Balasuriya, AS; (2013) An Investigation of DNA Integrity Biomarkers in Gametogenesis and Pre-Implantation Embryo Development to Predict Reproductive Potential. Doctoral thesis (PhD), UCL (University College London).

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Abstract

Unsuccessful fertilization, aberrant embryo development, implantation failure and recurrent miscarriages can occur despite any obvious reasons during assisted reproduction. DNA fragmentation and aneuploidy in gametes have been implicated as a possible cause for infertility, but the use of DNA fragmentation tests is controversial. The SCD-FISH test claims to analyse DNA fragmentation and aneuploidy in the same cell. In this thesis SCD-FISH was compared to single FISH and SCD, which showed that SCD-FISH was an unreliable method to study these parameters. Sperm preparation techniques in assisted reproduction technologies (ART) potentially generate exogenous stresses that cause additional DNA damage. This study subjected mature sperm to environmental insults that normally occur during ART, and highlighted the significant increase in sperm DNA fragmentation due to heat, freezing and oxidative stress. Since it is not possible to investigate the level of DNA damage in the egg or sperm, and still maintain its viability for use in fertilization, DNA damage in cumulus and granulosa cells were studied. There was no relationship between DNA fragmentation in these cells and fertilization or pregnancy outcome. DNA fragmentation was significantly higher in cumulus than granulosa cells. Allegedly minimising DNA damage, GM-CSF is a component added to IVF culture media. Its effect on murine embryo DNA fragmentation was studied and it was found to have no significant effect. The exposure of human embryonic stem cells to pre-tested toxins and its impact on DNA fragmentation and aneuploidy, and the correlation between these two parameters were also investigated. These studies emphasise the belief that the introduction of DNA fragmentation assays to the clinical arena is premature as its role is unsubstantiated, the lack of a transparent relationship between DNA fragmentation and pregnancy outcome and the importance of minimising damage to sperm and embryos due to external stresses during laboratory research and ART. Overall, the aim of this thesis was to examine the hypotheses that DNA fragmentation, aneuploidy and phosphatidylserine translocation are potential biomarkers of reproductive potential that exist in variable degrees of at different stages of gametogenesis and preimplantation embryo development, and are susceptible to environmental stresses.

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