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Electrospray mass spectrometry analysis of blastocoel fluid as a potential tool for bovine embryo selection

  • Embryo Biology
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Abstract

Purpose

The aim of this study was to analyze the metabolic profiles of blastocoel fluid (BF) obtained from bovine embryos produced in vivo and in vitro.

Methods

Expanded blastocysts (20/group) that were in vitro and in vivo derived at day 7 were used. BF was collected and analyzed under direct infusion conditions using a microTOF-Q® mass spectrometer with electrospray ionization and a mass range of 50–650 m/z.

Results

The spectrometry showed an evident difference in the metabolic profiles of BF from in vivo and in vitro produced embryos. These differences were very consistent between the samples of each group suggesting that embryo fluids can be used to identify the origin of the embryo. Ions 453.15 m/z, 437.18 m/z, and 398.06 m/z were identified as biomarkers for the embryo’s origin with 100% sensitivity and specificity. Although it was not possible to unveil the molecular identity of the differential ions, the resulting spectrometric profiles provide a phenotype capable of differentiating embryos and hence constitute a potential parameter for embryo selection.

Conclusion

To the best of our knowledge, our results showed, for the first time, an evident difference between the spectrometric profiles of the BF from bovine embryos produced in vivo and in vitro.

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References

  1. Petersen B. Basics of genome editing technology and its application in livestock species. Reproduction in Domestic Animals. 2017;52(S3):4–13. https://doi.org/10.1111/rda.13012.

    Article  CAS  PubMed  Google Scholar 

  2. Viana JH. 2018 Statistics of embryo production and transfer in domestic farm animals. Embryo Technology Newsletter. 2019;36(4):14.

    Google Scholar 

  3. Ealy AD, Wooldridge LK, SR MC. BOARD INVITED REVIEW: Post-transfer consequences of in vitro-produced embryos in cattle. Journal of Animal Science. 2019;97(6):2555–68. https://doi.org/10.1093/jas/skz116.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Corcoran D, Fair T, Park S, Rizos D, Patel OV, Smith GW, et al. Suppressed expression of genes involved in transcription and translation in in vitro compared with in vivo cultured bovine embryos. Reproduction. 2006;131(4):651–60. https://doi.org/10.1530/rep.1.01015.

    Article  CAS  PubMed  Google Scholar 

  5. Fair T, Lonergan P, Dinnyes A, Cottell DC, Hyttel P, Ward FA, et al. Ultrastructure of bovine blastocysts following cryopreservation: effect of method of blastocyst production. Molecular Reproduction and Development. 2001;58(2):186–95. https://doi.org/10.1002/1098-2795(200102)58:2<186::aid-mrd8>3.0.co;2-n.

    Article  CAS  PubMed  Google Scholar 

  6. Rizos D, Fair T, Papadopoulos S, Boland MP, Lonergan P. Developmental, qualitative, and ultrastructural differences between ovine and bovine embryos produced in vivo or in vitro. Molecular Reproduction and Development. 2002;62(3):320–7. https://doi.org/10.1002/mrd.10138.

    Article  CAS  PubMed  Google Scholar 

  7. Sudano MJ, Santos VG, Tata A, Ferreira CR, Paschoal DM, Machado R, et al. Phosphatidylcholine and sphingomyelin profiles vary in Bos taurus indicus and Bos taurus taurus in vitro- and in vivo-produced Blastocysts1. Biology of Reproduction. 2012;87(130):1–11. https://doi.org/10.1095/biolreprod.112.102897.

    Article  CAS  Google Scholar 

  8. Machado GM, Ferreira AR, Pivato I, Fidelis A, Spricigo JF, Paulini F, et al. Post-hatching development of in vitro bovine embryos from day 7 to 14 in vivo versus in vitro. Molecular Reproduction and Development. 2013;80(11):936–47. https://doi.org/10.1002/mrd.22230.

