Effect Of Sperm DNA Damage Research Articles

Sperm DNA Fragmentation: Causes, Effects and Relevance in IVF Outcomes

Sperm DNA fragmentation, characterized by damaged or broken DNA strands within sperm cells, has been recognized as an important factor in male infertility and poor ART outcome. This lecture aims to provide an overview of the causes, effects, and relevance of sperm DNA fragmentation in in vitro fertilization (IVF) outcomes and look into the latest evidences from the literatures. Causes: Sperm DNA fragmentation can result from various factors, including oxidative stress, advanced paternal age, lifestyle factors such as smoking and excessive alcohol consumption, environmental exposures, genotoxic agents, and underlying genetic abnormalities. These factors can induce DNA damage during sperm development, during the transit of sperm through the male reproductive tract or during the storage of the sperm in epididymis. Relevance in IVF Outcomes: The assessment of sperm DNA fragmentation has emerged as a valuable tool in evaluating male infertility and predicting IVF success. Several methods, including the sperm chromatin structure assay (SCSA), terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL), comet assay, and other commercial detection kits, are available to measure sperm DNA fragmentation. Previous small cohort studies have demonstrated that higher levels of sperm DNA fragmentation are associated with lower pregnancy rates, decreased embryo quality, and an increased risk of pregnancy loss in IVF cycles. Therefore, incorporating sperm DNA fragmentation analysis into the routine assessment of male infertility can aid in optimizing treatment strategies and improving IVF outcomes. Recent evidences: Latest large clinical studies showed a limited association to IVF outcome. The DFI remains controversial as a predictive tool. Nonetheless, sperm DNA fragmentation is still believed to be a significant factor affecting male fertility and IVF outcomes. Further research is warranted to explore the mechanisms underlying DNA fragmentation, develop standardized assessment methods, and explore potential interventions to mitigate the adverse effects of sperm DNA damage on IVF success.

The effect of paternal and maternal factors on the prognosis of live birth in couples with recurrent pregnancy loss.

Currently, recurrent pregnancy loss (RPL) examinations focus on the woman, although paternal factors are also involved. Men in couples with RPL have higher sperm DNA fragmentation levels than fertile men, but the effect of sperm DNA damage on couple's later prognosis is unknown. Advanced maternal age and obesity are associated with RPL, but paternal lifestyle factors are less studied. Therefore, we aimed to study the associations of couples' lifestyle factors, causes of RPL, and sperm DNA fragmentation with their prognosis of future live birth. This descriptive cohort study comprised 506 couples investigated for RPL at Helsinki University Hospital, Finland, between 2007 and 2016, linked with national health and population registers. The primary outcome was couple's live birth after RPL investigations. Data on couple's background factors, including age, body mass index, smoking, and alcohol use, were collected from medical records. Sperm DNA fragmentation index was analyzed from 211 men using the sperm chromatin dispersion test. The associations between background factors, sperm DNA fragmentation, and cumulative probability of live birth over time were analyzed using cross-tabulations and age-adjusted Cox regression. In all, 352 of 506 couples (69.6%) achieved live birth. Maternal age, unexplained RPL, prolonged pregnancy attempts before investigations, paternal obesity, and maternal smoking were associated with prognosis: unadjusted hazard ratio for couple's live birth for women aged 35-39 vs younger than 30 years was 0.63 (95% confidence interval [CI] 0.47-0.84), and for 40 years or older was 0.36 (95% CI 0.22-0.58). Age-adjusted hazard ratio for unexplained vs explained RPL was 1.39 (95% CI 1.12-1.72), for couple's pregnancy attempt at least 4 years vs less than 2 years was 0.50 (95% CI 0.33-0.76), for paternal body mass index at least 30 kg/m2 vs less than 25 kg/m2 was 0.67 (95% CI 0.46-0.98), and for maternal smoking was 0.71 (95% CI 0.51-0.99). Altogether, 96/135 (71.1%) couples with normal (<15%), 38/60 (63.3%) with intermediate (15-30%), and 11/16 (68.8%) with high sperm DNA fragmentation index achieved live birth (p =0.56). In couples with RPL, prolonged pregnancy attempts, a cause found in RPL examinations, lifestyle factors, and maternal age are negatively associated with their prognosis of future live birth. Sperm DNA fragmentation was not associated, but the number of men with damaged spermatozoa was small. We suggest that clinicians include women and men in RPL counseling because couple's joint lifestyle seems to determine their later prognosis.

