Differential resistance of mammalian sperm chromatin to oxidative stress as assessed by a two-tailed comet assay

2011 ◽  
Vol 23 (5) ◽  
pp. 633 ◽  
Author(s):  
María Enciso ◽  
Stephen D. Johnston ◽  
Jaime Gosálvez

Protamines of eutherian species are cysteine-rich molecules that become cross-linked by disulfide bonds during epididymal transit, whereas the protamines of most marsupial species lack cysteine residuals. The present study made use of the differences in protamine structure between eutherian and metatherian mammal spermatozoa to examine the comparative resistance of sperm DNA to oxidative damage in three eutherian species (Mus musculus, Homo sapiens, Sus domesticus) and three metatherian species (Vombatus ursinus, Phascolarctos cinereus, Macropus giganteus). Sperm DNA fragmentation of samples exposed to increasing concentrations of hydrogen peroxide was assessed by means of the two-tailed comet assay. The sperm DNA of the marsupial species studied were significantly more sensitive to oxidative stress than the spermatozoa of eutherian species. Such susceptibility is consistent with the lack of disulfide cross-linking in marsupial sperm chromatin and suggests that the oxidation of thiols to disulfides for chromatin condensation during epididymal transit in eutherian mammals is likely to be important in order to provide stability and protect these cells from the genotoxic effects of adverse environments.

2015 ◽  
Vol 27 (8) ◽  
pp. 1168 ◽  
Author(s):  
K. Pollock ◽  
J. Gosálvez ◽  
F. Arroyo ◽  
C. López-Fernández ◽  
M. Guille ◽  
...  

The integrity of sperm DNA is becoming increasingly recognised as an important parameter of semen quality, but there are no published reports of this procedure for any amphibian. The primary aim of this study was to apply a modified sperm chromatin dispersion (SCD) test (Halomax) to an amphibian sperm model (African clawed frog; Xenopus laevis) and to validate the assay against in situ nick translation (ISNT) and the double-comet assay procedure. Inactivated spermatozoa were collected from fresh testes (n = 3). Sperm DNA fragmentation (SDF) for each sperm sample was conducted immediately following activation (T0) and again after 1 h (T1) and 24 h (T24) of incubation at room temperature in order to produce a range of spermatozoa with differing levels of DNA damage. The SCD procedure resulted in the production of three nuclear morphotypes; amphibian sperm morphotype 1 (ASM-1) and ASM-2 showed no evidence of DNA damage, whereas ASM-3 spermatozoa were highly fragmented with large halos of dispersed DNA fragments and a reduced nuclear core. ISNT confirmed that ASM-3 nuclei contained damaged DNA. There was a significant correlation (r = 0.9613) between the levels of ASM-3 detected by the SCD test and SDF revealed by the double-comet assay.


Reproduction ◽  
2013 ◽  
Vol 146 (5) ◽  
pp. 433-441 ◽  
Author(s):  
Renata Simões ◽  
Weber Beringui Feitosa ◽  
Adriano Felipe Perez Siqueira ◽  
Marcilio Nichi ◽  
Fabíola Freitas Paula-Lopes ◽  
...  

Sperm chromatin fragmentation may be caused by a number of factors, the most significant of which is reactive oxygen species. However, little is known about the effect of sperm oxidative stress (OS) on DNA integrity, fertilization, and embryonic development in cattle. Therefore, the goal of this study was to evaluate the influence of sperm OS susceptibility on the DNA fragmentation rate and in vitro embryo production (IVP) in a population of bulls. Groups of cryopreserved sperm samples were divided into four groups, based on their susceptibility to OS (G1, low OS; G2, average OS; G3, high OS; and G4, highest OS). Our results demonstrated that the sperm DNA integrity was compromised in response to increased OS susceptibility. Furthermore, semen samples with lower susceptibility to OS were also less susceptible to DNA damage (G1, 4.06%; G2, 6.09%; G3, 6.19%; and G4, 6.20%). In addition, embryo IVP provided evidence that the embryo cleavage rate decreased as the OS increased (G1, 70.18%; G2, 62.24%; G3, 55.85%; and G4, 50.93%), but no significant difference in the blastocyst rate or the number of blastomeres was observed among the groups. The groups with greater sensitivity to OS were also associated with a greater percentage of apoptotic cells (G1, 2.6%; G2, 2.76%; G3, 5.59%; and G4, 4.49%). In conclusion, we demonstrated that an increased susceptibility to OS compromises sperm DNA integrity and consequently reduces embryo quality.


