Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Normal human mammary epithelial cells spontaneously escape senescence and acquire genomic changes

Nature volume 409, pages 633–637 (2001)Cite this article

Abstract

Senescence and genomic integrity are thought to be important barriers in the development of malignant lesions1. Human fibroblasts undergo a limited number of cell divisions before entering an irreversible arrest, called senescence2. Here we show that human mammary epithelial cells (HMECs) do not conform to this paradigm of senescence. In contrast to fibroblasts, HMECs exhibit an initial growth phase that is followed by a transient growth plateau (termed selection or M0; refs 3,4,5), from which proliferative cells emerge to undergo further population doublings (∼20–70), before entering a second growth plateau (previously termed senescence or M1; refs 4,5,6). We find that the first growth plateau exhibits characteristics of senescence but is not an insurmountable barrier to further growth. HMECs emerge from senescence, exhibit eroding telomeric sequences and ultimately enter telomere-based crisis to generate the types of chromosomal abnormalities seen in the earliest lesions of breast cancer. Growth past senescent barriers may be a pivotal event in the earliest steps of carcinogenesis, providing many genetic changes that predicate oncogenic evolution. The differences between epithelial cells and fibroblasts provide new insights into the mechanistic basis of neoplastic transformation.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: HMF and HMEC growth curves and cell morphologies in vitro.
Figure 2: Spontaneous genomic instability in human mammary epithelial cells.
Figure 3: Post-selection HMECs continue to shorten telomeres beyond the length detected in senescent HMFs and HMECs at the first growth plateau.

Similar content being viewed by others

References

  1. Shay, J. W., Wright, W. E. & Werbin, H. Toward a molecular understanding of human breast cancer: a hypothesis. Breast Cancer Res. Treatment. 25 83–94 (1993).

    Article  CAS  Google Scholar 

  2. Hayflick, L. The limited in vitro lifetime of human diploid cell strains. Exp. Cell Res. 37, 614–636 (1965).

    Article  CAS  Google Scholar 

  3. Hammond, S. L., Ham, R. G. & Stampfer, M. R. Serum-free growth of human mammary epithelial cells: rapid clonal growth in defined medium and extended passage with pituitary extract. Proc. Natl Acad. Sci. USA 81, 5435–5439 (1984).

    Article  ADS  CAS  Google Scholar 

  4. Foster, S. A. & Galloway, D. A. Human papillomavirus type 16 E7 alleviates a proliferation block in early passage human mammary epithelial cells. Oncogene 12, 1773–1779 (1996).

    CAS  PubMed  Google Scholar 

  5. Huschtscha, L. I. et al. Loss of p16INK4 expression by methylation is associated with lifespan extension of human mammary epithelial cells. Cancer Res. 58, 3508–3512 (1998).

    CAS  PubMed  Google Scholar 

  6. Kiyono, T. et al. Both Rb/p16INK4a inactivation and telomerase activity are required to immortalize human epithelial cells. Nature 396, 84–88 (1998).

    Article  ADS  CAS  Google Scholar 

  7. Meyer, K. M., Hess, S. M., Tlsty, T. D. & Leadon, S. A. Human mammary epithelial cells exhibit a differential p53-mediated response following exposure to ionizing or UV light. Oncogene 18, 5792–57805 (1999).

    Article  Google Scholar 

  8. Walen, K. H. & Stampfer, M. R. Chromosome analyses of human mammary epithelial cells at stages of chemical-induced transformation progression to immortality. Cancer Genet. Cytogenet. 37, 249–261 (1989).

    Article  CAS  Google Scholar 

  9. Brenner, A. J., Stampfer, M. R. & Aldaz, C. M. Increased p16 expression with first senescence arrest in human mammary epithelial cells and extended growth capacity with p16 inactivation. Oncogene 17, 199–205 (1998).

    Article  CAS  Google Scholar 

  10. Dimri, G. P. et al. A biomarker that identifies senescent human cells in culture and in aging skin in vivo. Proc. Natl Acad. Sci. USA 92, 9363–9367 (1995).

    Article  ADS  CAS  Google Scholar 

  11. Pignolo, R. J., Rotenberg, M. O. & Cristofalo, V. J. Alterations in contact and density-dependent arrest state in senescent WI-38 cells. In Vitro Cell. Dev. Biol. Anim. 30A, 471–476 (1994).

    Article  CAS  Google Scholar 

  12. Shay, J. W. & Wright, W. E. Quantitation of the frequency of immortalization of normal human diploid fibroblasts by SV40 large T-antigen. Exp. Cell Res. 184, 109–118 (1989).

    Article  CAS  Google Scholar 

  13. Taylor-Papadimitriou, J. et al. Keratin expression in human mammary epithelial cells cultured from normal and malignant tissue: relation to in vivo phenotypes and influence of medium. J. Cell Sci. 94, 403–413 (1989).