    Article  CAS  PubMed  Google Scholar 

  9. Noguchi T, Aizawa T, Munakata Y, Iwata H. Comparison of gene expression and mitochondria number between bovine blastocysts obtained in vitro and in vivo. J Reprod Dev. 2020;66(1):35–9. https://doi.org/10.1262/jrd.2019-100.

    Article  CAS  PubMed  Google Scholar 

  10. Peterson AJ, Lee RSF. Improving successful pregnancies after embryo transfer. Theriogenology. 2003;59(2):687–97. https://doi.org/10.1016/S0093-691X(02)01248-7.

    Article  CAS  PubMed  Google Scholar 

  11. Muñoz M, Uyar A, Correia E, Díez C, Fernandez-Gonzalez A, Caamaño JN, et al. Prediction of pregnancy viability in bovine in vitro-produced embryos and recipient plasma with Fourier transform infrared spectroscopy. Journal of Dairy Science. 2014;97(9):5497–507. https://doi.org/10.3168/jds.2014-8067.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Seli E, Vergouw CG, Morita H, Botros L, Roos P, Lambalk CB, et al. Noninvasive metabolomic profiling as an adjunct to morphology for noninvasive embryo assessment in women undergoing single embryo transfer. Fertility and Sterility. 2010;94(2):535–42. https://doi.org/10.1016/j.fertnstert.2009.03.078.

    Article  PubMed  Google Scholar 

  13. Leese HJ. Metabolism of the preimplantation embryo. 40 years on. 2012;143(4):417. https://doi.org/10.1530/rep-11-0484.

    Article  CAS  Google Scholar 

  14. Vera-Rodriguez M, Diez-Juan A, Jimenez-Almazan J, Martinez S, Navarro R, Peinado V, et al. Origin and composition of cell-free DNA in spent medium from human embryo culture during preimplantation development. Human Reproduction. 2018;33(4):745–56. https://doi.org/10.1093/humrep/dey028.

    Article  CAS  PubMed  Google Scholar 

  15. Juliano CS, Ana Clara FCMÁ, Hannah LG, Jason EB, Quinton AW, Gerrit JB. Cell-secreted vesicles containing microRNAs as regulators of gamete maturation. Journal of Endocrinology. 2018;236(1):R15–27. https://doi.org/10.1530/joe-17-0200.

    Article  CAS  Google Scholar 

  16. Palini S, Galluzzi L, De Stefani S, Bianchi M, Wells D, Magnani M, et al. Genomic DNA in human blastocoele fluid. Reproductive BioMedicine Online. 2013;26(6):603–10. https://doi.org/10.1016/j.rbmo.2013.02.012.

    Article  CAS  PubMed  Google Scholar 

  17. Leaver M, Wells D. Non-invasive preimplantation genetic testing (niPGT): the next revolution in reproductive genetics? Human Reproduction Update. 2020;26(1):16–42. https://doi.org/10.1093/humupd/dmz033.

    Article  CAS  PubMed  Google Scholar 

  18. Gopichandran N, Leese H. Metabolic characterization of the bovine blastocyst, inner cell mass, trophectoderm and blastocoel fluid. Reproduction (Cambridge, England). 2003;126:299–308. https://doi.org/10.1530/reprod/126.3.299.

    Article  CAS  Google Scholar 

  19. D'Alessandro A, Federica G, Palini S, Bulletti C, Zolla L. A mass spectrometry-based targeted metabolomics strategy of human blastocoele fluid: a promising tool in fertility research. Molecular BioSystems. 2012;8(4):953–8. https://doi.org/10.1039/c1mb05358b.

    Article  CAS  PubMed  Google Scholar 

  20. Jensen PL, Beck HC, Petersen J, Hreinsson J, Wånggren K, Laursen SB, et al. Proteomic analysis of human blastocoel fluid and blastocyst cells. Stem Cells and Development. 2012;22(7):1126–35. https://doi.org/10.1089/scd.2012.0239.