Andrology ◽  
2020 ◽  
Vol 8 (3) ◽  
pp. 610-617 ◽  
Author(s):  
Haitham Elbardisi ◽  
Renata Finelli ◽  
Ashok Agarwal ◽  
Ahmad Majzoub ◽  
Ralf Henkel ◽  
...  

Genes ◽  
2019 ◽  
Vol 10 (3) ◽  
pp. 236 ◽  
Author(s):  
Sheryl Homa ◽  
Anna Vassiliou ◽  
Jesse Stone ◽  
Aideen Killeen ◽  
Andrew Dawkins ◽  
...  

Oxidative stress (OS) is a significant cause of DNA fragmentation and is associated with poor embryo development and recurrent miscarriage. The aim of this study was to compare two different methods for assessing seminal OS and their ability to predict sperm DNA fragmentation and abnormal semen parameters. Semen samples were collected from 520 men attending for routine diagnostic testing following informed consent. Oxidative stress was assessed using either a chemiluminescence assay to measure reactive oxygen species (ROS) or an electrochemical assay to measure oxidation reduction potential (sORP). Sperm DNA fragmentation (DFI) and sperm with immature chromatin (HDS) were assessed using sperm chromatin structure assay (SCSA). Semen analysis was performed according to WHO 2010 guidelines. Reactive oxygen species sORP and DFI are negatively correlated with sperm motility (p = 0.0012, 0.0002, <0.0001 respectively) and vitality (p < 0.0001, 0.019, <0.0001 respectively). The correlation was stronger for sORP than ROS. Reactive oxygen species (p < 0.0001), sORP (p < 0.0001), DFI (p < 0.0089) and HDS (p < 0.0001) were significantly elevated in samples with abnormal semen parameters, compared to those with normal parameters. Samples with polymorphonuclear leukocytes (PMN) have excessive ROS levels compared to those without (p < 0.0001), but sORP and DFI in this group are not significantly increased. DNA fragmentation was significantly elevated in samples with OS measured by ROS (p = 0.0052) or sORP (p = 0.004). The results demonstrate the multi-dimensional nature of oxidative stress and that neither assay can be used alone in the diagnosis of OS, especially in cases of leukocytospermia.


Reproduction ◽  
2010 ◽  
Vol 140 (3) ◽  
pp. 445-452 ◽  
Author(s):  
Paola Villani ◽  
Patrizia Eleuteri ◽  
Maria Giuseppa Grollino ◽  
Michele Rescia ◽  
Pierluigi Altavista ◽  
...  

Sperm DNA damage may have adverse effects on reproductive outcome. Sperm DNA breaks can be detected by several tests, which evaluate DNA integrity from different and complementary perspectives and offer a new class of biomarkers of the male reproductive function and of its possible impairment after environmental exposure. The remodeling of sperm chromatin produces an extremely condensed nuclear structure protecting the nuclear genome from adverse environments. This nuclear remodeling is species specific, and differences in chromatin structure may lead to a dissimilar DNA susceptibility to mutagens among species. In this study, the capacity of the comet assay in its two variants (alkaline and neutral) to detect DNA/chromatin integrity has been evaluated in human, mouse, and bull sperm. The hypothesis that chromatin packaging might influence the amount of induced and detectable DNA damage was tested by treating sperm in vitro with DNAse I, whose activity is strictly dependent upon its DNA accessibility. Furthermore, hydrogen peroxide (H2O2) was used to assess whether spermatozoa of the three species showed a different sensitivity to oxidative stress. DNAse I-induced damage was also assessed by the sperm chromatin structure assay and the TUNEL assay, and the performances of these two assays were compared and correlated with the comet assay results. Results showed a different sensitivity to DNAse I treatment among the species with human sperm resulting the most susceptible. On the contrary, no major differences among species were observed after H2O2 treatment. Furthermore, the three tests show a good correlation in revealing sperm with DNA strand breaks.


