Abstract
Purpose
To provide a comprehensive analysis of mtDNA quantity in D5 and D6 blastocysts, as well as a further insight to the origin of delayed blastocyst development.
Methods
A retrospective cohort analysis of 829 D5 and 472 D6 blastocysts from 460 patients who underwent in vitro fertilization (IVF) with next-generation sequencing (NGS)–based preimplantation genetic testing for aneuploidy (PGT-A). The quantity of trophectoderm mtDNA was extrapolated from the NGS data, followed by the analysis of mean mtDNA levels between D5 and D6 blastocysts of the same ploidy (aneuploid/euploid) and transfer outcomes (positive/negative clinical pregnancy).
Results
D5 blastocysts had significantly higher euploidy rate and clinical pregnancy rate when compared with D6 blastocysts. The proportion of blastocysts derived from patients ≧ 40 years old were similar between the D5 and D6 cohorts. When blastocysts with identical ploidy were analyzed, the D5 cohorts all had significantly higher mean mtDNA levels than their D6 counterparts. Similarly, when embryo transfers with identical outcome were analyzed, the D5 cohorts also had significantly higher mean mtDNA levels than the D6 cohorts. Trophectoderm mtDNA level was independent of maternal age and blastocyst morphology grades.
Conclusions
Our data provided further evidence D5 blastocysts contained significantly greater mtDNA quantity than D6 blastocysts, and mtDNA quantity could be a key factor that affects the development rate of blastocysts. Furthermore, one must avoid using an arbitrary threshold when incorporating mtDNA quantity into the embryo selection criteria, as the observed value may have vastly different clinical implication when blastulation rate is also considered.
Similar content being viewed by others
Data availability
All data and material are available upon request.
References
Bourdon M, Pocate-Cheriet K, Finet de Bantel A, Grzegorczyk-Martin V, Amar Hoffet A, Arbo E, et al. Day 5 versus day 6 blastocyst transfers: a systematic review and meta-analysis of clinical outcomes. Hum Reprod. 2019;34:1948–64.
Barash OO, Ivani KA, Willman SP, Rosenbluth EM, Wachs DS, Hinckley MD, et al. Association between growth dynamics, morphological parameters, the chromosomal status of the blastocysts, and clinical outcomes in IVF PGS cycles with single embryo transfer. J Assist Reprod Genet. 2017;34:1007–16.
Irani M, O’Neill C, Palermo GD, Xu K, Zhang C, Qin X, et al. Blastocyst development rate influences implantation and live birth rates of similarly graded euploid blastocysts. Fertil Steril. 2018;110:95–102.e1.
Hashimoto S, Amo A, Hama S, Ito K, Nakaoka Y, Morimoto Y. Growth retardation in human blastocysts increases the incidence of abnormal spindles and decreased implantation potential after verification. Hum Reprod. 2013;28:1528–35.
Fragouli E, Spath K, Alfarawati S, Kaper F, Craig A, Michel CE, et al. Altered levels of mitochondrial DNA are associated with female age, aneuploidy, and provide an independent measure of embryonic implantation potential. PLoS Genet. 2015;11:e1005241.
Diez-Juan A, Rubio C, Marin C, Martinez S, Al-Asmar N, Riboldi M, et al. Mitochondrial DNA content as a viability score in human euploid embryos: less is better. Fertil Steril. 2015;104:534–41.
Ravichandran K, McCaffrey C, Grifo J, Morales A, Perloe M, Munne S, et al. Mitochondrial DNA quantification as a tool for embryo viability assessment: retrospective analysis of data from single euploid blastocysts transfers. Hum Reprod. 2017;32:1282–92.
Fragouli E, McCaffrey C, Ravichandran K, Spath K, Grifo JA, Munné S, et al. Clinical implications of mitochondrial DNA quantification on pregnancy outcomes: a blinded prospective non-selection study. Hum Reprod. 2017;32:2340–7.
Lledo B, Ortiz JA, Morales R, García-Hernández E, Ten J, Bernabeu A, et al. Comprehensive mitochondrial DNA analysis and IVF outcome. Hum Reprod Open. 2018;2018(4):hoy023.
Victor AR, Brake AJ, Tyndall JC, Griffin DK, Zouves CG, Barnes FL, et al. Accurate quantitation of mitochondrial DNA reveals uniform levels in human blastocysts irrespective of ploidy, age, or implantation potential. Fertil Steril. 2017;107:34–42.
Treff NR, Zhan Y, Tao X, Olcha M, Han M, Rajchel J, et al. Levels of trophectoderm mitochondrial DNA do not predict the reproductive potential of sibling embryos. Hum Reprod. 2017;32:954–62.
Klimczak AM, Pacheco LE, Lewis KE, Massahi N, Richards JP, Kearns WG, et al. Embryonal mitochondrial DNA: relationship to embryo quality and transfer outcomes. J Assist Reprod Genet. 2018;35:871–7.
