS  Google Scholar 

  45. McGrath, J. J. (1997) Quantitative measurement of cell membrane transport: technology and applications, Cryobiology 34, 315–334.

    Article  PubMed  CAS  Google Scholar 

  46. Anchordoguy, T. J., Cecchini, C. A., Crowe, J. H., and Crowe, L. M. (1991) Insights into the cryoprotective mechanism of dimethyl sulf­oxide for phospholipid bilayers, Cryobiology 28, 467–473.

    Article  PubMed  CAS  Google Scholar 

  47. Anchordoguy, T., Carpenter, J. F., Loomis, S. H., and Crowe, J. H. (1988) Mechanisms of interaction of amino acids with phospholipid bilayers during freezing, Biochim Biophys Acta 946, 299–306.

    Article  PubMed  CAS  Google Scholar 

  48. Yu, Z. W., and Quinn, P. J. (1994) Dimethyl sulphoxide: a review of its applications in cell biology, Biosci Rep 14, 259–281.

    Article  PubMed  CAS  Google Scholar 

  49. Fahy, G. M., Lilley, T. H., Linsdell, H., Douglas, M. S., and Meryman, H. T. (1990) Cryoprotectant toxicity and cryoprotectant toxicity reduction: in search of molecular mechanisms, Cryobiology 27, 247–268.

    Article  PubMed  CAS  Google Scholar 

  50. Fahy, G. M. (2010) Cryoprotectant toxicity neutralization, Cryobiology 60, S45–S53.

    Article  PubMed  CAS  Google Scholar 

  51. Fahy, G. M., Wowk, B., Wu, J., and Paynter, S. (2004) Improved vitrification solutions based on the predictability of vitrification solution toxicity, Cryobiology 48, 22–35.

    Article  PubMed  CAS  Google Scholar 

  52. Fahy, G. M. (1986) The relevance of cryoprotectant “toxicity” to cryobiology, Cryobiology 23, 1–13.

    Article  PubMed  CAS  Google Scholar 

  53. Farrant, J. (1969) Is there a common mechanism of protection of living cells by polyvinylpyrrolidone and glycerol ding freezing?, Nature 222, 1175–1176.

    Article  PubMed  CAS  Google Scholar 

  54. Hey, J. M., and MacFarlane, D. R. (1998) Crystallization of ice in aqueous solutions of glycerol and dimethyl sulfoxide 2: ice crystal growth kinetics, Cryobiology 37, 119–130.

    Article  PubMed  CAS  Google Scholar 

  55. Shaw, J. M., Kuleshova, L. L., MacFarlane, D. R., and Trounson, A. O. (1997) Vitrification properties of solutions of ethylene glycol in saline containing PVP, Ficoll, or dextran, Cryobiology 35, 219–229.

    Article  PubMed  CAS  Google Scholar 

  56. Alapati, R., Stout, M., Saenz, J., Gentry, G. T., Jr., Godke, R. A., and Devireddy, R. V. (2009) Comparison of the permeability properties and post-thaw motility of ejaculated and epididymal bovine spermatozoa, Cryobiology 59, 164–170.

    Article  PubMed  CAS  Google Scholar 

  57. Hagiwara, M., Choi, J. H., Devireddy, R. V., Roberts, K. P., Wolkers, W. F., Makhlouf, A., and Bischof, J. C. (2009) Cellular biophysics during freezing of rat and mouse sperm predicts postthaw motility, Biol Reprod 81, 700–706.

    Article  PubMed  CAS  Google Scholar 

  58. Alapati, R., Goff, K., Kubisch, H. M., and Devireddy, R. V. (2008) Water transport in epididymal and ejaculated rhesus monkey (Macaca mulatta) sperm during freezing, Cryobiology 57, 182–185.

    Article  PubMed  CAS  Google Scholar 

  59. Kardak, A., Leibo, S. P., and Devireddy, R. (2007) Membrane transport properties of equine and macaque ovarian tissues frozen in mixtures of dimethylsulfoxide and ethylene glycol, J Biomech Eng 129, 688–694.

    Article  PubMed  CAS  Google Scholar 

  60. Li, G., Saenz, J., Godke, R. A., and Devireddy, R. V. (2006) Effect of glycerol and cholesterol-loaded cyclodextrin on freezing-induced water loss in bovine spermatozoa, Reproduction 131, 875–886.

    Article  PubMed  CAS  Google Scholar 

  61. Pinisetty, D., Huang, C., Dong, Q., Tiersch, T. R., and Devireddy, R. V. (2005) Subzero water permeability parameters and optimal freezing rates for sperm cells of the southern platyfish, Xiphophorus maculatus, Cryobiology 50, 250–263.