Clinical implications of sperm DNA damage in IVF and ICSI: updated systematic review and meta-analysis.

The clinical effect of sperm DNA damage in assisted reproduction has been a controversial topic during recent decades, leading to a variety of clinical practice recommendations. While the latest European Society of Human Reproduction and Embryology (ESHRE) position report concluded that DNA damage negatively affects assisted reproduction outcomes, the Practice Committee of the American Society for Reproductive Medicine (ASRM) does not recommend the routine testing of DNA damage for in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). Herein, our aim was to perform a systematic review and meta-analysis of studies investigating whether sperm DNA damage affects clinical outcomes in IVF and ICSI, in order to contribute objectively to a consistent clinical recommendation. A comprehensive systematic search was conducted according to PRISMA guidelines from the earliest available online indexing year until March 2020, using the MEDLINE-PubMed and EMBASE databases. We included studies analysing IVF and/or ICSI treatments performed in infertile couples in which sperm DNA damage was well defined and assessed. Studies also had to include information about pregnancy, implantation or live birth rates as primary outcomes. The NHLBI-NIH quality assessment tool was used to assess the quality of each study. Meta-analyses were conducted using the Mantel-Haenszel method with random-effects models to evaluate the Risk Ratio (RR) between high-DNA-damage and control groups, taking into account the 95% confidence intervals. Heterogeneity among studies was evaluated using the I2 statistic. We also conducted sensitivity analyses and post-hoc subgroup analyses according to different DNA fragmentation assessment techniques. We identified 78 articles that met our inclusion and quality criteria and were included in the qualitative analysis, representing a total of 25639 IVF/ICSI cycles. Of these, 32 articles had sufficient data to be included in the meta-analysis, comprising 12380 IVF/ICSI cycles. Meta-analysis revealed that, considering IVF and ICSI results together, implantation rate (RR=0.74; 95% CI=0.61-0.91; I2 = 69) and pregnancy rate (RR=0.83; 0.73-0.94; I2 = 58) are negatively influenced by sperm DNA damage, although after adjustment for publication bias the relationship for pregnancy rate was no longer significant. The results showed a non-significant but detrimental tendency (RR=0.78; 0.58-1.06; I2 = 72) on live birth rate. Meta-analysis also showed that IVF outcomes are negatively influenced by sperm DNA damage, with a statistically significant impact on implantation (RR=0.68; 0.52-0.89; I2 = 50) and pregnancy rates (RR=0.72; 0.55-0.95; I2 = 72), although the latter was no longer significant after correction for publication bias. While it did not quite meet our threshold for significance, a negative trend was also observed for live birth rate (RR=0.48; 0.22-1.02; I2 = 79). In the case of ICSI, non-significant trends were observed for implantation (RR=0.79; 0.60-1.04; I2 = 72) or pregnancy rates (RR=0.89; 0.78-1.02; I2 = 44), and live birth rate (RR=0.92; 0.67-1.27; I2 = 70). The current review provides the largest evidence to date supporting a negative association between sperm DNA damage and conventional IVF treatments, significantly reducing implantation and pregnancy rates. The routine use of sperm DNA testing is therefore justified, since it may help improve the outcomes of IVF treatments and/or allow a given couple to be advised on the most suitable treatment. Further well-designed controlled studies on a larger number of patients are required to allow us to reach more precise conclusions, especially in the case of ICSI treatments.

The effect of sperm DNA damage on the metabolomic of pre-implantation mouse embryos

The effect of sperm DNA damage on the metabolomic of pre-implantation mouse embryos

The effect of sperm DNA damage on the quantitative secretome of murine blastocysts

The effect of sperm DNA damage on the quantitative secretome of murine blastocysts

A systematic review and meta-analysis to determine the effect of sperm DNA damage on in vitro fertilization and intracytoplasmic sperm injection outcome.

