Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

Isolation, culture and characterisation of somatic cells derived from semen and milk of endangered sheep and eland antelope

L. Nel-Themaat A B , M. C. Gómez A B , P. Damiani B , G. Wirtu B C , B. L. Dresser B D , K. R. Bondioli A F , L. A. Lyons E , C. E. Pope B and R. A. Godke A C
+ Author Affiliations
- Author Affiliations

A Department of Animal Sciences, Louisiana State University Agricultural Center, Baton Rouge, LA 70803, USA.

B Audubon Center for Research of Endangered Species, 14001 River Road, New Orleans, LA 70131, USA.

C Department of Veterinary Clinical Sciences, Louisiana State University School of Veterinary Medicine, Baton Rouge, LA 70803, USA.

D Department of Biological Sciences, University of New Orleans, New Orleans, LA 70148, USA.

E School of Veterinary Medicine, University of California—Davis, Davis, CA 95616, USA.

F Corresponding author. Email: Kbondioli@agcenter.lsu.edu

Reproduction, Fertility and Development 19(4) 576-584 https://doi.org/10.1071/RD06153
Submitted: 8 November 2006  Accepted: 19 March 2007   Published: 7 May 2007

Abstract

Semen and milk are potential sources of somatic cells for genome banks. In the present study, we cultured and characterised cells from: (1) cooled sheep milk; (2) fresh, cooled and frozen–thawed semen from Gulf Coast native (GCN) sheep (Ovis aries); and (3) fresh eland (Taurotragus oryx) semen. Cells attached to the culture surface from fresh (29%), cooled (43%) and slow-frozen (1°C/min; 14%) ram semen, whereas no attachment occurred in the fast-frozen (10°C/min) group. Proliferation occurred in fresh (50%) and cooled (100%) groups, but no cells proliferated after passage 1 (P1). Eland semen yielded cell lines (100%) that were cryopreserved at P1. In samples from GCN and cross-bred milk, cell attachment (83% and 95%, respectively) and proliferation (60% and 37%, respectively) were observed. Immunocytochemical detection of cytokeratin indicated an epithelial origin of semen-derived cells, whereas milk yielded either fibroblasts, epithelial or a mixture of cell types. Deoxyribonucleic acid microsatellite analysis using cattle-derived markers confirmed that eland cells were from the semen donor. Eland epithelial cells were transferred into eland oocytes and 12 (71%), six (35%) and two (12%) embryos cleaved and developed to morulae or blastocyst stages, respectively. In conclusion, we have developed a technique for obtaining somatic cells from semen. We have also demonstrated that semen-derived cells can serve as karyoplast donors for nuclear transfer.

Additional keywords: cell culture, cryopreservation, ejaculate, epithelial, Gulf Coast native sheep, nuclear transfer.


Acknowledgements

The authors extend their gratitude to Tray Harding and the LSU Agricultural Center Sheep Research Station team for providing the sheep and helping with sample collection. The authors also acknowledge Dr Alex Cole, Contessa Knight and the animal keepers from the Audubon Research Center Species Survival Center for helping with the eland semen collections.


References

Arase, S. , Sadamoto, Y. , Kato, S. , Fujie, K. , Nakanishi, H. , and Takeda, K. (1990). Three-dimensional culture of plucked human hair follicles inside the collagen gel matrix. Nippon Hifuka Gakkai Zasshi 100, 879–882.
PubMed |

Ashwood-Smith M. J. 1980. Low temperature preservation of cells, tissues and organs. In ‘Low Temperature Preservation in Medicine and Biology’. (Eds M. J. Ashwood-Smith and J. Farrant.) pp. 19–44. (Pitman Medical: Kent.)

