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Spermatogonial Stem Cell Transplantation, Testicular Function, and Restoration of Male Fertility in Mice

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Germline Stem Cells

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 450))

Summary

Mammalian spermatogonial stem cells, sometimes called male germline stem cells, are a small population of adult tissue-specific stem cells present in the testis. Formation of the spermatogonial stem cell population early in life and differentiation of spermatogonial stem cells in adults are responsible for continual production of sperm in the testis. Unfortunately, there are no specific biochemical or morphological markers for spermatogonial stem cells, so investigation of this cell type requires specific and consistent approaches to ensure valid data are obtained. Currently, the only assay for the presence of spermatogonial stem cells in a cell suspension is the spermatogonial stem cell transplantation technique. This requires the transfer of cells from a donor animal into the testis of a recipient animal, in which the spermatogonial stem cells will colonize and initiate donor-derived spermatogenesis. Although there is no specific marker for spermatogonial stem cells, several cell surface markers have been used to enrich for these cells prior to transplantation. Thus, selection and transplantation of spermatogonial stem cells can be used to investigate basic mechanisms regulating them. Successful transplantation and donor-derived spermatogenesis in recipient animals can lead to the restoration of fertility in infertile males. In combination with spermatogonial stem cell culture, this transplantation technique can also be used for the purpose of generating transgenic animals through the male germline. This chapter describes the methods for spermatogonial stem cell transplantation and how this approach is used to investigate testicular function.

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References

  1. 1. Sharpe, R. (1994) Regulation of spermatogenesis, in The physiology of reproduction (E. K. Knobil and J. D. Neill, eds.), Raven, New York, pp. 1363–1434.

    Google Scholar 

  2. 2. de Rooij, D. G., and Russell, L. D. (2000) All you wanted to know about spermatogonia but were afraid to ask. J. Androl. 21, 776–798.

    PubMed  Google Scholar 

  3. 3. Spradling, A., Drummond-Barbosa, D., and Kai, T. (2001) Stem cells find their niche. Nature. 414, 98–104.

    Article  CAS  PubMed  Google Scholar 

  4. 4. Shinohara, T., Orwig, K. E., Avarbock, M. R., and Brinster, R. L. (2001) Remodeling of the postnatal mouse testis is accompanied by dramatic changes in stem cell number and niche accessibility. Proc. Natl. Acad. Sci. U. S. A. 98, 6186–6191.

    Article  CAS  PubMed  Google Scholar 

  5. 5. McLean, D. J., Russell, L .D., and Griswold, M. D. (2002) Biological activity and enrichment of spermatogonial stem cells in vitamin A-deficient and hyperthermia-exposed testes from mice based on colonization following germ cell transplantation. Biol. Reprod. 66, 1374–1379.

    Article  CAS  PubMed  Google Scholar 

  6. 6. Brinster, R. L. (2002) Germline stem cell transplantation and transgenesis. Science. 296, 2174–2176.

    Article  CAS  PubMed  Google Scholar 

  7. 7. McLean, D. J. (2005) Spermatogonial stem cell transplantation and testicular function. Cell Tissue Res. 322, 21–31.

    Article  PubMed  Google Scholar 

  8. 8. Brinster, R. L., and Zimmermann, J. W. (1994) Spermatogenesis following male germ-cell transplantation. Proc. Natl. Acad. Sci. U. S. A. 91, 11298–11302.

    Article  CAS  PubMed  Google Scholar 

  9. 9. Kanatsu-Shinohara, M., Toyokuni, S., and Shinohara, T. (2004) CD9 is a surface marker on mouse and rat male germline stem cells. Biol. Reprod. 70, 70–75.

    Article  CAS  PubMed  Google Scholar 

  10. 10. Kubota, H., Avarbock, M. R., and Brinster, R. L. (2003) Spermatogonial stem cells share some, but not all, phenotypic and functional characteristics with other stem cells. Proc. Natl. Acad. Sci. U. S. A. 100, 6487–6492.

    Article  CAS  PubMed  Google Scholar 

  11. 11. Shinohara, T., Avarbock, M.R., and Brinster, R.L. (1999) beta1- and alpha6-integrin are surface markers on mouse spermatogonial stem cells. Proc. Natl. Acad. Sci. U. S. A. 96, 5504–5509.