Sperm DNA damage is prevalent among infertile men and is known to influence natural reproduction. However, the impact of sperm DNA damage on assisted reproduction outcomes remains controversial. Here, we conducted a meta-analysis of studies on sperm DNA damage (assessed by SCSA, TUNEL, SCD, or Comet assay) and clinical pregnancy after IVF and/or ICSI treatment from MEDLINE, EMBASE, and PUBMED database searches for this analysis. We identified 41 articles (with a total of 56 studies) including 16 IVF studies, 24 ICSI studies, and 16 mixed (IVF + ICSI) studies. These studies measured DNA damage (by one of four assays: 23 SCSA, 18 TUNEL, 8 SCD, and 7 Comet) and included a total of 8068 treatment cycles (3734 IVF, 2282 ICSI, and 2052 mixed IVF + ICSI). The combined OR of 1.68 (95% CI: 1.49–1.89; P < 0.0001) indicates that sperm DNA damage affects clinical pregnancy following IVF and/or ICSI treatment. In addition, the combined OR estimates of IVF (16 estimates, OR = 1.65; 95% CI: 1.34–2.04; P < 0.0001), ICSI (24 estimates, OR = 1.31; 95% CI: 1.08–1.59; P = 0.0068), and mixed IVF + ICSI studies (16 estimates, OR = 2.37; 95% CI: 1.89–2.97; P < 0.0001) were also statistically significant. There is sufficient evidence in the existing literature suggesting that sperm DNA damage has a negative effect on clinical pregnancy following IVF and/or ICSI treatment.

Should we evaluate and treat sperm DNA fragmentation?

The clinical utility of sperm DNA fragmentation tests needs to be revisited in light of increasing evidence of detrimental effect of sperm DNA damage on reproductive outcomes. Current evidence supports the association between high sperm DNA fragmentation and poor outcomes with regards to natural conception and intrauterine insemination. The relationship between high sperm DNA fragmentation and impaired outcomes after in-vitro fertilization and intracytoplasmic sperm injection are more equivocal. However, recent studies indicate that poor sperm chromatin content is associated with an increased risk of early pregnancy loss after in-vitro fertilization and intracytoplasmic sperm injection. Several strategies are proposed to alleviate sperm DNA fragmentation and/or select sperm with higher quality chromatin content for assisted reproductive techniques. The intake of oral antioxidants, varicocele repair, use of recurrent ejaculations alone or combined with micromanipulation-based sperm selection techniques, and the use of testicular sperm for intracytoplasmic sperm injection have been attempted with promising results. Sperm DNA fragmentation tests provide clinically relevant information for natural conception and artificial reproduction independent of those derived from conventional semen parameters. The increasing knowledge of paternal factors on pregnancy outcome and the improvement in treatment strategies should prompt routine evaluation of sperm DNA fragmentation in infertile couples.

A novel antioxidant formulation designed to treat male infertility associated with oxidative stress: promising preclinical evidence from animal models.

