Hostname: page-component-586b7cd67f-rdxmf Total loading time: 0 Render date: 2024-11-30T20:12:53.925Z Has data issue: false hasContentIssue false
  • English
  • Français

Production of bovine cloned embryos with donor cells frozen at a slow cooling rate in a conventional freezer (−20 °C)

Published online by Cambridge University Press:  08 June 2009

Liliana Chacón ,
Martha C. Gómez ,
Jill A. Jenkins ,
Stanley P. Leibo ,
Gemechu Wirtu ,
Betsy L. Dresser  and
C. Earle Pope
Liliana Chacón
Affiliation:
School of Veterinary Medicine, Colombian National University, Bogotá, Colombia. Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA. Department of Biological Sciences, University of New Orleans, New Orleans, Louisiana, USA.
Martha C. Gómez*
Affiliation:
Audubon Center for Research of Endangered Species, 14001 River Road, New Orleans, Louisiana 70131, USA. Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA. Institute of Genetics, Colombian National University, Bogotá, Colombia.
Jill A. Jenkins
Affiliation:
National Wetlands Research Center, US Geological Survey, Lafayette, Louisiana, USA.
Stanley P. Leibo
Affiliation:
Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA. Department of Biological Sciences, University of New Orleans, New Orleans, Louisiana, USA.
Gemechu Wirtu
Affiliation:
Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA.
Betsy L. Dresser
Affiliation:
Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA. Department of Biological Sciences, University of New Orleans, New Orleans, Louisiana, USA.
C. Earle Pope
Affiliation:
Audubon Center for Research of Endangered Species, New Orleans, Louisiana, USA.
*
All correspondence to: Martha C. Gómez. Audubon Center for Research of Endangered Species, 14001 River Road, New Orleans, Louisiana 70131, USA. Tel: +1 504 398 3159. Fax: +1 504 391 7707. e-mail: [email protected]
Get access Rights & Permissions [Opens in a new window]

Summary

Usually, fibroblasts are frozen in dimethyl sulphoxide (DMSO, 10% v/v) at a cooling rate of 1 °C/min in a low-temperature (−80 °C) freezer (LTF) before storage in liquid nitrogen (LN2); however, a LTF is not always available. The purpose of the present study was to evaluate apoptosis and viability of bovine fibroblasts frozen in a LTF or conventional freezer (CF; −20 °C) and their subsequent ability for development to blastocyst stage after fusion with enucleated bovine oocytes. Percentages of live cells frozen in LTF (49.5%) and CF (50.6%) were similar, but significantly less than non-frozen control (88%). In both CF and LTF, percentages of live apoptotic cells exposed to LN2 after freezing were lower (4% and 5%, respectively) as compared with unexposed cells (10% and 18%, respectively). Cells frozen in a CF had fewer cell doublings/24 h (0.45) and required more days (9.1) to reach 100% confluence at the first passage (P) after thawing and plating as compared with cells frozen in a LTF (0.96 and 4.0 days, respectively). Hypoploidy at P12 was higher than at P4 in cells frozen in either a CF (37.5% vs. 19.2%) or in a LTF (30.0% vs. 15.4%). A second-generation cryo-solution reduced the incidence of necrosis (29.4%) at 0 h after thawing as compared with that of a first generation cryo-solution (DMEM + DMSO, 60.2%). The percentage of apoptosis in live cells was affected by cooling rate (CF = 1.9% vs. LFT = 0.7%). Development of bovine cloned embryos to the blastocyst stage was not affected by cooling rate or freezer type.

