Letter | Published:

Human oocytes reprogram adult somatic nuclei of a type 1 diabetic to diploid pluripotent stem cells

Nature volume 510, pages 533536 (26 June 2014) | Download Citation

Subjects

Abstract

The transfer of somatic cell nuclei into oocytes can give rise to pluripotent stem cells that are consistently equivalent to embryonic stem cells1,2,3, holding promise for autologous cell replacement therapy4,5. Although methods to induce pluripotent stem cells from somatic cells by transcription factors6 are widely used in basic research, numerous differences between induced pluripotent stem cells and embryonic stem cells have been reported7,8,9,10,11, potentially affecting their clinical use. Because of the therapeutic potential of diploid embryonic stem-cell lines derived from adult cells of diseased human subjects, we have systematically investigated the parameters affecting efficiency of blastocyst development and stem-cell derivation. Here we show that improvements to the oocyte activation protocol, including the use of both kinase and translation inhibitors, and cell culture in the presence of histone deacetylase inhibitors, promote development to the blastocyst stage. Developmental efficiency varied between oocyte donors, and was inversely related to the number of days of hormonal stimulation required for oocyte maturation, whereas the daily dose of gonadotropin or the total number of metaphase II oocytes retrieved did not affect developmental outcome. Because the use of concentrated Sendai virus for cell fusion induced an increase in intracellular calcium concentration, causing premature oocyte activation, we used diluted Sendai virus in calcium-free medium. Using this modified nuclear transfer protocol, we derived diploid pluripotent stem-cell lines from somatic cells of a newborn and, for the first time, an adult, a female with type 1 diabetes.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Accessions

Primary accessions

Gene Expression Omnibus

Data deposits

Microarray data are available at GEO under accession numbers GSE54849 and GSE54876.

References

  1. 1.

    et al. Epigenetic memory in induced pluripotent stem cells. Nature 467, 285–290 (2010)

  2. 2.

    , , & ES cells derived from cloned and fertilized blastocysts are transcriptionally and functionally indistinguishable. Proc. Natl Acad. Sci. USA 103, 933–938 (2006)

  3. 3.

    et al. Equivalency of nuclear transfer-derived embryonic stem cells to those derived from fertilized mouse blastocysts. Stem Cells 24, 2023–2033 (2006)

  4. 4.

    et al. Therapeutic cloning in individual parkinsonian mice. Nature Med. 14, 379–381 (2008)

  5. 5.

    , , , & Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy. Cell 109, 17–27 (2002)

  6. 6.

    et al. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131, 861–872 (2007)

  7. 7.

    et al. Differentiation-defective phenotypes revealed by large-scale analyses of human pluripotent stem cells. Proc. Natl Acad. Sci. USA 110, 20569–20574 (2013)

  8. 8.

    et al. Subtelomeric hotspots of aberrant 5-hydroxymethylcytosine-mediated epigenetic modifications during reprogramming to pluripotency. Nature Cell Biol. 15, 700–711 (2013)

  9. 9.

    et al. Incomplete DNA methylation underlies a transcriptional memory of somatic cells in human iPS cells. Nature Cell Biol. 13, 541–549 (2011)

  10. 10.

    et al. Identification of a specific reprogramming-associated epigenetic signature in human induced pluripotent stem cells. Proc. Natl Acad. Sci. USA 109, 16196–16201 (2012)

  11. 11.

    et al. Clone- and gene-specific aberrations of parental imprinting in human induced pluripotent stem cells. Stem Cells 27, 2686–2690 (2009)

  12. 12.

    et al. Human oocytes reprogram somatic cells to a pluripotent state. Nature 478, 70–75 (2011)

  13. 13.

    et al. The histone deacetylase inhibitor scriptaid enhances nascent mRNA production and rescues full-term development in cloned inbred mice. Reproduction 138, 309–317 (2009)

  14. 14.

    et al. Nuclear genome transfer in human oocytes eliminates mitochondrial DNA variants. Nature 493, 632–637 (2013)

  15. 15.

    Stability of cyclin B protein during meiotic maturation and the first mitotic cell division in mouse oocytes. Biol. Cell 89, 211–219 (1997)

  16. 16.

    , & Incorporation of amino acids during maturation in vitro by the mouse oocyte: effect of puromycin on protein synthesis. Biol. Reprod. 7, 341–346 (1972)

  17. 17.

    , & Human gene expression first occurs between the four- and eight-cell stages of preimplantation development. Nature 332, 459–461 (1988)

  18. 18.

    et al. Human embryonic stem cells derived by somatic cell nuclear transfer. Cell 153, 1228–1238 (2013)

  19. 19.

    et al. Derivation of cloned human blastocysts by histone deacetylase inhibitor treatment after somatic cell nuclear transfer with beta-thalassemia fibroblasts. Stem Cells Dev. 20, 1951–1959 (2011)

  20. 20.

    , , , & Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei. Nature 394, 369–374 (1998)

  21. 21.

    , , & A reliable, noninvasive technique for spindle imaging and enucleation of mammalian oocytes. Nature Biotechnol. 18, 223–225 (2000)

  22. 22.

    et al. Borealin: a novel chromosomal passenger required for stability of the bipolar mitotic spindle. J. Cell Biol. 166, 179–191 (2004)

  23. 23.