Does a novel antioxidant formulation designed to restore redox balance within the male reproductive tract, reduce sperm DNA damage and increase pregnancy rates in mouse models of sperm oxidative stress? Oral administration of a novel antioxidant formulation significantly reduced sperm DNA damage in glutathione peroxidase 5 (GPX5), knockout mice and restored pregnancy rates to near-normal levels in mice subjected to scrotal heat stress. Animal and human studies have documented the adverse effect of sperm DNA damage on fertilization rates, embryo quality, miscarriage rates and the transfer of de novo mutations to offspring. Semen samples of infertile men are known to be deficient in several key antioxidants relative to their fertile counterparts. Antioxidants alone or in combination have demonstrated limited efficacy against sperm oxidative stress and DNA damage in numerous human clinical trials, however these studies have not been definitive and an optimum combination has remained elusive. The efficacy of the antioxidant formulation was evaluated in two well-established mouse models of oxidative stress, scrotal heating and Gpx5 knockout (KO) mice, (n = 12 per experimental group), by two independent laboratories. Mice were provided the antioxidant product in their drinking water for 2-8 weeks and compared with control groups for sperm DNA damage and pregnancy rates. In the Gpx5 KO model, oxidative DNA damage was monitored in spermatozoa by immunocytochemical detection of 8-hydroxy-2'-deoxyguanosine (8OHdG). In the scrotal heat stress model, male fertility was tested by partnering with three females for 5 days. The percentage of pregnant females, number of vaginal plugs, resorptions per litter, and litter size were recorded. Using immunocytochemical detection of 8OHdG as a biomarker of DNA oxidation, analysis of control mice revealed that around 30% of the sperm population was positively stained. This level increased to about 60% in transgenic mice deficient in the antioxidant enzyme, GPX5. Our results indicate that an 8 week pretreatment of Gpx5 KO mice with the antioxidant formulation provided complete protection of sperm DNA against oxidative damage. In mouse models of scrotal heat stress, only 35% (19/54) of female mice became pregnant resulting in 169 fetuses with 18% fetal resorption (30/169). This is in contrast to the antioxidant pretreated group where 74% (42/57) of female mice became pregnant, resulting in 427 fetuses with 9% fetal resorption (38/427). In both animal models the protection provided by the novel antioxidant was statistically significant (P < 0.01 for the reduction of 8OHdG in the spermatozoa of Gpx5 KO mice and P < 0.05 for increase in fertility in the scrotal heat stress model). It was not possible to determine the exact level of antioxidant consumption for each mouse during the treatment period. Recent clinical studies confirm moderate to severe sperm DNA damage in about 60% of all men visiting IVF centers and in about 80% of men diagnosed with idiopathic male infertility. Our results, if confirmed in humans, will impact clinical fertility practice because they support the concept of using an efficacious antioxidant supplementation as a preconception therapy, in order to optimize fertilization rates, help to maintain a healthy pregnancy and limit the mutational load carried by children. The study was funded by the Clermont Université and the University of Madrid. P.G. is the Managing Director of CellOxess LLC, which has a commercial interest in the detection and resolution of oxidative stress. A.M. and A.P. are employees of CellOxess, LLC. J.R.D., A.G.-A. and R.J.A. are honorary members of the CellOxess advisory board.

Paternal influence of sperm DNA integrity on early embryonic development

Does sperm DNA damage affect early embryonic development? Increased sperm DNA damage adversely affects embryo quality starting at Day 2 of early embryonic development and continuing after embryo transfer, resulting in reduced implantation rates and pregnancy outcomes. Abnormalities in the sperm DNA in the form of single and double strand breaks can be assessed by an alkaline Comet assay. Some prior studies have shown a strong paternal effect of sperm DNA damage on IVF outcome, including reduced fertilization, reduced embryo quality and cleavage rates, reduced numbers of embryos developing into blastocysts, increased percentage of embryos undergoing developmental arrest, and reduced implantation and pregnancy rates. A cross-sectional study of 215 men from infertile couples undergoing assisted reproduction techniques at the University of Utah Center for Reproductive Medicine. Sperm from men undergoing ART were analyzed for DNA damage using an alkaline Comet assay and classified into three groups: 'low damage' (0-30%), 'intermediate damage' (31-70%) and 'high damage' (71-100%). The cause of couples' infertility was categorized into one of the three types (male, female or unexplained). Each embryo was categorized as 'good', 'fair' or 'poor' quality, based on the number and grade of blastomeres. The influence of sperm DNA damage on early embryonic development was observed and classified into four stages: peri-fertilization effect (fertilization rate), early paternal effect (embryonic days 1-2), late paternal effect (embryonic days 3-5) and implantation stage effect. The paternal effect of sperm DNA damage was observed at each stage of early embryonic development. The peri-fertilization effect was higher in oocytes from patients with female infertility (20.85%) compared with male (8.22%; P < 0.001) and unexplained (7.30%; P < 0.001) infertility factors. In both the early and late paternal effect stages, the low DNA damage group had a higher percentage of good quality embryos (P < 0.05) and lower percentage of poor quality embryos (P < 0.05) compared with the high DNA damage group. Implantation was lower in the high DNA damage (33.33%) compared with intermediate DNA damage (55.26%; P < 0.001) and low DNA damage (65.00%; P < 0.001) groups. The implantation rate was higher following blastocyst transfer (58.33%), when compared with early stage blastocyst (53.85%; P = 0.554) and cavitating morula transfers (34.40%; P < 0.001). Implantation was higher when the female partner age was ≤35 years when compared with >35 year age group (52.75 versus 35.44%; P = 0.008). A potential limitation of this study is that it is cross-sectional. Generally in such studies more than one variable could affect the outcome. Analyzing sperm is one part of the equation but a number of environmental and female factors also have the potential to influence embryo development and implantation. Furthermore, the selection of morphologically normal and physiologically motile sperm may result in isolation of sperm with reduced DNA damage. Therefore, selecting the best available sperm for ICSI may lead to experimental bias, as the selected sperm do not represent the overall sperm population in which the DNA damage is measured. Similar studies on selected sperm and with a larger sample size are now required. The paternal influence of damaged chromatin is more prominent after zygotic transcriptional activation. A prolonged paternal effect on the developing embryo may be due to the active repair mechanism present in oocytes that tends to overcome the damaged paternal chromatin. The probability of eliminating an embryo fertilized by a sperm with damaged DNA is higher at the blastocyst stage than the cleavage stage; therefore blastocyst transfer could be recommended for better implantation success. Finally, we recommend ICSI treatment for patients with a higher percentage of sperm with DNA damage as well as additional studies with a larger sample size aimed at assessing DNA damage analysis as a diagnostic tool for IVF. This work was supported by the University of Utah internal funds. The authors declare no competing interests. N/A.

