Telomerase reactivation reverses tissue degeneration in aged telomerase-deficient mice

Journal name:
Nature
Volume:
469,
Pages:
102–106
Date published:
DOI:
doi:10.1038/nature09603
Received
Accepted
Published online

An ageing world population has fuelled interest in regenerative remedies that may stem declining organ function and maintain fitness. Unanswered is whether elimination of intrinsic instigators driving age-associated degeneration can reverse, as opposed to simply arrest, various afflictions of the aged. Such instigators include progressively damaged genomes. Telomerase-deficient mice have served as a model system to study the adverse cellular and organismal consequences of wide-spread endogenous DNA damage signalling activation in vivo1. Telomere loss and uncapping provokes progressive tissue atrophy, stem cell depletion, organ system failure and impaired tissue injury responses1. Here, we sought to determine whether entrenched multi-system degeneration in adult mice with severe telomere dysfunction can be halted or possibly reversed by reactivation of endogenous telomerase activity. To this end, we engineered a knock-in allele encoding a 4-hydroxytamoxifen (4-OHT)-inducible telomerase reverse transcriptase-oestrogen receptor (TERT-ER) under transcriptional control of the endogenous TERT promoter. Homozygous TERT-ER mice have short dysfunctional telomeres and sustain increased DNA damage signalling and classical degenerative phenotypes upon successive generational matings and advancing age. Telomerase reactivation in such late generation TERT-ER mice extends telomeres, reduces DNA damage signalling and associated cellular checkpoint responses, allows resumption of proliferation in quiescent cultures, and eliminates degenerative phenotypes across multiple organs including testes, spleens and intestines. Notably, somatic telomerase reactivation reversed neurodegeneration with restoration of proliferating Sox2+ neural progenitors, Dcx+ newborn neurons, and Olig2+ oligodendrocyte populations. Consistent with the integral role of subventricular zone neural progenitors in generation and maintenance of olfactory bulb interneurons2, this wave of telomerase-dependent neurogenesis resulted in alleviation of hyposmia and recovery of innate olfactory avoidance responses. Accumulating evidence implicating telomere damage as a driver of age-associated organ decline and disease risk1, 3 and the marked reversal of systemic degenerative phenotypes in adult mice observed here support the development of regenerative strategies designed to restore telomere integrity.

At a glance

Figures

  1. 4-OHT-dependent induction of telomerase activity in TERT-ER cells.
    Figure 1: 4-OHT-dependent induction of telomerase activity in TERT-ER cells.

    a, Telomerase activity in eNSCs (*, telomerase products) (top); real-time quantification of reactions above (bottom). b, Representative G4TERT-ER splenocyte metaphases. c, Proliferation of adult G4TERT-ER fibroblasts (n = 3) in media with vehicle (black) or 4-OHT (red). d, Representative image of G4TERT-ER fibroblasts (passage 6) in media with 4-OHT (bottom) or vehicle (top). e, Signal-free ends in primary splenocyte metaphases, 15 metaphases per sample, n = 2 (*P<0.05). f, Mean telomere-FISH signal in primary splenocyte interphases, normalized to centromeric signal, n = 3 (***P<0.0001). Open bars correspond to vehicle-treated and filled bars to 4-OHT-treated, error bars represent s.d.

  2. Telomerase activation in adult TERT-ER mice.
    Figure 2: Telomerase activation in adult TERT-ER mice.

    ac, Representative images of tissues from experimental and control mice. a, Haematoxylin and eosin-stained sections of testes. b, Primary splenocytes stained for 53BP1. c, Small intestine sections stained for 53BP1. d, Testes weight of adult males (30–50-week-old, n10). e, 53BP1 nuclear foci per 100 nuclei (n = 3). f, 53BP1 nuclear foci per 100 crypts (n = 4). g, Litter sizes (n = 3); h, Spleen weights (n6). i, Apoptotic cells per 100 intestinal crypts (n20). ***P = 0.0001, **P<0.005, *P<0.05. Open bars correspond to vehicle-treated and filled bars to 4-OHT-treated groups, error bars represent s.d.

