Impact of Antioxidant Treatment on DNA Fragmentation Index

In the Western world, approximately 15-20% of couples experience infertility problems during their reproductive life. In this context ´'infertility' is defined as 12 months of unprotected intercourse without getting pregnant.

Although there is limited access to reliable diagnostic methods, it is assumed that in at least 50% of all cases 'male subfertility' is a contributing or the main cause of the infertility of the couple. The gold standard in assessment of male reproductive function is standard semen analysis including evaluation of sperm number, motility and morphology. Although efforts have been made in to improve and standardise the methodology for semen analysis, there are no well established cut off levels for sperm concentration, motility or morphology which accurately predict the chance of pregnancy or indicate which assisted reproductive technique (ART) - the most frequent therapy for infertility- will ultimately be the most effective. This lack of diagnostic tools is not only hampering proper management of infertility but also represents a serious limitation in relation to the understanding of biological underpinnings of the disorder and as a consequence, the development of cause related treatment modalities.

Although various novel alternative methods of assessing semen quality have been developed, so far, none have contributed to or altered our clinical approach to the treatment of infertile couples. During the passed two decades a lot of attention has been paid to impairment of sperm DNA integrity as a possible cause of male subfertility. There are different techniques currently available for the assessment of sperm DNA integrity e.g. Sperm Chromatin Structure Assay (SCSA®), Comet, TUNEL and sperm chromatin dispersion test. Generally, the use of these tests has shown an increased proportion of sperm with fragmented DNA structure among subfertile men as compared to proven fertile or men from general population. Although measuring different characteristics of the status of sperm DNA (e.g. presence of single and double strand breaks, propensity to damage and probable cause of the fragmentation), comparisons of the results obtained with two or more of these tests show a correlation coefficient of a magnitude 0.4 to 0.6, indicating that, although not completely overlapping, the tests to a certain extent assess some of the same characteristics of sperm DNA. Another interesting finding is that there is only a weak to moderate correlation between standard sperm parameters and measures of DNA integrity, the level of association (correlation coefficient \~ 0.6) the most pronounced being for motility. This means that assessment of sperm chromatin integrity adds to the information obtained by standard semen analysis.

The most appraised, standardised and studied of the sperm nDNA tests is the SCSA® which is based on a very well defined protocol for handling of samples and subsequent data analysis. The robustness of the techniques was exemplified by a comparison of the SCSA® analysis of almost 200 samples results performed by two independent laboratories which found a correlation coefficient of 0.9 with mean difference between the DNA Fragmentation Index (DFI) results of about 1%. A further advantage of the SCSA® method over the remaining assessment techniques is the relatively large number of studies that have utilised the technique in clinical settings constituting a considerable record of the method's performance in predicting fertility outcome.

Briefly, these data indicate that the chance of spontaneous pregnancy decreases at DFI levels above 20% and approaches zero if DFI exceeds 30%. This is also true for Intrauterine insemination (IUI). However, it seems that even spermatozoa from samples with high DFI can be used for in vitro fertilisation by standard IVF or ICSI, with some data suggesting that ICSI might actually be more efficient using samples with high (>30%) DFI. A finding, which may at first appear paradoxical but may be a reflection of the relative probability of fortuitously selecting a sperm with intact DNA. The data regarding fertilisation rate, embryo quality and risk of miscarriage in relation to DFI are also conflicting.

The biological mechanisms responsible for DNA strand breaks and other types of impairment of sperm DNA integrity are not completely known. Amongst others incomplete DNA repair during spermiogenesis, abortive apoptosis and/or increased level of oxidative stress have been suggested as possible options .

The overall aim of this project is to evaluate effect of restoring oxidant balance using Androferti, (Q Pharma Laboratories; Alicante, Spain) on sperm DNA integrity in subfertile men with high level of DFI.