Phospho-Cdc25 correlates with activating G2/M checkpoint in mouse zygotes fertilized with hydrogen peroxide-treated mouse sperm.

The presence of oxidative stress in sperm cryopreservation induces sperm DNA damage. Our previous study has discovered that γH2AX, the DNA-damaged marker, was activated in the early mouse embryos fertilized with hydrogen peroxide (H2O2)-treated sperm. Furthermore, we found that checkpoint proteins ATM and Chk1 were phosphorylated and activated in the early mouse embryos. On the basis of previous researches, we examined the effects of sperm DNA damage on cell cycle arrest in mouse zygotes fertilized with H2O2-treated sperm. Development of fertilized eggs arrested at the PN disappearance stage. At 19 and 24 hours post-insemination (hpi), the percentage of zygotes at the PN disappearance stage was higher in H2O2-treated group compared to the control group. Immunofluorescence staining revealed Phospho-Cdc25C (Ser216) and Phospho-Cdc25B (Ser323) in or surrounding a single pronucleus, following insemination with H2O2-treated sperm. Our study suggests that fertilization with DNA-damaged sperm results in cell cycle arrest mediated by G2/M checkpoint activation in one of the pronuclei in mouse zygotes fertilized with H2O2-treated sperm; Phospho-Cdc25C and Phospho-Cdc25B correlate with activating G2/M checkpoint in zygotes fertilized with H2O2-treated sperm.

Endocannabinoids as Markers of Sperm Quality: Hot Spots

Endocannabinoids as Markers of Sperm Quality: Hot Spots

The effect of sperm DNA fragmentation on miscarriage rates: a systematic review and meta-analysis

Is there an association between high levels of sperm DNA damage and miscarriage? Miscarriage rates are positively correlated with sperm DNA damage levels. Most ejaculates contain a subpopulation of sperm with DNA damage, also referred to as DNA fragmentation, in the form of double or single-strand breaks which have been induced in the DNA prior to or following ejaculation. This DNA damage may be particularly elevated in some subfertile men, hence several studies have examined the link between sperm DNA damage levels and conception and miscarriage rates. A systematic review and meta-analysis of studies which examined the effect of sperm DNA damage on miscarriage rates was performed. Searches were conducted on MEDLINE, EMBASE and the Cochrane Library without any language restrictions from database inception to January 2012. We used the terms 'DNA damage' or 'DNA fragmentation' combined with 'miscarriage', 'abortion' or 'pregnancy' to generate a set of relevant citations. Data extraction was performed by two reviewers. Study quality was assessed using the Newcastle-Ottawa Scale. Meta-analysis of relative risks of miscarriage was performed with a random effects model. Subgroup analyses were performed by the type of DNA damage test, whether the sperm examined were prepared or from raw semen and for pregnancies resulting from IVF or ICSI treatment. We identified 16 cohort studies (2969 couples), 14 of which were prospective. Eight studies used acridine orange-based assays, six the TUNEL assay and two the COMET assay. Meta-analysis showed a significant increase in miscarriage in patients with high DNA damage compared with those with low DNA damage [risk ratio (RR) = 2.16 (1.54, 3.03), P < 0.00001)]. A subgroup analysis showed that the miscarriage association is strongest for the TUNEL assay (RR = 3.94 (2.45, 6.32), P < 0.00001). There is some variation in study characteristics, including the use of different assays and different thresholds for DNA damage and the definition of pregnancy loss. The use of methods which select sperm without DNA damage for use in assisted conception treatment may reduce the risk of miscarriage. This finding indicates that assays detecting DNA damage could be considered in those suffering from recurrent pregnancy loss. Further research is necessary to study the mechanisms of DNA damage and the potential therapeutic effects of antioxidant therapy. None.

