Mammalian Genome

, Volume 20, Issue 9–10, pp 664–673 | Cite as

Effect of IVF and laser zona dissection on DNA methylation pattern of mouse zygotes

  • Dominika D. Peters
  • Konstantin Lepikhov
  • Karsten Rodenacker
  • Susan Marschall
  • Auke Boersma
  • Peter Hutzler
  • Hagen Scherb
  • Jörn Walter
  • Martin Hrabé de Angelis
Article

Abstract

In vitro fertilization (IVF) and zona pellucida laser microdissection-facilitated IVF (Laser-IVF) are presently routine procedures in human assisted reproduction. The safety of these methods at the epigenetic level is not fully understood. Studies on mouse Laser-IVF embryos provide evidence that the use of Laser-IVF leads to reduced birth rate, indicating a potential harm of this technique for the embryo. Hence, the aim of this study was to examine the difference in DNA methylation pattern between IVF- and Laser-IVF-derived mouse zygotes. We examined two experimental groups of C3HeB/FeJ oocytes: (1) zona-intact and (2) laser-microdissected oocytes that were fertilized in vitro with freshly collected spermatozoa. Zygotes were fixed 5, 8, and 12 h after fertilization, and indirect immunofluorescence staining was studied using an anti-5-methylcytidine (5-MeC) antibody. The fluorescence intensities of paternal and maternal pronuclei were evaluated using the computer-assisted analysis of digital images. In addition, we performed a semiquantitative RT-PCR analysis of the presence of transcripts of three developmental marker genes, Oct4, Dab2, and Dnmt3b, in IVF- and Laser-IVF-derived blastocysts. We observed no significant differences in methylation status of the paternal genome and in the transcripts of the developmental marker genes after IVF and Laser-IVF. In conclusion, epigenetic patterns and early embryonic development are not altered by laser-assisted IVF techniques and another explanation must be sought for the poor implantation rates observed in mice.

