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Abstract

Maternally inherited mitochondrial (mt)DNA mutations can cause fatal or severely debilitating syndromes in children1,2,3, with disease severity dependent on the specific gene mutation and the ratio of mutant to wild-type mtDNA (heteroplasmy) in each cell and tissue4. Pathogenic mtDNA mutations are relatively common, with an estimated 778 affected children born each year in the United States5. Mitochondrial replacement therapies or techniques (MRT) circumventing mother–to–child mtDNA disease transmission involve replacement of oocyte maternal mtDNA6,7,8. Here we report MRT outcomes in several families with common mtDNA syndromes. The mother’s oocytes were of normal quality and mutation levels correlated with those in existing children. Efficient replacement of oocyte mutant mtDNA was performed by spindle transfer8, resulting in embryos containing >99% donor mtDNA. Donor mtDNA was stably maintained in embryonic stem cells (ES cells) derived from most embryos. However, some ES cell lines demonstrated gradual loss of donor mtDNA and reversal to the maternal haplotype. In evaluating donor–to–maternal mtDNA interactions, it seems that compatibility relates to mtDNA replication efficiency rather than to mismatch or oxidative phosphorylation dysfunction. We identify a polymorphism within the conserved sequence box II region of the D-loop as a plausible cause of preferential replication of specific mtDNA haplotypes. In addition, some haplotypes confer proliferative and growth advantages to cells. Hence, we propose a matching paradigm for selecting compatible donor mtDNA for MRT.

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Change history

  • 27 February 2019

    In this Letter, there are several errors regarding the assignments of mtDNA haplotypes for a subset of egg donors from our study. These errors have not been corrected online.

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Acknowledgements

The authors acknowledge the OHSU Embryonic Stem Cell Research Oversight Committee and the Institutional Review Board for oversight and guidance. We thank all study participants for tissue donations and the Women’s Health Research Unit staff, University Fertility Consultants and the Reproductive Endocrinology and Infertility Division in the Department of Obstetrics and Gynecology, Oregon Health and Science University for support and procurement of human gametes. Studies were supported by the Leducq Foundation, OHSU institutional funds and Cincinnati Children’s Hospital Research Foundation. Work in the laboratory of J.C.I.B. was supported by the G. Harold and Leila Y. Mathers Charitable Foundation, the Leona M. and Harry B. Helmsley Charitable Trust and the Moxie Foundation. A.P.L. was partially supported by a fellowship from the Hewitt Foundation. P.M.R. was partially supported by a fellowship from Fundación Alfonso Martín Escudero.

Author information

Author notes

    • Eunju Kang
    •  & Yeonmi Lee

    Present address: Stem Cell Center, ASAN Institute for Life Sciences, ASAN Medical Center, Olympic-ro 43-gil, Songpa-gu, Seoul 138-736, South Korea.

Affiliations

  1. Center for Embryonic Cell and Gene Therapy, Oregon Health & Science University, 3303 SW Bond Avenue, Portland, Oregon 97239, USA

    • Eunju Kang
    • , Nuria Marti Gutierrez
    • , Amy Koski
    • , Rebecca Tippner-Hedges
    • , Hong Ma
    • , Yeonmi Lee
    • , Tomonari Hayama
    • , Crystal Van Dyken
    • , Riffat Ahmed
    • , Ying Li
    • , Dongmei Ji
    • , Don P. Wolf
    •  & Shoukhrat Mitalipov
  2. Division of Reproductive & Developmental Sciences, Oregon National Primate Research Center, Oregon Health & Science University, 505 NW 185th Avenue, Beaverton, Oregon 97006, USA

    • Eunju Kang
    • , Nuria Marti Gutierrez
    • , Amy Koski
    • , Rebecca Tippner-Hedges
    • , Hong Ma
    • , Yeonmi Lee
    • , Tomonari Hayama
    • , Crystal Van Dyken
    • , Riffat Ahmed
    • , Ying Li
    • , Don P. Wolf
    •  & Shoukhrat Mitalipov
  3. Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines Road, La Jolla, California 92037, USA

    • Jun Wu
    • , Aida Platero-Luengo
    • , Paloma Martinez-Redondo
    •  & Juan Carlos Izpisua Belmonte
  4. Department of Cell Biology School of Osteopathic Medicine, Rowan University, 2 Medical Center Drive, Stratford, New Jersey 08084, USA

