Sperm DNA integrity is vital for fertilization and the development of healthy descendants, while it affects natural and assisted reproduction equally.
During the final stages of spermatogenesis, each sperm cell undergoes extensive molecular remodeling of its nucleus to compress and protect its genetic material. Possible failure of cellular mechanisms during sperm maturation, as well as oxidative stress in the environment of spermatogenesis, can cause breaks in one or both DNA strands compacted in the sperm heads. The presence of breaks in the DNA strands is called sperm DNA fragmentation and can result in chromosomal alterations and genome instability.
More specifically, there are three main mechanisms that cause sperm DNA fragmentation and these are (1) failure of programmed cell death resulting in sperm with fragmented DNA being released at ejaculation, (2) defective maturation of chromatin in which potential nicks in sperm DNA are not repaired before condensation making the genetic material more susceptible to exogenous factors and (3) increased production of reactive oxygen forms which directly affects chromatin quality by causing DNA fragmentation.
Sperm DNA fragmentation affects all stages of human reproduction. In particular, a direct relationship between sperm DNA damage and male infertility has been established with a meta-analysis of the data published to date suggesting that increased levels of segmentation (>20%) are predictive of male infertility with increased sensitivity (79%) and specificity (86%) (Santi et al., 2018). It has also been demonstrated that men with unexplained infertility have increased levels of DNA fragmentation compared to fertile men and that 1 in 4 men with unexplained infertility have a sperm DNA fragmentation rate greater than 20%. In addition, data have been presented suggesting that men with oligoasthenozoospermia have significantly higher rates of DNA fragmentation than men who had normal sperm counts, motility and morphology in their semen analysis.
Different meta-analyses of existing data from qualitative studies provide clear evidence as to the effect of sperm DNA fragmentation on reproductive outcomes; either through natural conception or through assisted reproduction by the route of intrauterine insemination or IVF. According to this metadata, increased sperm DNA fragmentation rates (1) reduce the probability of natural conception, (2) reduce the probability of pregnancy and birth through intrauterine insemination, (3) reduce fertilization rates, implantation and pregnancy but also negatively affect embryo quality in conventional in vitro fertilization (IVF) procedures; (4) negatively affect the quality of embryos derived from intracytoplasmic sperm injection (ICSI); and (5) are associated with a higher risk of miscarriage or spontaneous abortion after either natural conception or assisted conception (IVF and ICSI). It is important to note that emerging literature has reported that sperm DNA fragmentation in addition to its negative impact on embryonic development may also affect the health and well-being of descendants (Aitken RJ, 2017).
Several clinical and environmental factors can negatively affect the integrity of sperm DNA. Studies have shown that oxidative stress caused by an imbalance between the production of reactive oxygen species and the overall antioxidant capacity of sperm causes fragmentation of sperm genetic material and this can derive from both endogenous and exogenous factors.
Lifestyle factors such as smoking, alcohol, unbalanced diet, exposure to toxic substances, chemicals and radiation can significantly affect sperm genetic integrity. Men with varicocele have significantly higher levels of DNA fragmentation in their sperm, and there is an increase in scrotal temperature of 2-4 oC while at the same time oxidative stress is introduced into the process of spermatogenesis through apoptosic processes and correspondingly the proteins secreted. Obesity is another factor causing an increase in scrotal temperature and indeed an increased body weight may contribute to poorer reproductive outcomes.
Infections and inflammation of the urogenital tract can also trigger an increase in reactive oxygen species and lead to oxidative stress through the activation of white blood cells. Male ageing is an additional factor influencing the quality of sperm genetic material, as progressively older men show cumulative exposure to oxidative stress and reduced antioxidant capacity, which has been confirmed by scientific evidence, as an increased rate of DNA fragmentation is observed in the sperm of older men.
Indications and methods of assessing DNA fragmentation in semen
The most recent edition of the World Health Organization’s Laboratory Manual for Semen Testing (2021) notes that DNA fragmentation represents an important addition to the assessment of male fertility and is a promising biomarker in clinical andrology. The European Society for Human Reproduction and Embryology (ESHRE) recommends the assessment of sperm DNA fragmentation in cases of spontaneous abortions, while the European Association of Urology (EAU) additionally suggests that the test should also be performed in men with unexplained infertility. The European Academy of Andrology (EAU) has proposed the inclusion of DNA fragmentation in the baseline tests for men with oligoasthenozoospermia prior to the implementation of assisted reproduction protocols.