  3. Neural stem cell function following telomerase reactivation in vitro.
    Figure 3: Neural stem cell function following telomerase reactivation in vitro.

    ac, Representative images of experimental and control mice-derived NSCs. a, Secondary neurospheres. b, Differentiated NSCs stained with 53BP1 or c, GFAP and TUJ1 antibodies. d, Self-renewal capacity of secondary neurospheres (n = 4) ***P<0.0001, *P<0.001. e, 53BP1 nuclear foci per 100 cells (>400 nuclei per culture, n = 3). f, Multipotency (GFAP+/TUJ1+) of NSCs (n = 4; 308 wells per culture condition) **P = 0.0066. Scale bar, 100μm. Open bars correspond to vehicle-treated and filled bars to 4-OHT-treated groups, error bars represent s.d.

  4. NSC proliferation and differentiation following telomerase reactivation in vivo.
    Figure 4: NSC proliferation and differentiation following telomerase reactivation in vivo.

    NSC proliferation and neurogenesis were measured by Ki67, Sox-2 and Dcx expression in SVZ from experimental and control mice. Mature oligodendrocytes in the corpus callosum were stained with anti-Olig2 antibody. Equivalent coronal sections (n>10) were scored in a blinded fashion by laser scanning and plotted on the right panels. ×20 (SVZ) or ×40 (corpus callosum) objectives were used. *** P<0.0001, **P = 0.0022. Open bars correspond to vehicle-treated and filled bars to 4-OHT-treated groups, error bars represent s.d.

  5. Brain size, myelination, and olfactory function following telomerase reactivation.
    Figure 5: Brain size, myelination, and olfactory function following telomerase reactivation.

    a, Representative brains from age-matched experimental and control animals. b, Brain weights, n10, ***P = 0.0004, *P = 0.02. c, Representative electron micrographs of myelinated axonal tracts in corpus callosum, arrow heads indicate myelin sheath width (×12,000). Scale bars, 200nm. d, g ratios (inner/outer radii) (n = 2, >150 axons per mouse) ***P<0.0001. e, Representative tracings of experimental and control mice during 3-min exposure to water or 2-MB. f, g, Time spent in scent zone 3 with water or 2-MB for vehicle- or 4-OHT-treated G0TERT-ER (squares) and G4TERT-ER (circles) mice; n = 4. Error bars represent s.d., except in (d) (s.e.m.).

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Author information

Affiliations

  1. Belfer Institute for Applied Cancer Science and Departments of Medical Oncology, Medicine and Genetics, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Mariela Jaskelioff,
    • Florian L. Muller,
    • Ji-Hye Paik,
    • Emily Thomas,
    • Shan Jiang,
    • Ergun Sahin,
    • Maria Kost-Alimova,
    • Alexei Protopopov,
    • Juan Cadiñanos,
    • James W. Horner &
    • Ronald A. DePinho
  2. Division of Endocrinology, Diabetes & Metabolism, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA

    • Andrew C. Adams &
    • Eleftheria Maratos-Flier

Contributions

M.J. and R.A.D. designed and guided the research; M.J., F.L.M., J.-H.P., E.S., E.T., S.J. and M.K.-A. performed research. J.C. and J.W.H. generated the TERT-ER mouse. M.J., F.L.M., A.C.A., A.P., E.M.-F. and R.A.D. analysed data. M.J. and R.A.D. wrote the manuscript.

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The authors declare no competing financial interests.

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  1. Supplementary Information (751K)

    This file contains Supplementary Figures 1-4 with legends and Supplementary Tables 1-3.

Comments

  1. Report this comment #17235

    Rob Ord said:

    I believe that Robin Holiday (who also established the mechanism for DNA recombination) first proposed the 'loss-of chormosome'/ telomere model in a 1974 Fed. Proc. article, yet I never see him recognised. Is it now appropriate that he be recognised for his seminal contribution?

    Rob Ord PhD, LLB
    70 Russell St, Nelson 7010, New Zealand

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