Sperm DNA fragmentation : implications in assisted reproductive technologies

[摘要] ENGLISH ABSTRACT: Male fertility has for many years been defined in vitro as the ability of sperm to fertilizeoocytes and to obtain early cleavage-stage embryos. Spermatozoa comprise of anextraordinary high percentage of polyunsaturated fatty acids in their plasma membrane.Due to an extremely low content of cytoplasm, sperm cells have a particularly lowpotential to scavenge reactive oxygen species (ROS), and are therefore highly sensitiveto oxidative processes, which lead to sperm nucleus DNA damage/fragmentation.Normally, DNA fragmentation occurs in every ejaculate and can be induced by anexcessive ROS production of active leukocytes or the spermatozoa themselves. Underdistressed conditions, DNA fragmentation may also occur in the testis as a result ofoxidative processes in the apoptotic cascade. These DNA fragmentations can beregarded as late signs of programmed cell death (apoptosis).Clinically, DNA fragmentation in spermatozoa results in significantly decreasedimplantation and pregnancy rates especially in patients with oligo- and/orteratozoospermia. The p-pattern normal sperm morphology has been shown to givepoorer fertilization rates in vitro than the g- and n-patterns. In this study there isreported on the significant correlation found between the p-pattern normal spermmorphology and sperm DNA fragmentation as measured with the terminaldeoxynucleotidyl transferase-mediated dUDP-biotin end labeling (TUNEL) assay. Thisfinding further explains the lower fertility potential of patients presenting with p-patternnormal sperm morphology.In addition, this study explores the intricate relations between ROS in the semen, DNAfragmentation of the spermatozoa, as measured with the TUNEL assay and the spermchromatin structure assay (SCSA ), spermatozoa apoptotic status and spermparameters as measured with a standard semen analysis. Positive correlations werefound between ROS and the apoptotic status of the sperm, as well as between spermwith non-fragmented DNA and sperm concentration and percentage motility. Theresults emphasize the importance of sperm selection especially when the treatment ofchoice is intracytoplasmic sperm injection (ICSI). An early sign of programmed cell death, also known as apoptosis, is the externalizationof phosphatidylserine (PS) from the inner membrane leaflet to the outer leaflet. PSshows a high affinity to Annexin V. Apoptotic spermatozoa are able to fertilize oocytes,but embryo senescence may occur at the time when the paternal genes are activated.In this study there is reported on a novel method whereby spermatozoa can beseparated on the basis of their apoptotic status through flow cytometry. Results showedthat the normal sperm morphology, according to strict criteria, of the resultant nonapoptoticsperm fraction is significantly higher than that of the apoptotic counterpart.With refinement of this technique, it will be possible in future to use these separatednon-apoptotic sperm cells during ICSI for fertilization.From the above it is apparent that the spermatozoon has to play a vital role in thedevelopment of the embryo from fertilization to implantation and pregnancy. It is,however, important to note that besides the gametes, there are other critical factorswhich contribute to a successful in vitro fertilization (IVF) cycle, among these are the invitro culture conditions. In this regard, this study compared two sequential embryoculture systems. It was found that the more complex medium resulted in better daythree embryo quality and a better blastocyst formation rate and pregnancy rate.These findings highlight the importance of a holistic perspective towards the complexityof the factors involved in affecting embryo quality and pregnancy outcome.

[发布机构] Stellenbosch University