References

  1. Adenot PG, Mercier Y, Renard JP, Thompson EM (1997) Differential H4 acetylation of paternal and maternal chromatin precedes DNA replication and differential transcriptional activity in pronuclei of 1-cell mouse embryos. Development 124:4615–4625PubMedGoogle Scholar
  2. Boersma A, Marschall S, Hrabe de Angelis M (2007) Laser-assisted IVF—an alternative approach for successful cryopreservation of mutant mouse lines on C57BL/6 background. International Mouse Genome Conference, Kyoto, Japan, October 28 - November 1, 2007, P77Google Scholar
  3. Depypere HT, McLaughlin KJ, Seamark RF, Warnes GM, Matthews CD (1988) Comparison of zona cutting and zona drilling as techniques for assisted fertilization in the mouse. J Reprod Fertil 84:205–211PubMedCrossRefGoogle Scholar
  4. Doherty AS, Mann MR, Tremblay KD, Bartolomei MS, Schultz RM (2000) Differential effects of culture on imprinted H19 expression in the preimplantation mouse embryo. Biol Reprod 62:1526–1535CrossRefPubMedGoogle Scholar
  5. el-Danasouri I, Westphal LM, Neev Y, Gebhardt J, Louie D et al (1993) Zona opening with 308 nm XeCl excimer laser improves fertilization by spermatozoa from long-term vasectomized mice. Hum Reprod 8:464–466PubMedGoogle Scholar
  6. Enginsu ME, Schutze K, Bellanca S, Pensis M, Campo R et al (1995) Micromanipulation of mouse gametes with laser microbeam and optical tweezers. Hum Reprod 10:1761–1764PubMedGoogle Scholar
  7. Fulka H, Fulka J (2006) No differences in the DNA methylation pattern in mouse zygotes produced in vivo, in vitro, or by intracytoplasmic sperm injection. Fertil Steril 86:1534–1536CrossRefPubMedGoogle Scholar
  8. Germond M, Nocera D, Senn A, Rink K, Delacretaz G et al (1995) Microdissection of mouse and human zona pellucida using a 1.48-microns diode laser beam: efficacy and safety of the procedure. Fertil Steril 64:604–611PubMedGoogle Scholar
  9. Germond M, Nocera D, Senn A, Rink K, Delacretaz G et al (1996) Improved fertilization and implantation rates after non-touch zona pellucida microdrilling of mouse oocytes with a 1.48 microm diode laser beam. Hum Reprod 11:1043–1048PubMedGoogle Scholar
  10. Hirasawa R, Sasaki H (2009) Dynamic transition of Dnmt3b expression in mouse pre- and early post-implantation embryos. Gene Expr Patterns 9:27–30CrossRefPubMedGoogle Scholar
  11. Hollis A, Rastegar S, Descloux L, Delacretaz G, Rink K (1997) Zona pellucida microdrilling with a 1.48-micron diode laser. IEEE Eng Med Biol Mag 16:43–47CrossRefPubMedGoogle Scholar
  12. Howlett SK, Reik W (1991) Methylation levels of maternal and paternal genomes during preimplantation development. Development 113:119–127PubMedGoogle Scholar
  13. Hrabe de Angelis M, Balling R (1998) Large scale ENU screens in the mouse: genetics meets genomics. Mutat Res 400:25–32PubMedGoogle Scholar
  14. Hrabe de Angelis MH, Flaswinkel H, Fuchs H, Rathkolb B, Soewarto D et al (2000) Genome-wide, large-scale production of mutant mice by ENU mutagenesis. Nat Genet 25:444–447CrossRefPubMedGoogle Scholar
  15. Kawase Y, Iwata T, Ueda O, Kamada N, Tachibe T et al (2002) Effect of partial incision of the zona pellucida by piezo-micromanipulator for in vitro fertilization using frozen-thawed mouse spermatozoa on the developmental rate of embryos transferred at the 2-cell stage. Biol Reprod 66:381–385CrossRefPubMedGoogle Scholar
  16. Kawase Y, Aoki Y, Kamada N, Jishage K, Suzuki H (2004) Comparison of fertility between intracytoplasmic sperm injection and in vitro fertilization with a partial zona pellucida incision by using a piezo-micromanipulator in cryopreserved inbred mouse spermatozoa. Contemp Top Lab Anim Sci 43:21–25PubMedGoogle Scholar
  17. Khosla S, Dean W, Brown D, Reik W, Feil R (2001a) Culture of preimplantation mouse embryos affects fetal development and the expression of imprinted genes. Biol Reprod 64:918–926CrossRefPubMedGoogle Scholar
  18. Khosla S, Dean W, Reik W, Feil R (2001b) Culture of preimplantation embryos and its long-term effects on gene expression and phenotype. Hum Reprod Update 7:419–427CrossRefPubMedGoogle Scholar
  19. Kishigami S, Van Thuan N, Hikichi T, Ohta H, Wakayama S et al (2006) Epigenetic abnormalities of the mouse paternal zygotic genome associated with microinsemination of round spermatids. Dev Biol 289:195–205CrossRefPubMedGoogle Scholar
  20. Landel CP (2005) Archiving mouse strains by cryopreservation. Lab Anim 34:50–57CrossRefGoogle Scholar
  21. Liow SL, Bongso A, Ng SC (1996) Fertilization, embryonic development and implantation of mouse oocytes with one or two laser-drilled holes in the zona, and inseminated at different sperm concentrations. Hum Reprod 11:1273–1280PubMedGoogle Scholar
  22. Marschall S, Hrabe de Angelis M (1999) Cryopreservation of mouse spermatozoa: double your mouse space. Trends Genet 15:128–131CrossRefPubMedGoogle Scholar
  23. Marschall S, Huffstadt U, Balling R, Hrabe de Angelis M (1999) Reliable recovery of inbred mouse lines using cryopreserved spermatozoa. Mamm Genome 10:773–776CrossRefPubMedGoogle Scholar
  24. Mayer W, Niveleau A, Walter J, Fundele R, Haaf T (2000) Demethylation of the zygotic paternal genome. Nature 403:501–502CrossRefPubMedGoogle Scholar