Effects of sperm DNA damage on the levels of RAD51 and p53 proteins in zygotes and 2-cell embryos sired by golden hamsters without the major accessory sex glands

We previously reported that the male accessory sex gland (ASG) secretion is the main source of antioxidants to safeguard sperm genomic integrity and functional competence. Removal of all ASGs in the golden hamster can reduce male fertility by increasing embryo wastage. This study aims to investigate whether the oxidative DNA-damaged sperm from hamsters without all ASGs (TX) could successfully fertilize oocytes and to qualify the status of DNA repair by the expression of RAD51 and p53 proteins. Here we demonstrated a significantly higher DNA-base adduct formation (8-hydroxy-2′-deoxyguanosine) in sperm from TX males than those from sham-operated males. Comet assays demonstrated that all female pronuclei in both zygotes were intact, but single- and double-strand DNA damage was found in decondensed sperm in TX males only. DNA damage could also be detected in both nuclei of the TX 2-cell embryos. RAD51, a DNA repair enzyme, was found to be evenly distributed in the cytoplasm and nuclei in oocytes/zygotes, while at the 2-cell stage, a strong expression of p53 protein and a larger clear perinuclear area without RAD51 expression were found in TX embryos. In conclusion, we demonstrated for the first time DNA damage in decondensed sperm of zygotes and blastomeres of 2-cell stage embryos sired by TX males, resulting in the activation of DNA repair. Sperm DNA damage could induce the increase in p53 expression and the reduction of RAD51 expression in the TX 2-cell stage embryos.

Cannabinoid Receptor 1 Influences Chromatin Remodeling in Mouse Spermatids by Affecting Content of Transition Protein 2 mRNA and Histone Displacement

Marijuana smokers and animals treated with Δ9-tetrahydrocannabinol, the principal component of marijuana, show alterations of sperm morphology suggesting a role for cannabinoids in sperm differentiation and/or maturation. Because the cannabinoid receptor 1 (CNR1) activation appears to play a pivotal role in spermiogenesis, the developmental stage where DNA is remodeled, we hypothesized that CNR1 receptors might also influence chromatin quality in sperm. We used Cnr1 null mutant (Cnr1-/-) mice to study the possible role of endocannabinoids on sperm chromatin during spermiogenesis. We demonstrated that CNR1 activation regulated chromatin remodeling of spermatids by either increasing Tnp2 levels or enhancing histone displacement. Comparative analysis of wild-type, Cnr1+/-, and Cnr1-/- animals suggested the possible occurrence of haploinsufficiency for Tnp2 turnover control by CNR1, whereas histone displacement was disrupted to a lesser extent. Furthermore, flow cytometry analysis demonstrated that the genetic loss of Cnr1 decreased sperm chromatin quality and was associated with sperm DNA fragmentation. This damage increased during epididymal transit, from caput to cauda. Collectively, our results show that the expression/activity of CNR1 controls the physiological alterations of DNA packaging during spermiogenesis and epididymal transit. Given the deleterious effects of sperm DNA damage on male fertility, we suggest that the reproductive function of marijuana users may also be impaired by deregulation of the endogenous endocannabinoid system.

Paternal levels of DNA damage in spermatozoa and maternal parity influence offspring mortality in an endangered ungulate.

Understanding which factors influence offspring mortality rates is a major challenge since it influences population dynamics and may constrain the chances of recovery among endangered species. Most studies have focused on the effects of maternal and environmental factors, but little is known about paternal factors. Among most polygynous mammals, males only contribute the haploid genome to their offspring, but the possibility that sperm DNA integrity may influence offspring survival has not been explored. We examined several maternal, paternal and individual factors that may influence offspring survival in an endangered species (Gazella cuvieri). Levels of sperm DNA damage had the largest impact upon offspring mortality rates, followed by maternal parity. In addition, there was a significant interaction between these two variables, so that offspring born to primiparous mothers were more likely to die if their father had high levels of sperm DNA damage, but this was not the case among multiparous mothers. Thus, multiparous mothers seem to protect their offspring from the deleterious effects of sperm DNA damage. Since levels of sperm DNA damage seem to be higher among endangered species, more attention should be paid to the impact of this largely ignored factor among the viability of endangered species.