Keywords

BovineCloningConventional FreezerFibroblasts−20 °C
Type
Research Article
Information
Zygote , Volume 17 , Issue 4 , November 2009 , pp. 341 - 351
Copyright
Copyright © Cambridge University Press 2009

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Adams, R.M., Wang, M., Crane, A.M., Brown, B., Darlington, G.J. & Ledley, F.D. (1995). Effective cryopreservation and long-term storage of primary human hepatocytes with recovery of viability, differentiation and replicative potential. Cell Transplant 4, 579–86.CrossRefGoogle ScholarPubMed
Baust, J.M., Baust, J.G. & Van Buskirk, R. (1998). Cryopreservation outcome is enhanced by intracellular type medium and inhibition of apoptosis. Cryobiology 37, 410–1.Google Scholar
Baust, J.M., Van Buskirk, R. & Baust, J.G. (2000a). Cryopreservation-induced apoptotic gene activation. Cryobiology 41, 338 [abstract].Google Scholar
Baust, J.M., Van Buskirk, R. & Baust, J.G. (2000b). Cell viability improves following inhibition of cryopreservation-induced apoptosis. In Vitro Cell. Dev. Biol. Anim. 36, 262–70.2.0.CO;2>CrossRefGoogle ScholarPubMed
Baust, J.M., Van Buskirk, R. & Baust, J.G. (2002). Modulation of the cryopreservation cap: elevated survival with reduced dimethyl sulphoxide concentration. Cryobiology 45, 97–108.CrossRefGoogle Scholar
Baust, J.M. (2002). Molecular mechanisms of cellular demise associated with cryopreservation failure. Cell.Preserv. Technol. 1, 17–31.Google Scholar
Bhojwani, S., Vajta, G., Callesen, H., Roschlau, K., Kuwer, A., Becker, F., Alm, H., Torner, H., Kanitz, W. & Poehland, R. (2005). Developmental competence of HMC™ derived bovine cloned embryos obtained from somatic cell nuclear transfer of adult fibroblasts and granulosa cells. J. Reprod. Dev. 51, 465–75.Google Scholar
Coundouris, J.A., Grant, M.H., Engeset, J., Petrie, J.C. & Hawksworth, G.M. (1993). Cryopreservation of human adult hepatocytes for use in drug metabolism and toxicity studies. Xenobiotica 23, 1399–409.CrossRefGoogle ScholarPubMed
De Loecker, W., Koptelov, V.A., Grischenko, V.I. & De Loecker, P. (1998). Effects of cell concentration on viability and metabolic activity during cryopreservation. Cryobiology 37, 103–9.Google Scholar
Dong, Y.J., Bai, X.J., Barisanga, M.D., Matango, N.R. & Suzuki, T. (2003). Production of cloned calves following nuclear transfer with cultured frozen–thawed somatic cells using simple portable CO2 incubator. Theriogenology 59, 246 [abstract].Google Scholar
Du, F., Sung, L.Y., Tian, X.C. & Yang, X. (2002). Differential cytoplast requirement for embryonic and somatic cell nuclear transfer in cattle. Mol. Reprod. Dev. 63, 183–91.CrossRefGoogle ScholarPubMed
Estrada, J., Lee, E. & Piedrahita, J.A. (2006). Cryopreservation of donor cells for nuclear transfer: effect of cell freezing method on the efficiency of somatic cell nuclear transfer. Reprod. Fertil. Dev. 18, 125 [abstract].CrossRefGoogle Scholar
Freshney, R.I. & Freshney, M. (2002). Culture of Epithelial Cells. 2nd edn, p. 461New York: J. Wiley & Sons, Inc.CrossRefGoogle Scholar
Frim, J., Snyder, R.A., McGann, L.E. & Kruuv, J. (1978). Growth kinetics of cells following freezing in liquid nitrogen. Cryobiology 15, 502–16.Google Scholar
Giraldo, A.M., Lynn, J.W., Godke, R.A. & Bondioli, K.R. (2006). Proliferative characteristics and chromosomal stability of bovine donor cells for nuclear transfer. Mol. Reprod. Dev. 73, 1230–8.Google Scholar