2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Valeria Maria Iommiello ◽  
Elena Albani ◽  
Alessandra Di Rosa ◽  
Alessandra Marras ◽  
Francesca Menduni ◽  
...  

Oxidative stress (OS) plays an essential role in male infertility aetiology by affecting sperm quality, function, and also the integrity of sperm DNA. The assessment of oxidative stress in semen may be an important tool to improve the evaluation of sperm reproductive capacity. The purpose of this study was the evaluation of any possible relation between the unbalance of oxidative stress caused by superoxide anion in the ejaculate with the presence of sperm DNA fragmentation and high concentration of round cells. 56 semen samples from males from couples suffering from infertility were evaluated according to World Health Organisation (WHO) 2010 guidelines. Oxidative stress levels from N1 (low) to N4 (high) were assessed in ejaculates using oxiSperm; DFI (sperm DNA fragmentation index) as assessed by the SCSA (Sperm Chromatin Structure Assay) was used for evaluation of sperm chromatin integrity. Our data show that high oxidative stress (N3-N4 levels) correlated positively with aDFI≥30%(P=0.0379)and round cells≥1.500.000/mL(P=0.0084). In conclusion, OS increases sperm DNA damage. Thus evaluation of semen OS extent of sperm DNA damage in infertile man could be useful to develop new therapeutic strategies and improve success of assisted reproduction techniques (ART).


Zygote ◽  
2021 ◽  
pp. 1-8
Author(s):  
Shikai Wang ◽  
Weihong Tan ◽  
Yueyue Huang ◽  
Xianbao Mao ◽  
Zhengda Li ◽  
...  

Summary To determine the effects of sperm DNA fragmentation (SDF) on embryo morphokinetic parameters, cleavage patterns and embryo quality, this retrospective study analyzed 151 intracytoplasmic sperm injection (ICSI) cycles (1152 embryos collected) between November 2016 and June 2019. SDF was assessed using sperm chromatin dispersion. The cycles were divided into two groups based on the SDF rate: SDF < 15% (n = 114) and SDF ≥ 15% (n = 37). The embryo morphokinetic parameters, cleavage patterns, and embryo quality were compared between the two groups. The morphokinetic parameters tPNf, t2, t3, t4, t5, t6, and t8 were achieved significantly earlier in the SDF < 15% group compared with in the SDF ≥ 15% group. The fertilization and 2PN rates seemed to be significantly higher in the SDF < 15% group compared with in the SDF ≥ 15% group, while the abnormal cleavage rates were similar. However, a significantly higher rate of chaotic cleavage (CC) was observed in the SDF ≥ 15% group. The D3 high-quality embryo and available embryo rates were similar between the two groups. The blastocyst formation, high-quality blastocyst, and available blastocyst rates in the SDF < 15% group were significantly higher than those in the SDF ≥ 15% group. With an increase in SDF level, the chemical pregnancy, clinical pregnancy and implantation rates tended to decrease, while the miscarriage rate increased. This study demonstrated that SDF ≥ 15% reduces the fertilization rate of ICSI cycles and affects certain morphokinetic parameters. A higher SDF level can also induce a higher rate of CC, with subsequent decreases in the blastocyst formation rate and blastocyst quality.


2020 ◽  
Vol 98 (Supplement_4) ◽  
pp. 83-84
Author(s):  
Marina Fortes ◽  
Wei Liang Andre Tan ◽  
Laercio R Porto-Neto ◽  
Antonio Reverter ◽  
Gry B Boe-Hansen