    Article  PubMed  CAS  Google Scholar 

  62. Thirumala, S., Huang, C., Dong, Q., Tiersch, T. R., and Devireddy, R. V. (2005) A theoretically estimated optimal cooling rate for the cryopreservation of sperm cells from a live-bearing fish, the green swordtail Xiphophorus helleri, Theriogenology 63, 2395–2415.

    Article  PubMed  Google Scholar 

  63. Meryman, H. T. (1974) Freezing injury and its prevention in living cells, Annu Rev Biophys Bioeng 3, 341–363.

    Article  PubMed  CAS  Google Scholar 

  64. Meryman, H. T. (1956) Mechanics of freezing in living cells and tissues, Science 124, 515–521.

    Article  PubMed  CAS  Google Scholar 

  65. Meryman, H. T. (2007) Cryopreservation of living cells: principles and practice, Transfusion 47, 935–945.

    Article  PubMed  CAS  Google Scholar 

  66. Connor, W., and Ashwood-Smith, M. J. (1973) Cryoprotection of mammalian cells in tissue culture with polymers; possible mechanisms, Cryobiology 10, 488–496.

    Article  PubMed  CAS  Google Scholar 

  67. Holt, W. V., and North, R. D. (1994) Effects of temperature and restoration of osmotic equilibrium during thawing on the induction of plasma membrane damage in cryopreserved ram spermatozoa, Biol Reprod 51, 414–424.

    Article  PubMed  CAS  Google Scholar 

  68. Barrios, B., Perez-Pe, R., Gallego, M., Tato, A., Osada, J., Muino-Blanco, T., and Cebrian-Perez, J. A. (2000) Seminal plasma proteins revert the cold-shock damage on ram sperm membrane, Biol Reprod 63, 1531–1537.

    Article  PubMed  CAS  Google Scholar 

  69. Rodgers, F. G., and Davey, M. R. (1982) Ultrastructure of the cell envelope layers and surface details of Legionella pneumophila, J Gen Microbiol 128, 1547–1557.

    PubMed  CAS  Google Scholar 

  70. De Leeuw, F. E., De Leeuw, A. M., Den Daas, J. H., Colenbrander, B., and Verkleij, A. J. (1993) Effects of various cryoprotective agents and membrane-stabilizing compounds on bull sperm membrane integrity after cooling and freezing, Cryobiology 30, 32–44.

    Article  PubMed  Google Scholar 

  71. Barnard, T. (1987) Rapid freezing techniques and cryoprotection of biomedical specimens, Scanning Microsc 1, 1217–1224.

    PubMed  CAS  Google Scholar 

  72. Hirsh, A. G., Williams, R. J., and Meryman, H. T. (1985) A novel method of natural cryoprotection: intracellular glass formation in deeply frozen populus, Plant Physiol 79, 41–56.

    Article  PubMed  CAS  Google Scholar 

  73. Rall, W. F., Mazur, P., and McGrath, J. J. (1983) Depression of the ice-nucleation temperature of rapidly cooled mouse embryos by glycerol and dimethyl sulfoxide, Biophys J 41, 1–12.

    Article  PubMed  CAS  Google Scholar 

  74. Elliott, R., Szleifer, I., and Schick, M. (2007) A microscopic model calculation of the phase diagram of ternary mixtures of cholesterol and saturated and unsaturated phospholipids, Methods Mol Biol 398, 303–317.

    Article  PubMed  CAS  Google Scholar 

  75. Fahy, G. M. (1980) Analysis of “solution effects” injury. Equations for calculating phase diagram information for the ternary systems NaCl-dimethylsulfoxide-water and NaCl-glycerol-water, Biophys J 32, 837–850.

    Article  PubMed  CAS  Google Scholar 

  76. Guignon, B., Aparicio, C., Otero, L., and Sanz, P. D. (2009) Prediction of ice content in biological model solutions when frozen under high pressure, Biotechnol Prog 25, 454–460.

    Article  PubMed  CAS  Google Scholar 

  77. Kleinhans, F. W., and Mazur, P. (2007) Comparison of actual vs. synthesized ternary phase diagrams for solutes of cryobiological interest, Cryobiology 54, 212–222.

    Article  PubMed  CAS  Google Scholar 

  78. Morris, G. J., Goodrich, M., Acton, E., and Fonseca, F. (2006) The high viscosity encountered during freezing in glycerol solutions: effects on cryopreservation, Cryobiology 52, 323–334.