Ashwood-Smith, M. J. , and Lough, P. (1975). Cryoprotection of mammalian cells in tissue culture with methanol. Cryobiology 12, 517–518.
CrossRef | PubMed |

Bishop, M. D. , Kappes, S. M. , Keele, J. W. , Stone, R. T. , Sunden, S. L. , Hawkins, G. A. , Toldo, S. S. , Fries, R. , Grosz, M. D. , and Yoo, J. (1994). A genetic linkage map for cattle. Genetics 136, 619–639.
PubMed |

Blackburn, H. D. (2004). Development of national animal genetic resource programs. Reprod. Fertil. Dev. 16, 27–32.
CrossRef | PubMed |

Creighton, P. , Eggen, A. , Fries, R. , Jordan, S. A. , Hetzel, J. , Cunningham, E. P. , and Humphries, P. (1992). Mapping of bovine markers CYP21, PRL, and BOLA DRBP1 by genetic linkage analysis in reference pedigrees. Genomics 14, 526–528.
CrossRef | PubMed |

Elicha Gussin, H. A. , and Elias, S. (2002). Culture of fetal cells from maternal blood for prenatal diagnosis. Hum. Reprod. Update 8, 523–527.
CrossRef | PubMed |

Evans G., and Maxwell W. M. C. (1987). ‘Artificial Insemination of Sheep and Goats.’ (Butterworths: Sydney.)

Evenson, D. P. , and Melamed, M. R. (1983). Rapid analysis of normal and abnormal cell types in human semen and testis biopsies by flow cytometry. J. Histochem. Cytochem. 31, 248–253.
PubMed |

Farrant, J. , and Morris, G. J. (1973). Thermal shock and dilution shock as the causes of freezing injury. Cryobiology 10, 134–140.
CrossRef | PubMed |

Freshney R. I., and Freshney M. G. (Eds) (2002). ‘Culture of Epithelial Cells.’ (Wiley-Liss: New York.)

Gómez, M. C. , Pope, C. E. , Giraldo, A. , Lyons, L. A. , Harris, R. F. , King, A. L. , Cole, A. , Godke, R. A. , and Dresser, B. L. (2004). Birth of African wildcat cloned kittens born from domestic cats. Cloning Stem Cells 6, 247–258.
PubMed |

Green, A. E. (1967). Viability of cell cultures following extended preservation in liquid nitrogen. Proc. Soc. Exp. Biol. Med. 124, 1302–1307.
PubMed |

Janssen, D. L. , Edwards, M. L. , Koster, J. A. , Lanza, R. P. , and Ryder, O. A. (2004). Postnatal management of chryptordich banteng calves cloned by nuclear transfer utilizing frozen fibroblast cultures and enucleated cow ova. Reprod. Fertil. Dev. 16, 224.. [Abstract]
CrossRef |

Katska, L. , Bochenek, M. , Kania, G. , Rynska, B. , and Smorag, Z. (2002). Flow cytometric cell cycle analysis of somatic cells primary cultures established for bovine cloning. Theriogenology 58, 1733–1744.
CrossRef | PubMed |

Kishi, M. , Itagaki, Y. , Takakura, R. , Imamura, M. , Sudo, T. , Yoshinari, M. , Tanimoto, M. , Yasue, H. , and Kashima, N. (2000). Nuclear transfer in cattle using colostrum-derived mammary gland epithelial cells and ear-derived fibroblast cells. Theriogenology 54, 675–684.
CrossRef | PubMed |

Kossarek, L. M. , Grosse, W. M. , Finco, T. S. , and McGraw, R. A. (1993a). Rapid communication: bovine dinucleotide repeat polymorphism RM002. J. Anim. Sci. 71, 3173.
PubMed |

Kossarek, L. M. , Su, X. , Grosse, W. M. , Finlay, O. , Barendse, W. , Hetzel, D. J. , Dietz, A. B. , Womack, J. E. , and McGraw, R. A. (1993b). Rapid communication: bovine dinucleotide repeat polymorphism RM067. J. Anim. Sci. 71, 3178.
PubMed |

Krisher, R. L. , and Bavister, B. D. (1999). Enhanced glycolysis after maturation of bovine oocytes in vitro is associated with increased developmental competence. Mol. Reprod. Dev. 53, 19–26.
CrossRef | PubMed |

Kurata, S. , Itami, S. , Terashi, H. , and Takayasu, S. (1994). Successful transplantation of cultured human outer root sheath cells as epithelium. Ann. Plast. Surg. 33, 290–294.
CrossRef | PubMed |

Lanza, R. P. , Cibelli, J. B. , Diaz, F. , Moraes, C. T. , Farin, P. W. , Hammer, C. J. , West, M. D. , and Damiani, P. (2000). Cloning of an endangered species (Bos gaurus) using interspecies nuclear transfer. Cloning 2, 79–90.
CrossRef | PubMed |