    Article  CAS  PubMed  Google Scholar 

  12. 12. Shinohara, T., Orwig, K. E., Avarbock, M. R., and Brinster, R. L. (2000) Spermatogonial stem cell enrichment by multiparameter selection of mouse testis cells. Proc. Natl. Acad. Sci. U. S. A. 97, 8346–8351.

    Article  CAS  PubMed  Google Scholar 

  13. 13. Hamra, F. K., Gatlin, J., Chapman, K. M., et al. (2002) Production of transgenic rats by lentiviral transduction of male germ-line stem cells. Proc. Natl. Acad. Sci. U. S. A. 99, 14931–14936.

    Article  CAS  PubMed  Google Scholar 

  14. 14. Nagano, M., Brinster, C. J., Orwig, K. E., Ryu, B. Y., Avarbock, M. R., and Brinster, R. L. (2001) Transgenic mice produced by retroviral transduction of male germ-line stem cells. Proc. Natl. Acad. Sci. U. S. A. 98, 13090–13095.

    Article  CAS  PubMed  Google Scholar 

  15. 15. Kubota, H., Avarbock, M. R., and Brinster, R. L. (2004) Growth factors essential for self-renewal and expansion of mouse spermatogonial stem cells. Proc. Natl. Acad. Sci. U. S. A. 101, 16489–16494.

    Article  CAS  PubMed  Google Scholar 

  16. 16. Kubota, H., and Brinster, R. L. (2006) Technology insight: In vitro culture of spermatogonial stem cells and their potential therapeutic uses. Nat. Clin. Pract. Endocrinol. Metab. 2, 99–108.

    Article  CAS  PubMed  Google Scholar 

  17. 17. Boettger-Tong, H.L., Johnston, D.S., Russell, L.D., Griswold, M.D., and Bishop, C.E. (2000) Juvenile spermatogonial depletion (jsd) mutant seminiferous tubules are capable of supporting transplanted spermatogenesis. Biol. Reprod. 63, 1185–1191.

    Article  CAS  PubMed  Google Scholar 

  18. 18. Buaas, F. W., Kirsh, A. L., Sharma, M., et al. (2004) Plzf is required in adult male germ cells for stem cell self-renewal. Nat. Genet. 36, 647–652.

    Article  CAS  PubMed  Google Scholar 

  19. 19. Johnston, D. S., Russell, L. D., Friel, P. J., and Griswold, M. D. (2001) Murine germ cells do not require functional androgen receptors to complete spermatogenesis following spermatogonial stem cell transplantation. Endocrinology. 142, 2405–2408.

    Article  CAS  PubMed  Google Scholar 

  20. 20. Herrid, M., Vignarajan, S., Davey, R., Dobrinski, I., and Hill, J. R. (2006) Successful transplantation of bovine testicular cells to heterologous recipients. Reproduction. 132, 617–624.

    Article  CAS  PubMed  Google Scholar 

  21. 21. Honaramooz, A., Megee, S. O., and Dobrinski, I. (2002) Germ cell transplantation in pigs. Biol. Reprod. 66, 21–28.

    Article  CAS  PubMed  Google Scholar 

  22. 22. Ogawa, T., Dobrinski, I., Avarbock, M. R., and Brinster, R. L. (2000) Transplantation of male germ line stem cells restores fertility in infertile mice. Nat. Med. 6, 29–34.

    Article  CAS  PubMed  Google Scholar 

  23. 23. Shinohara, T., Avarbock, M. R., and Brinster, R. L. (2000) Functional analysis of spermatogonial stem cells in Steel and cryptorchid infertile mouse models. Dev. Biol. 220, 401–411.

    Article  CAS  PubMed  Google Scholar 

  24. 24. Brinster, C. J., Ryu, B. Y., Avarbock, M. R., Karagenc, L., Brinster, R. L., and Orwig, K. E. (2003) Restoration of fertility by germ cell transplantation requires effective recipient preparation. Biol. Reprod. 69, 412–420.

    Article  CAS  PubMed  Google Scholar 

  25. 25. Zhang, Z., Shao, S., and Meistrich, M. L. (2006) Irradiated mouse testes efficiently support spermatogenesis derived from donor germ cells of mice and rats. J. Androl. 27, 365–375.

    Article  PubMed  Google Scholar 

  26. 26. Oatley, J. M., de Avila, D. M., McLean, D. J., Griswold, M. D., and Reeves, J. J. (2002) Transplantation of bovine germinal cells into mouse testes. J. Anim. Sci. 80, 1925–1931.