    Article  PubMed  CAS  Google Scholar 

  79. Watson, P. F., and Duncan, A. E. (1988) Effect of salt concentration and unfrozen water fraction on the viability of slowly frozen ram spermatozoa, Cryobiology 25, 131–142.

    Article  PubMed  CAS  Google Scholar 

  80. Rall, W. F., Mazur, P., and Souzu, H. (1978) Physical-chemical basis of the protection of slowly frozen human erythrocytes by glycerol, Biophys J 23, 101–120.

    Article  PubMed  CAS  Google Scholar 

  81. Shepard, M. L., Goldston, C. S., and Cocks, F. H. (1976) The H2O-NaCl-glycerol phase diagram and its application in cryobiology, Cryobiology 13, 9–23.

    Article  PubMed  CAS  Google Scholar 

  82. Wolkers, W. F., Tablin, F., and Crowe, J. H. (2002) From anhydrobiosis to freeze-drying of eukaryotic cells, Comp Biochem Physiol A Mol Integr Physiol 131, 535–543.

    Article  PubMed  Google Scholar 

  83. Crowe, L. M. (2002) Lessons from nature: the role of sugars in anhydrobiosis, Comp Biochem Physiol A Mol Integr Physiol 131, 505–513.

    Article  PubMed  Google Scholar 

  84. Crowe, J. H., Carpenter, J. F., and Crowe, L. M. (1998) The role of vitrification in anhydrobiosis, Annu Rev Physiol 60, 73–103.

    Article  PubMed  CAS  Google Scholar 

  85. Crowe, J. H., Hoekstra, F. A., and Crowe, L. M. (1992) Anhydrobiosis, Annu Rev Physiol 54, 579–599.

    Article  PubMed  CAS  Google Scholar 

  86. Crowe, J. H., and Crowe, L. M. (1982) Induction of anhydrobiosis: membrane changes during drying, Cryobiology 19, 317–328.

    Article  PubMed  CAS  Google Scholar 

  87. Leslie, S. B., Israeli, E., Lighthart, B., Crowe, J. H., and Crowe, L. M. (1995) Trehalose and sucrose protect both membranes and proteins in intact bacteria during drying, Appl Environ Microbiol 61, 3592–3597.

    PubMed  CAS  Google Scholar 

  88. Leslie, S. B., Teter, S. A., Crowe, L. M., and Crowe, J. H. (1994) Trehalose lowers membrane phase transitions in dry yeast cells, Biochim Biophys Acta 1192, 7–13.

    Article  PubMed  CAS  Google Scholar 

  89. Guo, N., Puhlev, I., Brown, D. R., Mansbridge, J., and Levine, F. (2000) Trehalose expression confers desiccation tolerance on human cells, Nat Biotechnol 18, 168–171.

    Article  PubMed  CAS  Google Scholar 

  90. Potts, M. (2001) Desiccation tolerance: a simple process? Trends Microbiol 9, 553–559.

    Article  PubMed  CAS  Google Scholar 

  91. Potts, M. (1994) Desiccation tolerance of prokaryotes, Microbiol Rev 58, 755–805.

    PubMed  CAS  Google Scholar 

  92. Conrad, P. B., Miller, D. P., Cielenski, P. R., and de Pablo, J. J. (2000) Stabilization and preservation of Lactobacillus acidophilus in saccharide matrices, Cryobiology 41, 17–24.

    Article  PubMed  CAS  Google Scholar 

  93. Gordon, S. L., Oppenheimer, S. R., Mackay, A. M., Brunnabend, J., Puhlev, I., and Levine, F. (2001) Recovery of human mesenchymal stem cells following dehydration and rehydration, Cryobiology 43, 182–187.

    Article  PubMed  CAS  Google Scholar 

  94. Puhlev, I., Guo, N., Brown, D. R., and Levine, F. (2001) Desiccation tolerance in human cells, Cryobiology 42, 207–217.

    Article  PubMed  CAS  Google Scholar 

  95. Chen, T., Acker, J. P., Eroglu, A., Cheley, S., Bayley, H., Fowler, A., and Toner, M. (2001) Beneficial effect of intracellular trehalose on the membrane integrity of dried mammalian cells, Cryobiology 43, 168–181.

    Article  PubMed  CAS  Google Scholar 

  96. McGinnis, L. K., Zhu, L., Lawitts, J. A., Bhowmick, S., Toner, M., and Biggers, J. D. (2005) Mouse sperm desiccated and stored in trehalose medium without freezing, Biol Reprod 73, 627–633.