Loi, P. , Ptak, G. , Barboni, B. , Fulka, J. , Cappai, P. , and Clinton, M. (2001). Genetic rescue of an endangered mammal by cross-species nuclear transfer using post-mortem somatic cells. Nat. Biotechnol. 19, 962–964.
CrossRef | PubMed |

Lovelock, J. E. (1953). The haemolysis of human red blood cells by freezing and thawing. Biochim. Biophys. Acta 10, 414–426.
CrossRef | PubMed |

Luyet, B. J. , and Gonzales, F. (1952). Protective action of glycerol against freezing injury in embryonic tissues of chick. Biodynamica 7, 101–106.
PubMed |

Macpherson, I. , and Bryden, A. (1971). Mitomycin C treated cells as feeders. Exp. Cell Res. 69, 240–241.
CrossRef | PubMed |

Masui, T. , Wakefiels, J. F. , Lechner, M. A. , LaVech, M. B. , Sporn, M. B. , and Harris, C. C. (1986). Type beta transforming growth factor is the primary differentiation-inducing serum factor for normal human bronchial epithelial cells. Proc. Natl Acad. Sci. USA 83, 2438–2442.
CrossRef |

Mazur, P. , Leibo, S. P. , and Chu, E. H. Y. (1972). A two-factor hypothesis of freezing injury. Exp. Cell Res. 71, 345–355.
CrossRef | PubMed |

Moll, I. (1996). Differential epithelial outgrowth of plucked and microdissected human hair follicles in explant culture. Arch. Dermatol. Res. 288, 604–610.
PubMed |

Moses, H. L. , Tucker, E. B. , Leof, R. J. , Coffey, R. J. , Halper, J. , and Shipley, G. D. (1985). Type beta transforming growth factor is a growth stimulator and growth inhibitor. Cancer Cells 3, 67–71.


Paape, M. J. , and Tucker, H. A. (1966). Somatic cell content variation in fraction-collected milk. J. Dairy Sci. 49, 265–267.
PubMed |

Ryder, O. A. (2002). Cloning advances and challenges for conservation. Trends Biotechnol. 20, 231–232.
CrossRef | PubMed |

Ryder, O. A. , and Benirschke, K. (1997). The potential use of ‘cloning’ in the conservation effort. Zoo Biol. 16, 295–300.
CrossRef |

Stampfer M. R., Yaswen P., and Taylor-Papadimitriou J. 2002. Culture of human epithelial cells. In ‘Culture of Epithelial Cells’. (Eds R. I. Freshney and M. G. Freshney.) pp. 95–135. (Wiley-Liss: New York.)

Toonen, R. J. , and Hughes, S. (2001). Increased throughput for fragment analysis on an ABI PRISM 377 automated sequencer using a membrane comb and STRand software. Biotechniques 31, 1320–1324.
PubMed |

Tucker, R. F. , Shipley, G. D. , Moses, H. L. , and Holley, R. W. (1984). Growth inhibitor from BSC-1 cells closely related to platelet type beta transforming growth facor. Science 226, 705–707.
CrossRef | PubMed |

Wilmut, I. , Schnieke, A. E. , McWhir, J. , Kind, A. J. , and Campbell, K. H. S. (1997). Viable offspring derived from fetal and adult mammalian cells. Nature 385, 810–813.
CrossRef | PubMed |

Wilmut, I. , Beaujean, N. , de Sousa, P. A. , Dinnyes, A. , King, T. J. , Paterson, L. A. , Wells, D. N. , and Young, L. E. (2002). Somatic cell nuclear transfer. Nature 419, 583–586.
CrossRef | PubMed |

Wirtu G. (2004). Developing embryo technologies for the eland antelope (Taurotragus oryx). Ph.D. Thesis, Louisiana State University and Agricultural and Mechanical College, USA.

Wirtu G., Pope C. E., Lee B. C., Cole A., Vaccaro J., Sarrat E. L., Bavister B. D., Godke R. A., and Dresser B. L. (2002). Collection of semen by rectal massage from an eland (Taurotragu oryx) bull. In ‘Proceedings of the 2nd International Symposium on Assisted Reproductive Technology (ART) for the Conservation and Genetic Management of Wildlife’. pp. 318–320. (Omaha Zoo: Nebraska.)



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