    CAS  PubMed  Google Scholar 

  27. 27. Ogawa, T., Dobrinski, I., Avarbock, M. R., and Brinster, R. L. (1999) Xenogeneic spermatogenesis following transplantation of hamster germ cells to mouse testes. Biol. Reprod. 60, 515–521.

    Article  CAS  PubMed  Google Scholar 

  28. 28. Oatley, J. M., Reeves, J. J., and McLean, D. J. (2004) Biological activity of cryopreserved bovine spermatogonial stem cells during in vitro culture. Biol. Reprod. 71, 942–947.

    Article  CAS  PubMed  Google Scholar 

  29. 29. Franca, L. R., Ogawa, T., Avarbock, M. R., Brinster, R. L., and Russell, L. D. (1998) Germ cell genotype controls cell cycle during spermatogenesis in the rat. Biol. Reprod. 59, 1371–1377.

    Article  CAS  PubMed  Google Scholar 

  30. 30. Hill, J. R., and Dobrinski, I. (2006) Male germ cell transplantation in livestock. Reprod. Fertil. Dev. 18, 13–18.

    Article  CAS  PubMed  Google Scholar 

  31. 31. McLean, D. J., Friel, P. J., Johnston, D. S., and Griswold, M. D. (2003) Characterization of spermatogonial stem cell maturation and differentiation in neonatal mice. Biol. Reprod. 69, 2085–2091.

    Article  CAS  PubMed  Google Scholar 

  32. 32. Kanatsu-Shinohara, M., Toyokuni, S., and Shinohara, T. (2005) Genetic selection of mouse male germline stem cells in vitro: offspring from single stem cells. Biol. Reprod. 72, 236–240.

    Article  CAS  PubMed  Google Scholar 

  33. 33. Honaramooz, A., Behboodi, E., Megee, S. O., et al. (2003) Fertility and germline transmission of donor haplotype following germ cell transplantation in immunocompetent goats. Biol. Reprod. 69, 1260–1264.

    Article  CAS  PubMed  Google Scholar 

  34. 34. Oatley, J. M., Avarbock, M. R., Telaranta, A. I., Fearon, D. T., and Brinster, R. L. (2006) Identifying genes important for spermatogonial stem cell self-renewal and survival. Proc. Natl. Acad. Sci. U. S. A. 103, 9524–9529.

    Article  CAS  PubMed  Google Scholar 

  35. 35. Geijsen, N., Horoschak, M., Kim, K., Gribnau, J., Eggan, K., and Daley, G. Q. (2004) Derivation of embryonic germ cells and male gametes from embryonic stem cells. Nature. 427, 148–154.

    Article  CAS  PubMed  Google Scholar 

  36. 36. Oatley, J. M., and Brinster, R. L. (2006) Spermatogonial stem cells. Methods Enzymol. 419, 259–282.

    Article  CAS  PubMed  Google Scholar 

  37. 37. Zhang, Z., Shao, S., and Meistrich, M. L. (2006) The radiation-induced block in spermatogonial differentiation isdue to damage to the somatic environment, not the germ cells. J. Cell Physiol. 211, 149–158.

    Article  Google Scholar 

  38. 38. Hendrich, H. (2004) The laboratory mouse, Elsevier, Amsterdam.

    Google Scholar 

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Acknowledgments

I gratefully acknowledge the support from the National Institutes of Health (HD046521). I thank Drs. Daniel Johnston and Jon Oatley for helpful discussion on protocol development.

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Authors and Affiliations

  1. Department of Animal Sciences, Center for Reproductive Biology, Washington State University, Pullman, WA

    Derek J. McLean

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  1. Derek J. McLean
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Editors and Affiliations

  1. National Cancer Institute, Frederick, MD, USA

    Steven X. Hou  & Shree Ram Singh  & 

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© 2008 Humana Press, a part of Springer Science + Business Media, LLC

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McLean, D.J. (2008). Spermatogonial Stem Cell Transplantation, Testicular Function, and Restoration of Male Fertility in Mice. In: Hou, S.X., Singh, S.R. (eds) Germline Stem Cells. Methods in Molecular Biology™, vol 450. Humana Press. https://doi.org/10.1007/978-1-60327-214-8_11

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  • DOI: https://doi.org/10.1007/978-1-60327-214-8_11

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-60327-213-1

  • Online ISBN: 978-1-60327-214-8

  • eBook Packages: Springer Protocols

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