    Article  PubMed  CAS  Google Scholar 

  97. Eroglu, A., Bailey, S. E., Toner, M., and Toth, T. L. (2009) Successful cryopreservation of mouse oocytes by using low concentrations of trehalose and dimethylsulfoxide, Biol Reprod 80, 70–78.

    Article  PubMed  CAS  Google Scholar 

  98. Buchanan, S. S., Gross, S. A., Acker, J. P., Toner, M., Carpenter, J. F., and Pyatt, D. W. (2004) Cryopreservation of stem cells using trehalose: evaluation of the method using a human hematopoietic cell line, Stem Cells Dev 13, 295–305.

    Article  PubMed  CAS  Google Scholar 

  99. Erdag, G., Eroglu, A., Morgan, J., and Toner, M. (2002) Cryopreservation of fetal skin is improved by extracellular trehalose, Cryobiology 44, 218–228.

    Article  PubMed  CAS  Google Scholar 

  100. Eroglu, A., Toner, M., and Toth, T. L. (2002) Beneficial effect of microinjected trehalose on the cryosurvival of human oocytes, Fertil Steril 77, 152–158.

    Article  PubMed  Google Scholar 

  101. Eroglu, A., Russo, M. J., Bieganski, R., Fowler, A., Cheley, S., Bayley, H., and Toner, M. (2000) Intracellular trehalose improves the survival of cryopreserved mammalian cells, Nat Biotechnol 18, 163–167.

    Article  PubMed  CAS  Google Scholar 

  102. Bhowmick, S., Zhu, L., McGinnis, L., Lawitts, J., Nath, B. D., Toner, M., and Biggers, J. (2003) Desiccation tolerance of spermatozoa dried at ambient temperature: production of fetal mice, Biol Reprod 68, 1779–1786.

    Article  PubMed  CAS  Google Scholar 

  103. Guilak, F., Lott, K. E., Awad, H. A., Cao, Q., Hicok, K. C., Fermor, B., and Gimble, J. M. (2006) Clonal analysis of the differentiation potential of human adipose-derived adult stem cells, J Cell Physiol 206, 229–237.

    Article  PubMed  CAS  Google Scholar 

  104. DeLany, J. P., Floyd, Z. E., Zvonic, S., Smith, A., Gravois, A., Reiners, E., Wu, X., Kilroy, G., Lefevre, M., and Gimble, J. M. (2005) Proteomic analysis of primary cultures of human adipose-derived stem cells: modulation by Adipogenesis, Mol Cell Proteomics 4, 731–740.

    Article  PubMed  CAS  Google Scholar 

  105. Hicok, K. C., Du Laney, T. V., Zhou, Y. S., Halvorsen, Y. D., Hitt, D. C., Cooper, L. F., and Gimble, J. M. (2004) Human adipose-derived adult stem cells produce osteoid in vivo, Tissue Eng 10, 371–380.

    Article  PubMed  CAS  Google Scholar 

  106. Safford, K. M., Safford, S. D., Gimble, J. M., Shetty, A. K., and Rice, H. E. (2004) Characterization of neuronal/glial differentiation of murine adipose-derived adult stromal cells, Exp Neurol 187, 319–328.

    Article  PubMed  CAS  Google Scholar 

  107. Aust, L., Devlin, B., Foster, S. J., Halvorsen, Y. D., Hicok, K., du Laney, T., Sen, A., Willingmyre, G. D., and Gimble, J. M. (2004) Yield of human adipose-derived adult stem cells from liposuction aspirates, Cytotherapy 6, 7–14.

    Article  PubMed  CAS  Google Scholar 

  108. Awad, H. A., Wickham, M. Q., Leddy, H. A., Gimble, J. M., and Guilak, F. (2004) Chondrogenic differentiation of adipose-derived adult stem cells in agarose, alginate, and gelatin scaffolds, Biomaterials 25, 3211–3222.

    Article  PubMed  CAS  Google Scholar 

  109. Gimble, J. M., and Guilak, F. (2003) Differentiation potential of adipose derived adult stem (ADAS) cells, Curr Top Dev Biol 58, 137–160.

    Article  PubMed  Google Scholar 

  110. Gimble, J., and Guilak, F. (2003) Adipose-derived adult stem cells: isolation, characterization, and differentiation potential, Cytotherapy 5, 362–369.