    Article  CAS  PubMed  Google Scholar 

  21. Watson AJ, Barcroft LC. Regulation of blastocyst formation. Front Biosci. 2001;6:d708. https://doi.org/10.2741/watson.

    Article  CAS  PubMed  Google Scholar 

  22. Jensen PL, Grøndahl ML, Beck HC, Petersen J, Stroebech L, Christensen ST, et al. Proteomic analysis of bovine blastocoel fluid and blastocyst cells. Systems Biology in Reproductive Medicine. 2014;60(3):127–35. https://doi.org/10.3109/19396368.2014.894152.

    Article  CAS  PubMed  Google Scholar 

  23. Farra C, Choucair F, Awwad J. Non-invasive pre-implantation genetic testing of human embryos: an emerging concept. Human Reproduction. 2018;33(12):2162–7. https://doi.org/10.1093/humrep/dey314.

    Article  CAS  PubMed  Google Scholar 

  24. Magli MC, Pomante A, Cafueri G, Valerio M, Crippa A, Ferraretti AP, et al. Preimplantation genetic testing: polar bodies, blastomeres, trophectoderm cells, or blastocoelic fluid? Fertility and Sterility. 2016;105(3):676–83.e5. https://doi.org/10.1016/j.fertnstert.2015.11.018.

    Article  PubMed  Google Scholar 

  25. Tobler KJ, Zhao Y, Ross R, Benner AT, Xu X, Du L, et al. Blastocoel fluid from differentiated blastocysts harbors embryonic genomic material capable of a whole-genome&#xa0;deoxyribonucleic acid amplification and comprehensive chromosome microarray analysis. Fertility and Sterility. 2015;104(2):418–25. https://doi.org/10.1016/j.fertnstert.2015.04.028.

    Article  CAS  PubMed  Google Scholar 

  26. Hammond ER, Shelling AN, Cree LM. Nuclear and mitochondrial DNA in blastocoele fluid and embryo culture medium: evidence and potential clinical use. Human Reproduction. 2016;31(8):1653–61. https://doi.org/10.1093/humrep/dew132.

    Article  CAS  PubMed  Google Scholar 

  27. Zhang Y, Li N, Wang L, Sun H, Ma M, Wang H, et al. Molecular analysis of DNA in blastocoele fluid using next-generation sequencing. Journal of Assisted Reproduction and Genetics. 2016;33(5):637–45. https://doi.org/10.1007/s10815-016-0667-7.

    Article  PubMed  PubMed Central  Google Scholar 

  28. Rule K, Chosed RJ, Arthur Chang T, David Wininger J, Roudebush WE. Relationship between blastocoel cell-free DNA and day-5 blastocyst morphology. Journal of Assisted Reproduction and Genetics. 2018;35(8):1497–501. https://doi.org/10.1007/s10815-018-1223-4.

    Article  PubMed  PubMed Central  Google Scholar 

  29. Kuznyetsov V, Madjunkova S, Antes R, Abramov R, Motamedi G, Ibarrientos Z, et al. Evaluation of a novel non-invasive preimplantation genetic screening approach. PLoS One. 2018;13(5):e0197262–e. https://doi.org/10.1371/journal.pone.0197262.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Li P, Song Z, Yao Y, Huang T, Mao R, Huang J, et al. Preimplantation genetic screening with spent culture medium/blastocoel fluid for in vitro fertilization. Scientific Reports. 2018;8(1):9275. https://doi.org/10.1038/s41598-018-27367-4.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Tšuiko O, Zhigalina DI, Jatsenko T, Skryabin NA, Kanbekova OR, Artyukhova VG, et al. Karyotype of the blastocoel fluid demonstrates low concordance with both trophectoderm and inner cell mass. Fertility and Sterility. 2018;109(6):1127–34.e1. https://doi.org/10.1016/j.fertnstert.2018.02.008.