The assessment of DNA fragmentation rate in semen is performed by direct (TUNEL, Comet) or indirect (SCSA, SCD) tests which, while they may yield different results, correlate well with each other in terms of indications of the presence of lesions (Esteves et al.,2021). Although a single reference threshold for the percentage of DNA fragmentation in semen has not yet been established, a major study by Esteves and colleagues (2021) estimates that values of 20-30% appear to be associated with adverse reproductive outcomes, either through natural conception or assisted reproduction, while a recent review by Agarwal et al. (2022) and an earlier meta-analysis by Santi et al. (2018) suggests a threshold of 20% to distinguish between fertile and infertile men.
Ways to reduce the DNA fragmentation rate in sperm
There are different strategies to reduce DNA fragmentation in sperm and these vary depending on the underlying factor affecting the genetic quality of sperm. High risk factors such as smoking, alcohol consumption, exposure to chemical factors and other damaging elements should be limited, as should factors that lead to an increase in scrotal temperature. In particular, cessation of harmful exposures and weight loss has been studied and found to be decisive in reducing the average rate of DNA fragmentation in sperm. Correction of varicocele has provided significant evidence for potentially improved reproductive outcomes with a recent meta-analysis by Qui et al. (2021) reporting a significant increase in pregnancy and birth rates following varicocele correction and assisted reproduction with ICSI.
The administration of antioxidants has been shown to be beneficial in terms of balancing the redox potential in sperm and in improving the genetic integrity of sperm DNA, but should be accompanied by medical and laboratory monitoring as overexposure to antioxidants due to excessive or unjustified use can cause reductive stress which equally negatively affects the quality of DNA in spermatozoa. Reductive stress prevents the normal maturation of spermatozoa by preventing sufficient condensation of genetic material in the nucleus and making it more susceptible to damage.
In addition, a reduction in ejaculatory abstinence time has been associated with lower levels of DNA fragmentation and in selected reproductive assistance protocols an improvement in clinical outcomes has been found with the collection and use of a second ejaculate sample at 1-3 hours intervals. Finally, in cases where medically assisted reproduction is indicated although with persisting DNA fragmentation levels, sperm selection is often offered through advanced pre-fertilization screening or through specific morphological criteria (absence of cenotopes) or functional criteria (ability to bind to hyaluronic acid) during ICSI, methods which, although they do not substitute holistic sample optimization, provide improved clinical results with a higher probability of selecting spermatozoa with intact DNA.
- Santi D, Spaggiari G, Simoni M. Sperm DNA fragmentation index as a promising predictive tool for male infertility diagnosis and treatment management – meta-analyses. Reprod Biomed Online 2018;37:315-26.
- Aitken RJ. DNA damage in human spermatozoa; important contributor to mutagenesis in the offspring. TranslAndrolUrol 2017;6(Suppl 4):S761-4).
- Esteves SC, Zini A, Coward RM, Evenson DP, Gosálvez J, LewisSEM, etal. Sperm DNA fragmentation testing: summary evidence and clinical practice recommendations. Andrologia 2021;53:e13874.
- Agarwal A, Farkouh A, Parekh N, Zini A, Arafa M, Kandil H, Tadros N, Busetto GM, Ambar R, Parekattil S, Boitrelle F, Sallam H, Jindal S, Ko E, Simopoulou M, Park HJ, Sadighi MA, Saleh R, Ramsay J, Martinez M, Elbardisi H, Alvarez J, Colpi G, Gosalvez J, Evenson D, Shah R. Sperm DNA Fragmentation: A Critical Assessment of Clinical Practice Guidelines. World J Mens Health. 2022 Jan;40(1):30-37.
- Qiu D, Shi Q, Pan L. Efficacy of varicocelectomy for sperm DNA integrity improvement: A meta-analysis. Andrologia 2021;53:e13885.
Farkouh A, Salvio G, Kuroda S, Saleh R, Vogiatzi P, Agarwal A. Sperm DNA integrity and male infertility: a narrative review and guide for the reproductive physicians. Transl Androl Urol. 2022 Jul;11(7):1023-1044. doi: 10.21037/tau-22-149.