    Article  PubMed  Google Scholar 

  111. Safford, K. M., Hicok, K. C., Safford, S. D., Halvorsen, Y. D., Wilkison, W. O., Gimble, J. M., and Rice, H. E. (2002) Neurogenic differentiation of murine and human adipose-derived stromal cells, Biochem Biophys Res Commun 294, 371–379.

    Article  PubMed  CAS  Google Scholar 

  112. Gronthos, S., Franklin, D. M., Leddy, H. A., Robey, P. G., Storms, R. W., and Gimble, J. M. (2001) Surface protein characterization of human adipose tissue-derived stromal cells, J Cell Physiol 189, 54–63.

    Article  PubMed  CAS  Google Scholar 

  113. Thirumala, S., Forman, J. M., Monroe, W. T., and Devireddy, R. (2007) Freezing and post-thaw apoptotic behaviour of cells in the presence of palmitoyl nanogold particles, Nanotechnology 18, 195104.

    Article  CAS  Google Scholar 

  114. Rubinsky, B., and Ikeda, M. (1985) A cryomicroscope using directional solidification for the controlled freezing of biological mat, Cryobiology 22, 55–68.

    Article  Google Scholar 

  115. Garner, D. L., and Johnson, L. A. (1995) Viability assessment of mammalian sperm using SYBR-14 and propidium iodide, Biol Reprod 53, 276–284.

    Article  PubMed  CAS  Google Scholar 

  116. Takamatsu, H., and Rubinsky, B. (1999) Viability of deformed cells, Cryobiology 39, 243–251.

    Article  PubMed  CAS  Google Scholar 

  117. Takamatsu, H., Takeya, R., Naito, S., and Sumimoto, H. (2005) On the mechanism of cell lysis by deformation, J Biomech 38, 117–124.

    Article  PubMed  Google Scholar 

  118. Wolfe, J., and Bryant, G. (1999) Freezing, drying, and/or vitrification of membrane-solute-water systems, Cryobiology 39, 103–129.

    Article  PubMed  CAS  Google Scholar 

  119. Rubinsky, B. (2000) Cryosurgery, Annu Rev Biomed Eng 2, 157–187.

    Article  PubMed  CAS  Google Scholar 

  120. Devireddy, R. V., Swanlund, D. J., Roberts, K. P., and Bischof, J. C. (1999) Subzero water permeability parameters of mouse spermatozoa in the presence of extracellular ice and cryoprotective agents, Biol Reprod 61, 764–775.

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The author thanks Dr. Elizabeth Clubb and Dr. James Wade at the Pennington Biomedical Research Center (PBRC) for supplying the liposuction aspirates and their many patients for consenting to participate in this protocol; Marilyn Dietrick of the LSU School of Veterinary Medicine Flow Cytometry Core Facility; Prof. Jeffrey Gimble, Gang Yu, Xiying Wu, of the Stem Cell Biology Laboratory at the Pennington Biomedical Research Center (PBRC), and the clinical nutrition research unit (CNRU) Molecular Mechanism Core at PBRC for their technical assistance. In addition, acknowledgments are also due to Prof. Devireddy’s graduate students, Ryan Fuller (comparing the freezing devices, CRF, and DSS), and Surbhi Mittal (desiccation) for performing the experiments reported in this chapter. This work was supported in part by funding from the Louisiana Board of Regents and the Department of Mechanical Engineering at the Louisiana State University (LSU). S. Thirumala was supported by an Economic Development Assistantship (EDA) and a Dissertation Fellowship awarded by LSU.

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Authors and Affiliations

  1. Bioengineering Laboratory, Department of Mechanical Engineering, Louisiana State University, Baton Rouge, LA, USA

    Ram Devireddy & Sreedhar Thirumala

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  1. Ram Devireddy
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  2. Sreedhar Thirumala
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Correspondence to Ram Devireddy .

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Editors and Affiliations

  1. , Stem Cell Biology, Pennington Biomedical Research Center, Perkins Rd. 6400, Baton Rouge, 70808, Louisiana, USA

    Jeffrey M. Gimble

  2. School of Medicine, Department of Pharmacology, Tulane University, Tulane Ave. 1430, New Orleans, 70112-2699, Louisiana, USA

    Bruce A. Bunnell

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Devireddy, R., Thirumala, S. (2011). Preservation Protocols for Human Adipose Tissue-Derived Adult Stem Cells. In: Gimble, J., Bunnell, B. (eds) Adipose-Derived Stem Cells. Methods in Molecular Biology, vol 702. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61737-960-4_27

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  • DOI: https://doi.org/10.1007/978-1-61737-960-4_27

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61737-959-8

  • Online ISBN: 978-1-61737-960-4

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