    Article  PubMed  Google Scholar 

  32. Vajta G, Kuwayama M. Improving cryopreservation systems. Theriogenology. 2006;65(1):236–44. https://doi.org/10.1016/j.theriogenology.2005.09.026.

    Article  CAS  PubMed  Google Scholar 

  33. Min SH, Kim JW, Lee YH, Park SY, Jeong PS, Yeon JY, et al. Forced collapse of the blastocoel cavity improves developmental potential in cryopreserved bovine blastocysts by slow-rate freezing and vitrification. Reproduction in Domestic Animals. 2014;49(4):684–92. https://doi.org/10.1111/rda.12354.

    Article  CAS  PubMed  Google Scholar 

  34. Diaz F, Bondiolli K, Paccamonti D, Gentry GT. Cryopreservation of Day 8 equine embryos after blastocyst micromanipulation and vitrification. Theriogenology. 2016;85(5):894–903. https://doi.org/10.1016/j.theriogenology.2015.10.039.

    Article  CAS  PubMed  Google Scholar 

  35. Kazemi P, Dashtizad M, Shamsara M, Mahdavinezhad F, Hashemi E, Fayazi S, et al. Effect of blastocoel fluid reduction before vitrification on gene expression in mouse blastocysts. Molecular Reproduction and Development. 2016;83(8):735–42. https://doi.org/10.1002/mrd.22681.

    Article  CAS  PubMed  Google Scholar 

  36. Ochota M, Wojtasik B, Niżański W. Survival rate after vitrification of various stages of cat embryos and blastocyst with and without artificially collapsed blastocoel cavity. Reproduction in Domestic Animals. 2017;52(S2):281–7. https://doi.org/10.1111/rda.12826.

    Article  CAS  PubMed  Google Scholar 

  37. Tedeschi G, Albani E, Borroni EM, Parini V, Brucculeri AM, Maffioli E, et al. Proteomic profile of maternal-aged blastocoel fluid suggests a novel role for ubiquitin system in blastocyst quality. Journal of Assisted Reproduction and Genetics. 2017;34(2):225–38. https://doi.org/10.1007/s10815-016-0842-x.

    Article  PubMed  Google Scholar 

  38. Ruggeri RR, Watanabe Y, Meirelles F, Bressan FF, Frantz N, Bos-Mikich A. The use of parthenotegenetic and IVF bovine blastocysts as a model for the creation of human embryonic stem cells under defined conditions. Journal of Assisted Reproduction and Genetics. 2012;29(10):1039–43. https://doi.org/10.1007/s10815-012-9866-z.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Santos RR, Schoevers EJ, Roelen BAJ. Usefulness of bovine and porcine IVM/IVF models for reproductive toxicology. Reprod Biol Endocrinol. 2014;12:117. https://doi.org/10.1186/1477-7827-12-117.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Marei WFA, Van den Bosch L, Pintelon I, Mohey-Elsaeed O, Bols PEJ, Leroy JLMR. Mitochondria-targeted therapy rescues development and quality of embryos derived from oocytes matured under oxidative stress conditions: a bovine in vitro model. Human Reproduction. 2019;34(10):1984–98. https://doi.org/10.1093/humrep/dez161.

    Article  CAS  PubMed  Google Scholar 

  41. Parrish JJ, Krogenaes A, Susko-Parrish JL. Effect of bovine sperm separation by either swim-up or Percoll method on success of in vitro fertilization and early embryonic development. Theriogenology. 1995;44(6):859–69. https://doi.org/10.1016/0093-691X(95)00271-9.

    Article  CAS  PubMed  Google Scholar 

  42. Machado GM, Carvalho JO, Filho ES, Caixeta ES, Franco MM, Rumpf R, et al. Effect of Percoll volume, duration and force of centrifugation, on in vitro production and sex ratio of bovine embryos. Theriogenology. 2009;71(8):1289–97. https://doi.org/10.1016/j.theriogenology.2009.01.002.

