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Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts

An Erratum to this article was published on 01 September 2003

Abstract

Most mammalian cells do not divide indefinitely, owing to a process termed replicative senescence. In human cells, replicative senescence is caused by telomere shortening, but murine cells senesce despite having long stable telomeres1. Here, we show that the phenotypes of senescent human fibroblasts and mouse embryonic fibroblasts (MEFs) differ under standard culture conditions, which include 20% oxygen. MEFs did not senesce in physiological (3%) oxygen levels, but underwent a spontaneous event that allowed indefinite proliferation in 20% oxygen. The proliferation and cytogenetic profiles of DNA repair-deficient MEFs suggested that DNA damage limits MEF proliferation in 20% oxygen. Indeed, MEFs accumulated more DNA damage in 20% oxygen than 3% oxygen, and more damage than human fibroblasts in 20% oxygen. Our results identify oxygen sensitivity as a critical difference between mouse and human cells, explaining their proliferative differences in culture, and possibly their different rates of cancer and ageing.

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Figure 1: Senescent MEFs resemble oxidant-treated human fibroblasts.
Figure 2: Low oxygen abolishes replicative senescence of MEFs.
Figure 3: Growth of repair-deficient MEFs in 3% and 20% oxygen.
Figure 4: MEFs accumulate high levels of oxidative DNA damage in 20% oxygen.

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Acknowledgements

We thank D. Chen and his group for technical advice and DNA-PKcs−/− MEFs, P. Hasty for Ku80−/− MEFs and critical reading of the manuscript, S. Chang for mTR−/− MEFs, J. Hoeijmakers and H. van Steeg for xpa−/− MEFs, and J. Vijg and his group for helpful discussions. This work was supported by research grants from the National Institutes of Health (AG17242 to J.C.; AG18679 to S.M.), and training grants from the National Institutes of Health (AG00266 to J.G.) and Department of Defence Breast Cancer Research Program (8KB-0100 and BC010658 to S.P.), under contract AC03-76SF00098 to the University of California by the Department of Energy.

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Correspondence to Judith Campisi.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information, Fig. S1 Efficacy of GSE, levels of G1 cyclin-dependent kinases, and responsiveness to serum deprivation of MEFs cultured in 3% O2.

Supplementary Information, Fig. S2 Senescence of Balb/c and DNA-PKcs -/- MEFs in low oxygen. (PDF 398 kb)

Supplementary Information, Fig. S3 Telomere-FISH and growth of telomerase-deficient MEFs in 3% and 20% O2.

Supplementary Information, Table S1 Chromosomal aberrations in MEF cultures (DOC 31 kb)

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Parrinello, S., Samper, E., Krtolica, A. et al. Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts. Nat Cell Biol 5, 741–747 (2003). https://doi.org/10.1038/ncb1024

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