Barnes FL, Victor AR, Zouves CG, Viotti M. Mitochondrial DNA quantitation-making sense of contradictory reports. Hum Reprod. 2017;32:2149–50.
Wells D, Ravichandran K, McCaffrey C, Grifo J, Morales A, Perloe M, et al. Reply: mitochondrial DNA quantification-the devil in the detail. Hum Reprod. 2017;32:2150–1.
Gardner DK, Schoolcraft WB. In vitro culture of human blastocyst. In: Jansen R, Mortimer D, editors. Towards reproductive certainty: infertility and genetics beyond 1999. Carnforth: Parthenon Press; 1999. p. 378–88.
Li H, Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics. 2009;25:1754–60.
Ding J, Sidore C, Butler TJ, Wing MK, Qian Y, Meirelles O, et al. Assessing mitochondrial DNA variation and copy number in lymphocytes of ∼2,000 sardinians using tailored sequencing analysis tools. PLoS Genet. 2015;11:e1005306.
Barbuscia A, Martikainen P, Myrskylä M, Remes H, Somigliana E, Klemetti R, et al. Maternal age and risk of low birth weight and premature birth in children conceived through medically assisted reproduction. Evidence from Finnish population registers. Hum Reprod. 2020;35:212–20.
May-Panloup P, Boucret L, Chao de la Barca JM, Desquiret-Dumas V, Ferré-L'Hotellier V, Morinière C, et al. Ovarian ageing: the role of mitochondria in oocytes and follicles. Hum Reprod Update. 2016;22:725–43.
Brown WM. Polymorphism in mitochondrial DNA of humans as revealed by restriction endonuclease analysis. Proc Natl Acad Sci U S A. 1980;77:3605–9.
Thundathil J, Filion F, Smith LC. Molecular control of mitochondrial function in preimplantation mouse embryos. Mol Reprod Dev. 2005;71:405–13.
May-Panloup P, Vignon X, Chretien MF, Heymen Y, Tamassia M, Malthiery Y, et al. Increase of mitochondrial DNA content and transcripts in early bovine embryogenesis associated with upregulation of mtTFA and NRF1 transcription factors. Reprod Biol Endocrinol. 2005;3:65.
Spikings EC, Alderson J, St John JC. Regulated mitochondrial DNA replication during oocyte maturation is essential for successful porcine embryonic development. Biol Reprod. 2007;76:327–35.
Ho JR, Arrach N, Rhodes-Long K, Salem W, McGinnis LK, Chung K, et al. Blastulation timing is associated with differential mitochondrial content in euploid embryos. J Assist Reprod Genet. 2018;35(4):711–20.
Hashimoto S, Morimoto N, Yamanaka M, Matsumoto H, Yamochi T, Goto H, et al. Quantitative and qualitative changes of mitochondrial in human preimplantation embryos. J Assist Reprod Genet. 2017;34:573–80.
Van Blerkom J. Mitochondrial function in the human oocyte and embryo and their role in developmental competence. Mitochondrion. 2011;11:797–813.
Chan CC, Liu VW, Lau EY, Yeung WS, Ng EH, Ho PC. Mitochondrial DNA content and 4977 bp deletion in unfertilized oocytes. Mol Hum Reprod. 2005;11:843–6.
Duran HE, Simsek-Duran F, Oehninger SC, Jones HW Jr, Castora FJ. The association of reproductive senescence with mitochondrial quantity, function, and DNA integrity in human oocytes at different stages of maturation. Fertil Steril. 2011;96:384–8.
May-Panloup P, Chretien MF, Jacques C, Vasseur C, Malthiery Y, Reynier P. Low oocyte mitochondrial DNA content in ovarian insufficiency. Hum Reprod. 2005;20:593–7.
Murakoshi Y, Sueoka K, Takahashi K, Sato S, Sakurai T, Tajima H, et al. Embryo developmental capability and pregnancy outcome are related to the mitochondrial DNA copy number and ooplasmic volume. J Assist Reprod Genet. 2013;30:1367–75.
Reynier P, May-Panloup P, Chretien MF, Morgan CJ, Jean M, Savagner F, et al. Mitochondrial DNA content affects the fertilizability of human oocytes. Mol Hum Reprod. 2001;7:425–9.
Santos TA, El Shourbagy S, St John JC. Mitochondrial content reflects oocyte variability and fertilization outcome. Fertil Steril. 2006;85:584–91.
Lee SE, Kim EY, Choi HY, Moon JJ, Park MJ, Lee JB, et al. Rapamycin rescues the poor developmental capacity of aged porcine oocytes. Asian-Australas J Anim Sci. 2014;27:635–47.