    PubMed  Google Scholar 

  14. Foster, S. A., Wong, D. J., Barrett, M. T. & Galloway, D. A. Inactivation of p16 in human mammary epithelial cells by CpG island methylation. Mol. Cell. Biol. 18, 1793–1801 (1998).

    Article  CAS  Google Scholar 

  15. Lansdorp, P. M. et al. Heterogeneity in telomere length of human chromosomes. Hum. Mol. Genet. 5, 685–691 (1996).

    Article  CAS  Google Scholar 

  16. van Steensel, B., Smogorzewska, A. & de Lange, T. TRF2 protects human telomeres from end-to-end fusions. Cell 92, 401–413 (1998).

    Article  CAS  Google Scholar 

  17. Stampfer, M. R. et al. Gradual phenotypic conversion associated with immortalization of cultured human mammary epithelial cells. Mol. Biol. Cell 8, 2391–2405 (1997).

    Article  CAS  Google Scholar 

  18. Karlseder, J., Broccoli, D., Dai, Y., Hardy, S. & de Lange, T. p53- and ATM-dependent apoptosis induced by telomeres lacking TRF2. Science 283, 1321–1325 (1999).

    Article  CAS  Google Scholar 

  19. Artandi, S. E. et al. Telomere dysfunction promotes non-reciprocal translocations and epithelial cancers in mice. Nature 406, 641–645 (2000).

    Article  ADS  CAS  Google Scholar 

  20. Chin, L. et al. p53 Deficiency rescues the adverse effects of telomere loss and cooperates with telomere dysfunction to accelerate carcinogenesis. Cell 97, 527–538 (1999).

    Article  CAS  Google Scholar 

  21. Alcorta, D. A. et al. Involvement of the cyclin-dependent kinase inhibitor p16 (INK4A) in replicative senescence of normal human fibroblasts. Proc. Natl Acad. Sci. USA 92, 13742–13747 (1996).

    Article  ADS  Google Scholar 

  22. Hara, E. et al. Regulation of p16CDKN2 expression and its implications for cell immortalization and senescence. Mol. Cell. Biol. 16, 859–867 (1996).

    Article  CAS  Google Scholar 

  23. Burbano, R. R. et al. Cytogenetics of epithelial hyperplasias of the human breast. Cancer Genet. Cytogenet. 119, 62–66 (2000).

    Article  CAS  Google Scholar 

  24. Pandis, N. et al. Chromosome abnormalities in bilateral breast carcinomas. Cytogenetic evaluation of the clonal origin of multiple primary tumors. Cancer 76, 250–258 (1995).

    Article  CAS  Google Scholar 

  25. Berg, J. W. & Hutter, R. V. Breast cancer. Cancer 75, 257–269 (1995).

    Article  CAS  Google Scholar 

  26. Stoeber, K. et al. Cdc6 protein causes premature entry into S phase in a mammalian cell-free system. EMBO J. 17, 7219–7229 (1998).

    Article  CAS  Google Scholar 

  27. Wei, W. & Sedivy, J. M. Differentiation between senescence (M1) and crisis (M2) in human fibroblasts cultures. Exp. Cell Res. 253, 519–522 (1999).

    Article  CAS  Google Scholar 

  28. Tlsty, T. D. et al. Potentiation of genomic instability in normal human mammary epithelial cells by an epigenetic event. J. Mammary Gland Biol. Neoplasia (in the press).

Download references

Acknowledgements

We thank E. H. Blackburn, I. Herskowitz and J. Li for comments, criticism and reading the manuscript. Stimulating discussion and thoughtful critique were provided by Y. Crawford, G. Whitworth, M. Heiman, M. Springer, D. Crawford, P. Hein and J. Anderson. We thank S. Gilbert for assistance with the figures; P. Ortiz for library support; and G. Williams for MCM2 antibodies. This work was supported by NIH and NIH/NASA grants to T.D.T. and a DOE and NIH grant to M.R.S. C.R.H. is supported by a Howard Hughes Pre-doctoral Fellowship.

Author information

Authors and Affiliations

  1. Department of Pathology and UCSF Comprehensive Cancer Center, University of California at San Francisco, San Francisco, 94143-0506, California, USA

    Serguei R. Romanov, B. Krystyna Kozakiewicz, Charles R. Holst, Larisa M. Haupt & Thea D. Tlsty

  2. Lawrence Berkeley National Laboratory, Berkeley, 94720, California, USA

    Martha R. Stampfer

Authors
  1. Serguei R. Romanov
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. B. Krystyna Kozakiewicz
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Charles R. Holst
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Martha R. Stampfer
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Larisa M. Haupt
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Thea D. Tlsty
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Thea D. Tlsty.

Supplementary Information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Romanov, S., Kozakiewicz, B., Holst, C. et al. Normal human mammary epithelial cells spontaneously escape senescence and acquire genomic changes. Nature 409, 633–637 (2001). https://doi.org/10.1038/35054579

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/35054579

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing