Skip to main content
SpringerLink
Log in

Secretome of the preimplantation human embryo by bottom-up label-free proteomics

  • Original Paper
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript

Abstract

A bottom-up label-free mass spectrometric proteomic strategy was used to analyse the protein profiles of the human embryonic secretome. Culture media samples used for embryonic culture of patients undergoing intracytoplasmic sperm injection cycles were selected as a test case for this exploratory proof-of-principle study. The media were stored after embryo transfer and then pooled into positive (n = 8) and negative (n = 8) implantation groups. The absolute quantitative bottom-up technique employed a multidimensional protein identification technology based on separation by nano-ultra-high pressure chromatography and identification via tandem nano-electrospray ionization mass spectrometry with data-independent scanning in a hydrid QqTOF mass spectrometer. By applying quantitative bottom-up proteomics, unique proteins were found exclusively in both the positive- and negative-implantation groups, which suggest that competent embryos express and secrete unique biomarker proteins into the surrounding culture medium. The selective monitoring of these possible secretome biomarkers could make viable procedures using single-embryo transfer.

A bottom-up label-free proteomic mass spectrometric analysis of the human embryo secretome is described. This approach seems to allow quantification and identification of unique proteins related to positive- and negative-implantation groups, which can be further validated as biomarkers for selection and transfer of a single embryo.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  1. Palermo G, Joris H, Devroey P, Van Steirteghem AC (1992) Pregnancies after intracytoplasmic injection of single spermatozoon into an oocyte. Lancet 340:17–18

    Article  CAS  Google Scholar 

  2. Pandian Z, Bhattacharya S, Ozturk O, Serour G, Templeton A (2009) Number of embryos for transfer following in-vitro fertilisation or intra-cytoplasmic sperm injection. Cochrane Database Syst Rev 2:CD003416.

  3. Katz-Jaffe MG, McReynolds S, Gardner DK, Schoolcraft WB (2009) The role of proteomics in defining the human embryonic secretome. Mol Hum Reprod 15:271–277

    Article  CAS  Google Scholar 

  4. Hamel M, Dufort I, Robert C, Gravel C, Leveille MC, Leader A, Sirard MA (2008) Identification of differentially expressed markers in human follicular cells associated with competent oocytes. Hum Reprod 23:1118–1127

    Article  CAS  Google Scholar 

  5. Bromer JG, Seli E (2008) Assessment of embryo viability in assisted reproductive technology: shortcomings of current approaches and the emerging role of metabolomics. Curr Opin Obstet Gynecol 20:234–241

    Article  Google Scholar 

  6. Aydiner F, Yetkin CE, Seli E (2010) Perspectives on emerging biomarkers for non-invasive assessment of embryo viability in assisted reproduction. Curr Mol Med 10:206–215

    Article  CAS  Google Scholar 

  7. Katz-Jaffe MG, McCallie BR, Janesch A, Filipovits JA, Schoolcraft WB, Gardner DK (2010) Blastocysts from patients with polycystic ovaries exhibit altered transcriptome and secretome. Reprod Biomed Online 21:520–526

    Article  CAS  Google Scholar 

  8. Beardsley AJ, Li Y, O’Neill C (2010) Characterization of a diverse secretome generated by the mouse preimplantation embryo in vitro. Reprod Biol Endocrinol 8:71

    Article  Google Scholar 

  9. Katz-Jaffe MG, Schoolcraft WB, Gardner DK (2006) Analysis of protein expression (secretome) by human and mouse preimplantation embryos. Fertil Steril 86:678–685

    Article  CAS  Google Scholar 

  10. Katz-Jaffe MG, Gardner DK (2008) Symposium: innovative techniques in human embryo viability assessment. Can proteomics help to shape the future of human assisted conception? Reprod Biomed Online 17:497–501

    Article  CAS  Google Scholar 

  11. Katz-Jaffe MG, Gardner DK, Schoolcraft WB (2006) Proteomic analysis of individual human embryos to identify novel biomarkers of development and viability. Fertil Steril 85:101–107

    Article  CAS  Google Scholar 

  12. Chambery A, Colucci-D’Amato L, Vissers JP, Scarpella S, Langridge JI, Parente A (2009) Proteomic profiling of proliferating and differentiated neural mes-c-myc A1 cell line from mouse embryonic mesencephalon by LC-MS. J Proteome Res 8:227–238

    Article  CAS  Google Scholar 

  13. Levin Y, Wang L, Ingudomnukul E, Schwarz E, Baron-Cohen S, Palotas A, Bahn S (2009) Real-time evaluation of experimental variation in large-scale LC-MS/MS-based quantitative proteomics of complex samples. J Chromatogr B Analyt Technol Biomed Life Sci 877:1299–1305

    Article  CAS  Google Scholar 

  14. Patel VJ, Thalassinos K, Slade SE, Connolly JB, Crombie A, Murrell JC, Scrivens JH (2009) A comparison of labeling and label-free mass spectrometry-based proteomics approaches. J Proteome Res 8:3752–3759

    Article  CAS  Google Scholar 

  15. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  Google Scholar 

  16. Silva JC, Denny R, Dorschel CA, Gorenstein M, Kass IJ, Li GZ, McKenna T, Nold MJ, Richardson K, Young P, Geromanos S (2005) Quantitative proteomic analysis by accurate mass retention time pairs. Anal Chem 77:2187–2200

    Article  CAS  Google Scholar 

  17. Krämer-Albers E-M, Bretz N, Tenzer S, Winterstein C, Möbius W, Berger H, Nave K-A, Schild H, Trotter J (2007) Oligodendrocytes secrete exosomes containing major myelin and stress-protective proteins: Trophic support for axons? Proteomics Clin. Appl 1:1446–1461

    Google Scholar 

  18. Li GZ, Vissers JP, Silva JC, Golick D, Gorenstein MV, Geromanos SJ (2009) Database searching and accounting of multiplexed precursor and product ion spectra from the data independent analysis of simple and complex peptide mixtures. Proteomics 9:1696–1719

    Article  CAS  Google Scholar 

  19. Geromanos SJ, Vissers JP, Silva JC, Dorschel CA, Li GZ, Gorenstein MV, Bateman RH, Langridge JI (2009) The detection, correlation, and comparison of peptide precursor and product ions from data independent LC-MS with data dependant LC-MS/MS. Proteomics 9:1683–1695

    Article  CAS  Google Scholar 

  20. Chambery A, Vissers JP, Langridge JI, Lonardo E, Minchiotti G, Ruvo M, Parente A (2009) Qualitative and quantitative proteomic profiling of cripto(−/−) embryonic stem cells by means of accurate mass LC-MS analysis. J Proteome Res 8:1047–1058

    Article  CAS  Google Scholar 

  21. Delahunty CM, Yates JR (2007) MudPIT: multidimensional protein identification technology. Biotechniques 43:563, 565, 567 passim.

  22. Lo Turco EG, Souza GH, Garcia JS, Ferreira CR, Eberlin MN, Bertolla RP (2010) Effect of endometriosis on the protein expression pattern of follicular fluid from patients submitted to controlled ovarian hyperstimulation for in vitro fertilization. Hum Reprod 25:1755–1766

    Article  Google Scholar 

  23. Ferreira CR, Lo Turco EG, Saraiva SA, Bertolla RP, Perecin F, Souza GHFM, Murgu M, Garcia JS, Cortezzi SS, Meirelles FV, Klitzke CF, Cabral EC, Miglino MA, Porciuncula PM, Leal CLV EB Jr, Martins DdS, Ambrósio CE, D’Alexandri F, Smith LC, Eberlin MN (2010) Proteomics, metabolomis and lipidomics in reproductive biotechnologies: the MS solutions. Acta Scientiae Veterinariae 38:s277–s289

    Google Scholar 

  24. Dominguez F, Gadea B, Esteban FJ, Horcajadas JA, Pellicer A, Simon C (2008) Comparative protein-profile analysis of implanted versus non-implanted human blastocysts. Hum Reprod 23:1993–2000