  25. McGrath J, Solter D (1984) Completion of mouse embryogenesis requires both the maternal and paternal genomes. Cell 37:179–183CrossRefPubMedGoogle Scholar
  26. Nagy A, Gertsenstein M, Vintersten K, Behringer R (2003) Manipulating the mouse embryo. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
  27. Nakagata N (2000) Cryopreservation of mouse spermatozoa. Mamm Genome 11:572–576CrossRefPubMedGoogle Scholar
  28. Nakagata N, Takeshima T (1993) Cryopreservation of mouse spermatozoa from inbred and F1 hybrid strains. Jikken Dobutsu 42:317–320PubMedGoogle Scholar
  29. Nakagata N, Okamoto M, Ueda O, Suzuki H (1997) Positive effect of partial zona-pellucida dissection on the in vitro fertilizing capacity of cryopreserved C57BL/6 J transgenic mouse spermatozoa of low motility. Biol Reprod 57:1050–1055CrossRefPubMedGoogle Scholar
  30. Nicklas W, Baneux P, Boot R, Decelle T, Deeny AA (2002) Recommendations for the health monitoring of rodent and rabbit colonies in breeding and experimental units. Lab Anim 36:20–42CrossRefPubMedGoogle Scholar
  31. Nolan PM, Peters J, Strivens M, Rogers D, Hagan J et al (2000) A systematic, genome-wide, phenotype-driven mutagenesis programme for gene function studies in the mouse. Nat Genet 25:440–443CrossRefPubMedGoogle Scholar
  32. Obruca A, Strohmer H, Sakkas D, Menezo Y, Kogosowski A et al (1994) Use of lasers in assisted fertilization and hatching. Hum Reprod 9:1723–1726PubMedGoogle Scholar
  33. Oswald J, Engemann S, Lane N, Mayer W, Olek A et al (2000) Active demethylation of the paternal genome in the mouse zygote. Curr Biol 10:475–478CrossRefPubMedGoogle Scholar
  34. Payne D (1995) Micro-assisted fertilization. Reprod Fertil Dev 7:831–839CrossRefPubMedGoogle Scholar
  35. Peters DD, Marschall S, Mahabir E, Boersma A, Heinzmann U et al (2006) Risk assessment of mouse hepatitis virus infection via in vitro fertilization and embryo transfer by the use of zona-intact and laser-microdissected oocytes. Biol Reprod 74:246–252CrossRefPubMedGoogle Scholar
  36. Quinn P, Kerin JF, Warnes GM (1985) Improved pregnancy rate in human in vitro fertilization with the use of a medium based on the composition of human tubal fluid. Fertil Steril 44:493–498PubMedGoogle Scholar
  37. Reik W, Walter J (2001) Genomic imprinting: parental influence on the genome. Nat Rev Genet 2:21–32CrossRefPubMedGoogle Scholar
  38. Reik W, Dean W, Walter J (2001) Epigenetic reprogramming in mammalian development. Science 293:1089–1093CrossRefPubMedGoogle Scholar
  39. Ribas RC, Taylor JE, McCorquodale C, Mauricio AC, Sousa M et al (2006) Effect of zona pellucida removal on DNA methylation in early mouse embryos. Biol Reprod 74:307–313CrossRefPubMedGoogle Scholar
  40. Santos F, Hendrich B, Reik W, Dean W (2002) Dynamic reprogramming of DNA methylation in the early mouse embryo. Dev Biol 241:172–182CrossRefPubMedGoogle Scholar
  41. Scholer HR, Dressler GR, Balling R, Rohdewohld H, Gruss P (1990) Oct-4: a germline-specific transcription factor mapping to the mouse t-complex. EMBO J 9:2185–2195PubMedGoogle Scholar
  42. Szczygiel MA, Kusakabe H, Yanagimachi R, Whittingham DG (2002) Intracytoplasmic sperm injection is more efficient than in vitro fertilization for generating mouse embryos from cryopreserved spermatozoa. Biol Reprod 67:1278–1284CrossRefPubMedGoogle Scholar
  43. Sztein JM, Farley JS, Mobraaten LE (2000) In vitro fertilization with cryopreserved inbred mouse sperm. Biol Reprod 63:1774–1780CrossRefPubMedGoogle Scholar
  44. Thornton CE, Brown SD, Glenister PH (1999) Large numbers of mice established by in vitro fertilization with cryopreserved spermatozoa: implications and applications for genetic resource banks, mutagenesis screens, and mouse backcrosses. Mamm Genome 10:987–992CrossRefPubMedGoogle Scholar
  45. Wrenzycki C, Herrmann D, Carnwath JW, Niemann H (1999) Alterations in the relative abundance of gene transcripts in preimplantation bovine embryos cultured in medium supplemented with either serum or PVA. Mol Reprod Dev 53(1):8–18CrossRefPubMedGoogle Scholar
  46. Yanagimachi R (1998) Intracytoplasmic sperm injection experiments using the mouse as a model. Hum Reprod 13(Suppl 1):87–98PubMedGoogle Scholar
  47. Yang DH, Smith ER, Roland IH, Sheng Z, He J et al (2002) Disabled-2 is essential for endodermal cell positioning and structure formation during mouse embryogenesis. Dev Biol 251:27–44CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Dominika D. Peters
    • 1
  • Konstantin Lepikhov
    • 3
  • Karsten Rodenacker
    • 2
  • Susan Marschall
    • 1
  • Auke Boersma
    • 1
  • Peter Hutzler
    • 4
  • Hagen Scherb
    • 2
  • Jörn Walter
    • 3
  • Martin Hrabé de Angelis
    • 1
  1. 1.Institute of Experimental GeneticsHelmholtz Zentrum München, German Research Center for Environmental Health (GmbH)NeuherbergGermany
  2. 2.Institute of Biomathematics and BiometryHelmholtz Zentrum München, German Research Center for Environmental Health (GmbH)NeuherbergGermany
  3. 3.Department of Natural Sciences – Technical Faculty III FR 8.3, Biological Sciences, Institute of Genetics/EpigeneticsUniversity of SaarlandSaarbrückenGermany
  4. 4.Institute of PathologyHelmholtz Zentrum München, German Research Center for Environmental Health (GmbH)NeuherbergGermany

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