Zhang XM, Li L, Xu JJ, Wang N, Liu WJ, Lin XH, et al. Rapamycin preserves the follicle pool reserve and prolongs the ovarian lifespan of female rats via modulating mTOR activation and sirtuin expression. Gene. 2013;523:82–7.
Liu M, Yin Y, Ye X, Zeng M, Zhao Q, Keefe DL, et al. Resveratrol protects against age-associated infertility in mice. Hum Reprod. 2013;28:707–17.
Sugiyama M, Kawahara-Miki R, Kawana H, Shirasuna K, Kuwayama T, Iwata H. Resveratrol-induced mitochondrial synthesis and autophagy in oocytes derived from early antral follicles of aged cows. J Reprod Dev. 2015;61:251–9.
Stojkovic M, Westesen K, Zakhartchenko V, Stojkovic P, Boxhammer K, Wolf E. Coenzyme Q(10) in submicron-sized dispersion improves development, hatching, cell proliferation, and adenosine triphosphate content of in vitro-produced bovine embryos. Biol Reprod. 1999;61:541–7.
Ben-Meir A, Burstein E, Borrego-Alvarez A, Chong J, Wong E, Yavorska T, et al. Coenzyme Q10 restores oocyte mitochondrial function and fertility during reproductive aging. Aging Cell. 2015;14:887–95.
Zhang Y, Zhang C, Shu J, Guo J, Chang HM, Leung PCK, et al. Adjuvant treatment strategies in ovarian stimulation for poor responders undergoing IVF: a systematic review and network meta-analysis. Hum Reprod Update. 2020;28(26):247–63.
Alexeyev M, Shokolenko I, Wilson G, LeDoux S. The maintenance of mitochondrial DNA integrity–critical analysis and update. Cold Spring Harb Perspect Biol. 2013;5:a012641.
Leese HJ. Quiet please, do not disturb: a hypothesis of embryo metabolism and viability. Bioessays. 2002;24:845–9.
Van Blerkom J, Davis P, Lee J. ATP content of human oocytes and developmental potential and outcome after in vitro fertilization and embryo transfer. Hum Reprod. 1995;10:415–24.
Liu H, Trimarchi J, Keefe D. Involvement of mitochondria in oxidative stress induced cell death in mouse zygotes. Biol Reprod. 2000;62:1745–53.
Morimoto N, Hashimoto S, Yamanaka M, Nakano T, Satoh M, Nakaoka Y, et al. Mitochondrial oxygen consumption rate of human embryos declines with maternal age. J Assist Reprod Genet. 2020;37:1815–21.
Simon AL, Kiehl M, Fischer E, Proctor JG, Bush MR, Givens C, et al. Pregnancy outcomes from more than 1,800 in vitro fertilization cycles with the use of 24-chromosome single-nucleotide polymorphism-based preimplantation genetic testing for aneuploidy. Fertil Steril. 2018;110:113–21.
Vinals Gonzalez X, Odia R, Naja R, Serhal P, Saab W, Seshadri S, et al. Euploid blastocysts implant irrespective of their morphology after NGS-(PGT-A) testing in advanced maternal age patients. J Assist Reprod Genet. 2019;36:1623–9.
Viotti M, Victor AR, Zouves CG, Barnes FL. Is mitochondrial DNA quantitation in blastocyst trophectoderm cells predictive of developmental competence and outcome in clinical IVF? J Assist Reprod Genet. 2017;34:1581–5.
Acknowledgments
The authors would like to thank all of the patients who participated in the present study, as well as the IVF laboratory team members at IHMED Fertility center for their assistance.
Author information
Authors and Affiliations
Contributions
Study design and coordination: Frank Shao-Ying Wu and Ni-Chung Lee
Subject recruitment: Shao-Peng Weng, Meng-Shun Shen, and Pei-Chun Ma
Data collection: Shao-Peng Weng, Meng-Shun Shen, and Po-Kuan Wu
Data analysis: Frank Shao-Ying Wu, Ni-Chung Lee, Po-Kuan Wu, and Pei-Chun Ma
Drafting of manuscript: Frank Shao-Ying Wu
Study supervision and manuscript revision: Ni-Chung Lee
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflicts of interest.
Ethics approval
The study protocol was approved by the institutional review board of the National Taiwan University Hospital (IRB #201905053RIND).
Consent to participate
Not applicable due to the retrospective nature of the study.
Consent for publication
The authors transfer to Springer the publication rights and give full consent for all of the information about to be published in Journal of Assisted Reproduction and Genetics.
Code availability
Not applicable.
Additional information
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Wu, F.SY., Weng, SP., Shen, MS. et al. Suboptimal trophectoderm mitochondrial DNA level is associated with delayed blastocyst development. J Assist Reprod Genet 38, 587–594 (2021). https://doi.org/10.1007/s10815-020-02045-5
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10815-020-02045-5