Fertilization capacity with rainbow trout DNA-damaged sperm and embryo developmental success

Mammalian spermatozoa undergo a strong selection process along the female tract to guarantee fertilization by good quality cells, but risks of fertilization with DNA-damaged spermatozoa have been reported. In contrast, most external fertilizers such as fish seem to have weaker selection procedures. This fact, together with their high prolificacy and external embryo development, indicates that fish could be useful for the study of the effects of sperm DNA damage on embryo development. We cryopreserved sperm from rainbow trout using egg yolk and low-density lipoprotein as additives to promote different rates of DNA damage. DNA fragmentation and oxidization were analyzed using comet assay with and without digestion with restriction enzymes, and fertilization trials were performed. Some embryo batches were treated with 3-aminobenzamide (3AB) to inhibit DNA repair by the poly (ADP-ribose) polymerase, which is an enzyme of the base excision repair pathway. Results showed that all the spermatozoa cryopreserved with egg yolk carried more than 10% fragmented DNA, maintaining fertilization rates of 61.1+/-2.3 but a high rate of abortions, especially during gastrulation, and only 14.5+/-4.4 hatching success. Furthermore, after 3AB treatment, hatching dropped to 3.2+/-2.2, showing that at least 10% DNA fragmentation was repaired. We conclude that trout sperm maintains its ability to fertilize in spite of having DNA damage, but that embryo survival is affected. Damage is partially repaired by the oocyte during the first cleavage. Important advantages of using rainbow trout for the study of processes related to DNA damage and repair during development have been reported.

Sperm chromatin structure assay parameters are not related to fertilization rates, embryo quality, and pregnancy rates in in vitro fertilization and intracytoplasmic sperm injection, but might be related to spontaneous abortion rates

To investigate the relationship between sperm chromatin structure assay (SCSA) parameters, DNA fragmentation index (DFI) and high DNA stainability (HDS), and outcomes of IVF and intracytoplasmic sperm injection (ICSI). Retrospective review and prospective study. Academic human reproduction laboratory. Two hundred twenty-three couples undergoing conventional IVF (n = 137) and ICSI (n = 86). Testing with SCSA on a semen aliquot taken from ejaculate used for assisted reproductive technology (ART). Conventional semen parameters, DFI, HDS, outcomes of IVF and ICSI. There were no significant differences in IVF and ICSI fertilization rate, good embryo rate, and pregnancy rate (PR) between high, moderate, and low DFI or HDS groups. Men with HDS >15% had significantly higher IVF abortion rates. There was a statistically insignificant trend toward an increased abortion rate in the high DFI (>27%) group. The DFI correlated negatively with sperm motility, and HDS correlated negatively with sperm morphology and concentration. Neither DFI nor HDS scores can provide independent information about embryo quality, fertilization, and PRs for infertility patients undergoing ART. Sperm DNA fragmentation probably affects sperm motility. The relationship between HDS and IVF abortion rates provides preliminary evidence that ICSI may be indicated for men with HDS >15%. The potential adverse effect of sperm DNA damage on the quality of postimplantation embryo and spontaneous abortion should be a concern.

Correlation of sperm DNA damage with IVF and ICSI outcomes: A systematic review and meta-analysis