    • Karen Agaronyan
    •  & Dmitry Temiakov
  5. Division of Human Genetics, Cincinnati Children’s Hospital Medical Center, 3333 Burnet Avenue, Cincinnati, Ohio 45229, USA

    • Xinjian Wang
    • , Shiyu Luo
    •  & Taosheng Huang
  6. Reproductive Medical Centre, Anhui Medical University, No 218, Jixi Rd, Shushan District, Heifei, Anhui 230022, China

    • Dongmei Ji
  7. IviGen Los Angeles, 406 Amapola Avenue, Suite 215, Torrance, California 90501, USA

    • Refik Kayali
    •  & Cengiz Cinnioglu
  8. Research Cytogenetics Laboratory, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, USA

    • Susan Olson
  9. Division of Reproductive Endocrinology, Department of Obstetrics and Gynecology, Oregon Health & Science University, 3303 SW Bond Avenue, Portland, Oregon 97239, USA

    • Jeffrey Jensen
    • , David Battaglia
    • , David Lee
    • , Diana Wu
    • , Paula Amato
    •  & Shoukhrat Mitalipov
  10. Knight Cardiovascular Institute, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland, Oregon 97239, USA

    • Shoukhrat Mitalipov
  11. Department of Biomedical Engineering, Oregon Health & Science University, 3303 SW Bond Avenue, Portland, Oregon 97239, USA

    • Shoukhrat Mitalipov

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Contributions

E.K., N.M.G., P.A. and S.M. conceived the study and designed the experiments. T.Hu., P.A. and A.K. evaluated clinical genetic results and coordinated recruitment of study participants. P.A. and J.J. performed skin biopsies and blood collections. P.A., D.B., D.L. and D.W. performed ovarian stimulations and oocyte retrievals. N.M.G., R.T.H. and S.M. conducted spindle transfer, intracytoplasmic sperm injection, embryo culture and establishment of ES cell lines. E.K., R.T.H., H.M., Y.Le., Y.Li, R.A., T.Ha. and N.M.G. cultured ES cells. E.K., C.V.D., Y.Le. and T.Ha. performed teratoma studies. E.K., C.V.D., Y.Li and D.J. performed mitochondrial complex assays. J.W., P.M.R. and A.P.L. performed in vitro differentiation and Seahorse assays. K.A. and D.T. performed mtDNA transcription experiments. E.K., C.V.D., R.T.H. and A.K. prepared mtDNA and performed MiSeq assays. E.K. and Y.Le. performed NGS data analysis and interpretation. E.K., C.V.D. validated mtDNA mutations by Sanger sequencing. H.M. and R.T.H. performed ARMS–qPCR. X.W., S.L. and T.Hu. performed CNV data analysis. R.K. and C.C. performed the comparative genome hybridization array. S.O. performed G-banding karyotypes. E.K., J.W., D.P.W., J.C.I.B., P.A. and S.M. analysed data and wrote the paper.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Paula Amato or Shoukhrat Mitalipov.

Reviewer Information Nature thanks E. Shoubridge and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Extended data

Supplementary information

Excel files

  1. 1.

    Supplementary Table 1

    Sequence differences between egg donor haplotypes understudy by whole mtDNA sequencing (syn, synonymous; non-syn, nonsynonymous; frmshft, frameshift).

  2. 2.

    Supplementary Table 2

    mtDNA variants in oocytes and somatic tissues from mutant carriers and children and healthy oocyte donors (syn, synonymous; non-syn,non- synonymous; frmshft, frameshift).

  3. 3.

    Supplementary Table 3

    Maternal mtDNA analysis in ESCs derived from ST blastocysts (syn, synonymous; non-syn, non- synonymous; frmshft, frameshift).

  4. 4.

    Supplementary Table 4

    Analysis of maternal mtDNA heteroplasmy in ESCs derived from somatic cell nuclear transfer (SCNT) blastocysts (syn, synonymous; nonsyn, non- synonymous; frmshft, frameshift).

  5. 5.

    Supplementary Table 5

    Maternal mtDNA heteroplasmy changes in 18 ST-ES and 8NT-ES cell lines. Filtered reads were aligned to the human mitochondrial sequence reference NC_012920.1 followed by variant calling. Presented nucleotide positions indicate difference from the reference or between different human haplotypes.

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DOI

https://doi.org/10.1038/nature20592

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