Gómez, M.C., Pope, C.E., Lopez, M., Dumas, C., Giraldo, A. & Dresser, B.L. (2006). Chromosomal aneuploidy in African Wildcat somatic cells and cloned embryos. Cloning Stem Cells 8, 69–78.CrossRefGoogle ScholarPubMed
Green, A.L., Wells, D.N. & Oback, B. (2007). Cattle cloned from increasingly differentiated muscle cells. Biol. Reprod. 77, 395–406.CrossRefGoogle ScholarPubMed
Hayes, O., Rodriguez, L. L., Gonzalez, A., Falcon, V., Aguilar, A. & Castro, F.O. (2005). Effect of cryopreservation on fusion efficiency and in vitro development into blastocysts of bovine cell lines used in somatic cell cloning. Zygote 13, 277–82.CrossRefGoogle ScholarPubMed
Hill, J.R., Winger, Q.A., Long, C.R., Looney, C.R., Thompson, J.A. & Westhusin, M.E. (2000). Development rates of male bovine nuclear transfer embryos derived from adult and fetal cells. Biol. Reprod. 62, 1135–40.Google Scholar
Holm, P., Booth, P.J., Schmidt, M.H., Greve, T. & Callesen, H. (1999). High bovine blastocyst development in a static in vitro production system using SOFaa medium supplemented with sodium citrate and myo-inositol with or without serum-proteins. Theriogenology 52, 683–700.CrossRefGoogle ScholarPubMed
Jacobs, P.A., Court, B. & Doll, R. (1961). Distribution of human chromosome counts in relation to age. Nature 191, 1178–80.CrossRefGoogle ScholarPubMed
Karlsson, J.O., Cravalho, E.G., Borel Rinkes, I.H., Tompkins, R.G., Yarmush, M.L. & Toner, M. (1993). Nucleation and growth of ice crystals inside cultured hepatocytes during freezing in the presence of dimethyl sulfoxide. Biophys. J. 65, 2524–36.CrossRefGoogle ScholarPubMed
Kato, Y., Tani, T. & Tsunoda, Y. (2000). Cloning of calves from various somatic cell types of male and female adult, newborn and fetal cows. J. Reprod. Fertil. 120, 231–7.CrossRefGoogle ScholarPubMed
Kerr, J.F., Wyllie, A.H. & Currie, A.R. (1972). Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br. J. Cancer 26, 239–57.CrossRefGoogle ScholarPubMed
Kragh, P.M., Du, Y., Corydon, T.J., Purup, S., Bolund, L. & Vajta, G. (2005). Efficient in vitro production of porcine blastocysts by handmade cloning with a combined electrical and chemical activation. Theriogenology 64, 1536–45.Google Scholar
Kragh, P.M., Vajta, G., Corydon, T.J., Purup, S., Bolund, L. & Callesen, H. (2004). Production of transgenic porcine blastocysts by hand-made cloning. Reprod. Fertil. Dev. 16, 315–8.Google Scholar
Li, J. and Mombaerts, P. (2008). Nuclear transfer-mediated rescue of the nuclear genome of nonviable mouse cells frozen without cryoprotectant. Biol. Reprod. 79, 588–93.Google Scholar
Matsushita, T., Yagi, T., Hardin, J.A., Cragun, J.D., Crow, F.W., Bergen, H.R., Gores, G.J. & Nyberg, S.L. (2003). Apoptotic cell death and function of cryopreserved porcine hepatocytes in a bioartificial liver. Cell Transplant 12, 109–21.Google Scholar
Mazur, P. (1963). Kinetics of water loss from cells at subzero temperatures and the likelihood of intracellular freezing. J. Gen. Physiol. 47, 347–69.CrossRefGoogle ScholarPubMed
Mazur, P. (1984). Freezing of living cells: mechanisms and implications. Am. J. Physiol. Cell. Physiol. 247, C125–C142.CrossRefGoogle ScholarPubMed