Abstract Traits such as sperm morphology and motility are routine in veterinarian evaluations of bull fertility. However, they rarely are included in livestock breeding programs, which typically use only scrotal circumference (SC) and some female traits for fertility selection. We studied 25 male fertility traits measured in two research populations of bulls (1,099 Brahman, and 1,719 Tropical Composite) and one commercial population (2,490 Santa Gertrude bulls). Measurements included standard semen evaluation (e.g. sperm motility and morphology) and SC. In the research data, we also measured sperm DNA fragmentation and sperm protamine deficiency for about 50% of the bulls. Using a mixture of genomic and pedigree analyses, we estimated heritabilities and genetic correlations for all traits, in each population. Our analyses suggest that bull fertility traits have a heritable component, which makes selective breeding possible. The phenotype variation in sperm DNA fragmentation and sperm protamine deficiency traits also have a heritable component (h2 ~ 0.05–0.22). These first estimates for heritability of sperm chromatin phenotypes require further studies, with larger datasets, to corroborate present results. In all three populations, we observed genetic correlations across traits that were favorable, but not high. For example, the percentage of normal sperm (PNS) from the sperm morphology evaluation was positively correlated with SC. In the research data, sperm DNA fragmentation was negatively correlated with PNS (r2 ~ 0.23–0.33), meaning that bulls with a higher PNS had less DNA fragmentation, being therefore more fertile according to both indicators. Given the favorable and yet not high genetic correlations between traits, it is possible to envision that sperm chromatin phenotypes might form a panel, together with PNS and SC, for a comprehensive bull fertility index. Selection indices that include fertility traits are being implemented in the dairy industry and could be recommended for beef cattle, too. An index that benefits from the favorable genetic correlations between traits that describe different aspects of bull fertility is a sensible approach to selective breeding. The clinical use of complementary indicators for male fertility is largely accepted, when deciding on bull fitness for the mating season. We propose extending this rationale to create a multi-trait index that captures genetic merit for bull fertility. In addition, we performed genome-wide association analyses in the research data and identified eight QTLs in the X chromosome. Correlations and shared SNP associations support the hypothesis that these phenotypes have the same underlying cause: abnormal spermatogenesis. In conclusion, it is possible to improve bull fertility through selective breeding, by measuring complementary fertility traits. Genomic selection for bull fertility might be more accurate if the X chromosome mutations that underlie the discovered QTL are included in the analyses. Polymorphisms associated with fertility in the bull accumulate in the X chromosome, as they do in humans and mice, thus suggesting specialization of this chromosome.


Antioxidants ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 97
Author(s):  
Leila Rashki Ghaleno ◽  
AliReza Alizadeh ◽  
Joël R. Drevet ◽  
Abdolhossein Shahverdi ◽  
Mojtaba Rezazadeh Valojerdi

One important reason for male infertility is oxidative stress and its destructive effects on sperm structures and functions. The particular composition of the sperm membrane, rich in polyunsaturated fatty acids, and the easy access of sperm DNA to oxidative damage due to sperm cell specific cytologic and metabolic features (no cytoplasm left and cells unable to mount stress responses) make it the cell type in metazoans most susceptible to oxidative damage. In particular, oxidative damage to the spermatozoa genome is an important issue and a cause of male infertility, usually associated with single- or double-strand paternal DNA breaks. Various methods of detecting sperm DNA fragmentation have become important diagnostic tools in the prognosis of male infertility and such assays are available in research laboratories and andrology clinics. However, to date, there is not a clear consensus in the community as to their respective prognostic value. Nevertheless, it is important to understand that the effects of oxidative stress on the sperm genome go well beyond DNA fragmentation alone. Oxidation of paternal DNA bases, particularly guanine and adenosine residues, the most sensitive residues to oxidative alteration, is the starting point for DNA damage in spermatozoa but is also a danger for the integrity of the embryo genetic material independently of sperm DNA fragmentation. Due to the lack of a spermatozoa DNA repair system and, if the egg is unable to correct the sperm oxidized bases, the risk of de novo mutation transmission to the embryo exists. These will be carried on to every cell of the future individual and its progeny. Thus, in addition to affecting the viability of the pregnancy itself, oxidation of the DNA bases in sperm could be associated with the development of conditions in young and future adults. Despite these important issues, sperm DNA base oxidation has not attracted much interest among clinicians due to the lack of simple, reliable, rapid and consensual methods of assessing this type of damage to the paternal genome. In addition to these technical issues, another reason explaining why the measurement of sperm DNA oxidation is not included in male fertility is likely to be due to the lack of strong evidence for its role in pregnancy outcome. It is, however, becoming clear that the assessment of DNA base oxidation could improve the efficiency of assisted reproductive technologies and provide important information on embryonic developmental failures and pathologies encountered in the offspring. The objective of this work is to review relevant research that has been carried out in the field of sperm DNA base oxidation and its associated genetic and epigenetic consequences.


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