    Article  CAS  PubMed  Google Scholar 

  43. Holm P, Booth PJ, Schmidt MH, Greve T, Callesen H. High bovine blastocyst development in a static in vitro production system using sofaa medium supplemented with sodium citrate and myo-inositol with or without serum-proteins. Theriogenology. 1999;52(4):683–700. https://doi.org/10.1016/S0093-691X(99)00162-4.

    Article  CAS  PubMed  Google Scholar 

  44. Magli MC, Albanese C, Crippa A, Tabanelli C, Ferraretti AP, Gianaroli L. Deoxyribonucleic acid detection in blastocoelic fluid: a new predictor of embryo ploidy and viable pregnancy. Fertility and Sterility. 2019;111((1)):77–85. https://doi.org/10.1016/j.fertnstert.2018.09.016.

    Article  CAS  PubMed  Google Scholar 

  45. Gianaroli L, Magli MC, Pomante A, Crivello AM, Cafueri G, Valerio M, et al. Blastocentesis: a source of DNA for preimplantation genetic testing. Results from a pilot study. Fertility and Sterility. 2014;102(6):1692–9.e6. https://doi.org/10.1016/j.fertnstert.2014.08.021.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We would like to thank the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brazil (CAPES) for the scholarship granted to Fernandes GO. This research was only possible due to the assistance of the research team of the Mass Spectrometry Laboratory at Embrapa Genetic Resource and Dr. Marcelo Henrique Soller Ramada, University Catholica de Brasília. The authors thank Dr. José de Lima Cardozo Filho for his valuable advice regarding the analysis and his support during the laboratory work. The authors would also like to thank Dr. Carlos Frederico Martins and Dr. Andrei Fidelis (DVM).

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Funding

The Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Brazil for the scholarship of Fernandes GO. The Embrapa Recursos Genéticos e Biotecnologia, Brazil. The Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Brazil.

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Authors and Affiliations

  1. School of Agriculture and Veterinary Medicine, University of Brasilia, Brasília, DF, Brazil

    Gabriela de Oliveira Fernandes, Otávio Augusto Costa de Faria & Margot Alves Nunes Dode

  2. Laboratory of Mass Spectrometry, Embrapa Genetic Resources and Biotechnology, Brasília, DF, Brazil

    Daniel Nogoceke Sifuentes

  3. Laboratory of Animal Reproduction, Embrapa Genetic Resources and Biotechnology, Brasília, DF, Brazil

    Maurício Machaim Franco & Margot Alves Nunes Dode

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  1. Gabriela de Oliveira Fernandes
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  2. Otávio Augusto Costa de Faria
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  3. Daniel Nogoceke Sifuentes
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  4. Maurício Machaim Franco
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  5. Margot Alves Nunes Dode
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Contributions

All authors contributed to the study conception and design. Material preparation, data collection, and analysis were performed by Gabriela de Oliveira Fernandes, Otávio Augusto Costa de Faria, Daniel Nogoceke Sifuentes, and Maurício Machaim Franco. The first draft of the manuscript was written by Gabriela de Oliveira Fernandes and Margot Alves Nunes Dode and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Margot Alves Nunes Dode.

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Ethics approval

The experiment was approved by the Animal Use Ethics Committee of Embrapa Genetic Resources and Biotechnology (CEUA/CENARGEN) protocol number 004/2017.

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All authors agreed to participate in this work.

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All authors included in this study agree with its publication.

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The authors declare no competing interests.

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de Oliveira Fernandes, G., de Faria, O.A.C., Sifuentes, D.N. et al. Electrospray mass spectrometry analysis of blastocoel fluid as a potential tool for bovine embryo selection. J Assist Reprod Genet 38, 2209–2217 (2021). https://doi.org/10.1007/s10815-021-02189-y

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