Mazur, P. (2004). Principles of cryobiology. In Life in the Frozen State (eds Fuller, B.J., Laneg, N. & Benson, E.E.) pp. 3–65. Boca Raton: CRS Press.CrossRefGoogle Scholar
McGann, L.E. (1979). Optimal temperature ranges for control of cooling rate. Cryobiology 16, 211–6.Google Scholar
Muldrew, K., Acker, J.P., Elliot, J.A.W. & McGann, E. (2004). The water to ice transition: implications for living cells. In Life in the Frozen State (eds , B.J.Fuller, , Lane, N. & Benson, E.E.) pp. 67–108. Boca Raton: CRS Press.Google Scholar
Pegg, D.E. and Diaper, M.P. (1990). Freezing versus vitrification: basic principles. In Cryopreservation and Low Temperature Biology in Blood Transfusion (eds Smit-Sibinga, C.T., Das, P.C. & Meryman, H.T.) pp. 55–69. Dordrecht: Kluwer Academic Publishers.Google Scholar
Polge, C., Smith, A.U. & Parkes, A.S. (1949). Revival of spermatozoa after vitrification and dehydration at low temperatures. Nature 164, 666.CrossRefGoogle ScholarPubMed
Rapatz, G. & Luyet, B. (1968). Combined effects of freezing rates and of various protective agents on the preservation of human erythrocytes. Cryobiology 4, 215–22.Google Scholar
Saksela, E. and Moorhead, P.S. (1963). Aneuploidy in the degenerative phase of serial cultivation of human cell strains. Proc. Natl. Acad. Sci. USA 50, 390–5.Google Scholar
Stroh, C., Cassens, U., Samraj, A.K., Sibrowski, W., Schulze-Osthoff, K. & Los, M. (2002). The role of caspases in cryoinjury: caspase inhibition strongly improves the recovery of cryopreserved hematopoietic and other cells. FASEB J. 16, 1651–3.Google Scholar
Stylianou, J., Vowels, M. & Hadfield, K. (2006). Novel cryoprotectant significantly improves the post-thaw recovery and quality of HSC from CB. Cytotherapy 8, 57–61.CrossRefGoogle ScholarPubMed
Tani, T., Kato, Y. & Tsunoda, Y. (2000). Developmental potential of cumulus cell-derived culture frozen in a quiescent state after nucleus transfer. Theriogenology 53, 1623–9.Google Scholar
Thornberry, N.A. and Lazebnik, Y. (1998). Caspases: enemies within. Science 281, 1312–6.Google Scholar
Vajta, G., Peura, T.T., Holm, P., Paldi, A., Greve, T., Trounson, A.O. & Callesen, H. (2000). New method for culture of zona-included or zona-free embryos: the well of the well (WOW) system. Mol. Reprod. Dev. 55, 256–64.3.0.CO;2-7>CrossRefGoogle ScholarPubMed
Vajta, G., Lewis, I.M., Hyttel, P., Thouas, G.A. & Trounson, A.O. (2001). Somatic cell cloning without micromanipulators. Cloning 3, 89–95.Google Scholar
Vajta, G., Maddox-Hyttel, P., Skou, C.T., Tecirlioglu, R.T., Peura, T.T., Lai, L., Murphy, C.N., Prather, R.S., Kragh, P.M. & Callesen, H. (2005). Highly efficient and reliable chemically assisted enucleation method for handmade cloning in cattle. Reprod. Fertil. Develop. 17, 791–7.Google Scholar
Wakayama, S., Ohta, H., Hikichi, T., Mizutani, E., Iwaki, T., Kanagawa, O. & Wakayama, T. (2008). Production of healthy cloned mice from bodies frozen at –20 °C for 16 years. Proc. Natl. Acad. Sci. USA. 105, 17318–22.CrossRefGoogle Scholar
Wells, D.N., Misica, P.M. & Tervit, H.R. (1999). Production of cloned calves following nuclear transfer with cultured adult mural granulosa cells. Biol. Reprod. 60, 996–1005.Google Scholar
Wilmut, I., Schnieke, A.E., McWhir, J., Kind, A.J. & Campbell, K.H. (1997). Viable offspring derived from fetal and adult mammalian cells. Nature 385, 810–3.Google Scholar