To assess the effects of sperm DNA damage, as determined by the TUNEL assay and the SCSA respectively, on the outcomes of IVF/ICSI treatment. A Medline search (from Jan 1978 to Apr 2006) was performed, together with a manual search of the bibliographies of retrieved original papers and review articles. 8 articles met all inclusion/exclusion criteria, of which, 5 used the TUNEL assay and the other 3 used the SCSA. All these articles were included in separate meta-analysis. The meta-analysis was conducted using the RevMan software with fixed-effect model or random-effects model. As for articles using the TUNEL assay, the pooled results of IVF outcomes indicated that the clinical pregnancy rate (RR 0.68, 95% CI 0.54 to 0.85, P = 0.006), but not the fertilization rate (RR 0.79, 95% CI 0.54 to 1.16, P = 0.23) decreased significantly for patients with high degree of sperm DNA damage compared with those with low degree of sperm DNA damage. RRs of the ICSI outcomes indicated that there was no significant difference in either fertilization rate (RR 1.03, 95% CI 0.89 to1.18, P = 0.70) or clinical pregnancy rate (RR 0.76, 95% CI 0.55 to 1.04, P = 0.09) between these two groups. As for the SCSA papers, the pooled results showed no significant effects of sperm DNA damage on the clinical pregnancy rate after IVF (RR 0.58, 95% CI 0.25 to 1.31, P = 0.19) or ICSI (RR 1.18, 95% CI 0.81 to 1.74, P = 0.38). Our meta-analysis indicates that sperm DNA damage, as assessed by the TUNEL assay, significantly decreases only the chance of IVF clinical pregnancy, but not that of either IVF fertilization or ICSI fertilization or ICSI clinical pregnancy. Besides, our results also reveal that sperm DNA damage, when assessed by the SCSA, has no significant effect on the chance of clinical pregnancy after IVF or ICSI treatment.

DNA Integrity Is Compromised in Protamine‐Deficient Human Sperm

The objective of this study was to examine the relationship between DNA integrity and protamines in human sperm. One hundred forty-nine male infertility patients were included in an Institutional Review Board-approved study. Sperm were evaluated for DNA fragmentation using the DNA Integrity Assay, a test equivalent to the sperm chromatin structure assay (SCSA). Additionally, nuclear proteins were extracted and the protamine-1/protamine-2 ratio (P1/P2), protamine-1 (P1), protamine-2 (P2), and total protamine concentrations were evaluated. We identified 37 patients with abnormally low P1/P2 ratios, 99 patients with normal P1/P2 ratios, and 13 patients with abnormally high P1/P2 ratios. DNA fragmentation was significantly elevated in patients with low P1/P2 ratios (37.1 +/- 6.02) vs those with normal and high P1/P2 ratios (26.7 +/- 1.9 and 23.8 +/- 3.2, respectively; P < .05) and was inversely correlated with the P1/P2 ratio (R(s) -0.18, P < .05), P1 concentration (R(s) -0.29, P < .001), P2 concentration (R(s) - 0.24, P < .005), and total protamine concentration (R(s) -0.28, P < .001). Furthermore, chi2 analysis revealed a significant increase in the incidence of marked DNA fragmentation in patients with diminished levels of either P1 or P2. The present study is the first to report that human sperm protamine content is significantly related to DNA fragmentation. In particular, sperm P1 and P2 concentrations inversely correlate with DNA fragmentation, indicating a protective role of the protamines against sperm DNA damage. In light of recent studies highlighting the negative effect of sperm DNA damage on ART outcomes, these findings indicate a possible clinical significance for human sperm protamine levels.

Potential adverse effect of sperm DNA damage on embryo quality after ICSI

Sperm DNA damage is prevalent amongst infertile men and has been shown to strongly impact adversely natural reproduction, intrauterine insemination-assisted reproduction and to a lesser degree IVF/ICSI fertilization. The objective of this study was to examine further the relationship between sperm DNA denaturation (DD) and reproductive outcomes after ICSI. We evaluated infertile couples (n = 60) undergoing IVF/ICSI at a single centre. Sperm DD was assessed by flow cytometry analysis of Acridine Orange-treated sperm and expressed as the percentage of sperm with DD. Couples were sub-grouped according to sperm DD results: group 1: 0-15%; group 2: >15-30%; group 3: >30%. There were no differences between the three groups with regard to maternal age, sperm parameters, oocyte maturation, fertilization or pregnancy rates. Group 3 had a significantly higher rate of multinucleation among the embryo cohorts compared to either groups 1 or 2 (20% versus 10% and 8% respectively, P = 0.04). There was a statistically insignificant trend toward an increased spontaneous pregnancy loss rate in group 3 (P =0.50). Although we did not observe significant relationships between sperm DNA damage and either fertilization or pregnancy rates, the potential adverse effect of sperm DNA damage on embryo quality and spontaneous pregnancy loss is concerning.