    Article  CAS  Google Scholar 

  25. Kanka J (2003) Gene expression and chromatin structure in the pre-implantation embryo. Theriogenology 59:3–19

    Article  CAS  Google Scholar 

  26. Hamatani T, Carter MG, Sharov AA, Ko MS (2004) Dynamics of global gene expression changes during mouse preimplantation development. Dev Cell 6:117–131

    Article  CAS  Google Scholar 

  27. Misirlioglu M, Page GP, Sagirkaya H, Kaya A, Parrish JJ, First NL, Memili E (2006) Dynamics of global transcriptome in bovine matured oocytes and preimplantation embryos. Proc Natl Acad Sci U S A 103:18905–18910

    Article  CAS  Google Scholar 

  28. Toyoda M, Kojima M, Takeuchi T (2000) Jumonji is a nuclear protein that participates in the negative regulation of cell growth. Biochem Biophys Res Commun 274:332–336

    Article  CAS  Google Scholar 

  29. Takeuchi T, Watanabe Y, Takano-Shimizu T, Kondo S (2006) Roles of jumonji and jumonji family genes in chromatin regulation and development. Dev Dyn 235:2449–2459

    Article  CAS  Google Scholar 

  30. Pasini D, Cloos PA, Walfridsson J, Olsson L, Bukowski JP, Johansen JV, Bak M, Tommerup N, Rappsilber J, Helin K (2010) JARID2 regulates binding of the Polycomb repressive complex 2 to target genes in ES cells. Nature 464:306–310

    Article  CAS  Google Scholar 

  31. Modarressi MH, Cameron J, Taylor KE, Wolfe J (2001) Identification and characterisation of a novel gene, TSGA10, expressed in testis. Gene 262:249–255

    Article  CAS  Google Scholar 

  32. Behnam B, Modarressi MH, Conti V, Taylor KE, Puliti A, Wolfe J (2006) Expression of Tsga10 sperm tail protein in embryogenesis and neural development: from cilium to cell division. Biochem Biophys Res Commun 344:1102–1110

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We thank the Brazilian Science Foundations CNPQ, CAPES, FINEP, and FAPESP for financial assistance.

We thank to Kevin S. Kerian for English language review of this manuscript.

Author information

Authors and Affiliations

  1. Sapientiae Institute-Educational and Research Center in Assisted Reproduction, São Paulo, SP, Brazil, 04503-040

    Sylvia S. Cortezzi, Daniela P. A. F. Braga & Edson Borges Jr

  2. Institute of Exact Sciences, Alfenas Federal University, Alfenas, MG, Brazil, 37170-000

    Jerusa S. Garcia

  3. ThoMSon Mass Spectrometry Laboratory-Institute of Chemistry, University of Campinas-UNICAMP, Campinas, SP, Brazil, 13083-970

    Christina R. Ferreira & Marcos N. Eberlin

  4. Fertility-Assisted Fertilization Center, São Paulo, SP, Brazil, 01401-002

    Daniela P. A. F. Braga, Rita C. S. Figueira, Assumpto Iaconelli Jr & Edson Borges Jr

  5. Mass Spectrometry Applications Research and Development Laboratory, Waters Corporation, Barueri, SP, Brazil, 06455-020

    Gustavo H. M. F. Souza

Authors
  1. Sylvia S. Cortezzi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jerusa S. Garcia
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Christina R. Ferreira
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Daniela P. A. F. Braga
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Rita C. S. Figueira
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Assumpto Iaconelli Jr
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Gustavo H. M. F. Souza
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Edson Borges Jr
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Marcos N. Eberlin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Marcos N. Eberlin.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cortezzi, S.S., Garcia, J.S., Ferreira, C.R. et al. Secretome of the preimplantation human embryo by bottom-up label-free proteomics. Anal Bioanal Chem 401, 1331–1339 (2011). https://doi.org/10.1007/s00216-011-5202-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00216-011-5202-1

Keywords

Search

Navigation