    , & Donor age is paramount to success in oocyte donation. Hum. Reprod. 14, 2755–2758 (1999)

  24. 24.

    et al. Controlled ovarian hyperstimulation: does prolonged stimulation justify cancellation of in vitro fertilization cycles? Gynecol. Endocrinol. 21, 232–234 (2005)

  25. 25.

    et al. Hotspots of aberrant epigenomic reprogramming in human induced pluripotent stem cells. Nature 471, 68–73 (2011)

  26. 26.

    et al. Somatic coding mutations in human induced pluripotent stem cells. Nature 471, 63–67 (2011)

  27. 27.

    et al. Reprogramming within hours following nuclear transfer into mouse but not human zygotes. Nature Commun. 2, 488 (2011)

  28. 28.

    & A protocol for embryonic stem cell derivation by somatic cell nuclear transfer into human oocytes. Protocol Exch (2014)

  29. 29.

    & Payment of egg donors in stem cell research in the USA. Reprod. Biomed. Online 18, 603–608 (2009)

  30. 30.

    et al. Impracticality of egg donor recruitment in the absence of compensation. Cell Stem Cell 9, 293–294 (2011)

  31. 31.

    et al. Egg sharing for research: a successful outcome for patients and researchers. Cell Stem Cell 10, 239–240 (2012)

  32. 32.

    The Ethics Committee of the American Society for Reproductive Medicine Financial compensation of oocyte donors. Fertil. Steril. 88, 305–309 (2007)

  33. 33.

    et al. Ethics. The ISSCR guidelines for human embryonic stem cell research. Science 315, 603–604 (2007)

  34. 34.

    , & Chromosome transfer in mature oocytes. Nature Protocols 5, 1138–1147 (2010)

  35. 35.

    & Nuclear transfer into mouse oocytes. J. Vis. Exp. 30, 116 (2006)

  36. 36.

    et al. Highly efficient neural conversion of human ES and iPS cells by dual inhibition of SMAD signaling. Nature Biotechnol. 27, 275–280 (2009)

  37. 37.

    et al. iPSC-derived beta cells model diabetes due to glucokinase deficiency. J. Clin. Invest. 123, 3146–3153 (2013)

  38. 38.

    et al. Beta cell dysfunction due to increased ER stress in a stem cell model of Wolfram syndrome. Diabetes 63, 923–933 (2014

Download references

Acknowledgements

This research was supported the New York Stem Cell Foundation (NYSCF) and a New York State Stem Cell Science (NYSTEM) IIRP Award no. C026184, and the Russell Berrie Foundation Program in Cellular Therapies of Diabetes. We thank S. Mitalipov for helpful discussions and providing reagents, S. Micucci for counting cells in S-phase, and Z. Hall for critical reading of the manuscript. D.E. is a NYSCF-Robertson Investigator.

Author information

Author notes

    • Mitsutoshi Yamada
    •  & Bjarki Johannesson

    These authors contributed equally to this work.

Affiliations

  1. The New York Stem Cell Foundation Research Institute, New York, New York 10032, USA

    • Mitsutoshi Yamada
    • , Bjarki Johannesson
    • , Daniel Paull
    • , Michael W. Nestor
    • , Susan L. Solomon
    •  & Dieter Egli
  2. Stem Cell Unit, Department of Genetics, Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel

    • Ido Sagi
    •  & Nissim Benvenisty
  3. Naomi Berrie Diabetes Center, Department of Pediatrics, College of Physicians and Surgeons, Columbia University, New York, New York 10032, USA

    • Lisa Cole Burnett
    • , Matthew Freeby
    • , Ellen Greenberg
    • , Robin S. Goland
    •  & Rudolph L. Leibel
  4. Center for Women’s Reproductive Care, College of Physicians and Surgeons, Columbia University, New York 10019, USA

    • Daniel H. Kort
    • , Robert W. Prosser
    •  & Mark V. Sauer
  5. Department of Obstetrics and Gynecology, College of Physicians and Surgeons, Columbia University, New York 10032, USA

    • Daniel H. Kort
    • , Robert W. Prosser
    •  & Mark V. Sauer

Authors

  1. Search for Mitsutoshi Yamada in:

  2. Search for Bjarki Johannesson in:

  3. Search for Ido Sagi in:

  4. Search for Lisa Cole Burnett in:

  5. Search for Daniel H. Kort in:

  6. Search for Robert W. Prosser in:

  7. Search for Daniel Paull in:

  8. Search for Michael W. Nestor in:

  9. Search for Matthew Freeby in:

  10. Search for Ellen Greenberg in:

  11. Search for Robin S. Goland in:

  12. Search for Rudolph L. Leibel in:

  13. Search for Susan L. Solomon in:

  14. Search for Nissim Benvenisty in:

  15. Search for Mark V. Sauer in:

  16. Search for Dieter Egli in:

Contributions

M.V.S. supervised the research oocyte donation program and retrieved oocytes. D.E. designed, performed and interpreted nuclear transfer experiments, derived ES cells with M.Y., and wrote the paper with input from all authors. M.Y. performed statistical analysis, M.Y. and B.J. performed stem cell characterization and differentiation, L.C.B. performed neuronal differentiation, I.S. and N.B. performed gene expression analysis, M.W.N. assisted with calcium experiments, D.H.K. performed data analysis, D.P. assisted in nuclear transfer experiments, R.W.P. collected developmental data of IVF embryos, M.F. made the skin biopsy, E.G. coordinated human subjects research, R.S.G. wrote the IRB protocol, R.L.L. contributed project planning, S.L.S. created the environment specifically for this work.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Dieter Egli.

Extended data

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nature13287

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.