Impaired protamination and sperm DNA damage in a Nellore bull with high percentages of morphological sperm defects in comparison to normospermic bulls

The routine semen evaluation assessing sperm concentration, motility and morphology, does not identify subtle defects in sperm chromatin architecture. Bulls appear to have stable chromatin, with low levels of DNA fragmentation. However, the nature of fragmentation and its impact on fertility remain unclear and there are no detailed reports characterizing the DNA organization and damage in this species. The intensive genetic selection, the use of artificial insemination and in vitro embryo production associated to the cryopreservation process can contribute to the chromatin damage and highlights the importance of sperm DNA integrity for the success of these technologies. Frozen-thawed semen samples from three ejaculates from a Nellore bull showed high levels of morphological sperm abnormalities (55.8±5.1%), and were selected for complementary tests. Damage of acrosomal (76.9±8.9%) and plasma membranes (75.7±9.3%) as well as sperm DNA strand breaks (13.8±9.5%) and protamination deficiency (3.7±0.6%) were significantly higher compared to the values measured in the semen of five Nellore bulls with normospermia (24.3±3.3%; 24.5±6.1%; 0.6±0.5%; 0.4±0.6% for acrosome, plasma membrane, DNA breaks and protamine deficiency, respectively) (P<0.05). Motility and percentage of spermatozoa with low mitochondrial potential showed no differences between groups. This study shows how routine semen analyses (in this case morphology) may point to the length and complexity of sperm cell damage emphasizing the importance of sperm function testing.

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Effects of disulfide bond reducing agents on sperm chromatin structural integrity and developmental competence of in vitro matured oocytes after intracytoplasmic sperm injection in pigs

Reproduction ◽

10.1530/rep-08-0143 ◽

2009 ◽

Vol 137 (4) ◽

pp. 633-643 ◽

Cited By ~ 11

Author(s):

Wen-Min Cheng ◽

Lei An ◽

Zhong-Hong Wu ◽

Yu-Bo Zhu ◽

Jing-Hao Liu ◽

...

Keyword(s):

Disulfide Bond ◽

Intracytoplasmic Sperm Injection ◽

Chemical Activation ◽

Reducing Agents ◽

Developmental Competence ◽

Sperm Chromatin ◽

Dna Integrity ◽

Acridine Orange Staining ◽

Sperm Dna

We recently reported that electrical activation followed by secondary chemical activation greatly enhanced the developmental competence ofin vitromatured porcine oocytes fertilized by intracytoplasmic sperm injection (ICSI). We hypothesized that sperm treatment with disulfide bond reducing agents will enhance the development competence of porcine embryos produced by this ICSI procedure. We examined the effects of glutathione (GSH), dithiothreitol (DTT), GSH or DTT in combination with heparin on sperm DNA structure, paternal chromosomal integrity, pronuclear formation, and developmental competence ofin vitromatured porcine oocytes after ICSI. Acridine orange staining and flow cytometry based sperm chromatin structure assay were used to determine sperm DNA integrity by calculating the cells outside the main population (COMP αT). No differences were observed in COMP αT values among GSH-treated and control groups. COMP αT values in GSH-treated groups were significantly lower than that in DTT-treated groups. Following ICSI, GSH treatments did not significantly alter paternal chromosomal integrity. Paternal chromosomal integrity in sperm treated with DTT plus or minus heparin was also the lowest among all groups. GSH-treated sperm yielded the highest rates of normal fertilization and blastocyst formation, which were significantly higher than that of control and DTT-treated groups. The majority of blastocysts derived from control and GSH-treated spermatozoa were diploid, whereas blastocysts derived from DTT-treated spermatozoa were haploid. In conclusion, sperm treatment with GSH enhanced the developmental capacity of porcine embryos produced by our optimized ICSI procedure.

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Species-Specific Differences in Sperm Chromatin Decondensation Between Eutherian Mammals Underlie Distinct Lysis Requirements

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.669182 ◽

2021 ◽

Vol 9 ◽

Author(s):

Jordi Ribas-Maynou ◽

Estela Garcia-Bonavila ◽

Carlos O. Hidalgo ◽

Jaime Catalán ◽

Jordi Miró ◽

...

Keyword(s):

Dna Damage ◽

Dna Condensation ◽

Proteinase K ◽

Sperm Chromatin ◽

Dna Integrity ◽

Dna Breaks ◽

Chromatin Decondensation ◽

The Third ◽

Core Length ◽

Sperm Dna

Sperm present a highly particular DNA condensation that is acquired during their differentiation. Protamines are key elements for DNA condensation. However, whereas the presence of protamine 1 (P1) is conserved across mammalian species, that of protamine 2 (P2) has evolved differentially, existing only few species that use both protamines for sperm DNA condensation. In addition, altered P1/P2 ratios and alterations in the expression of P1 have previously been associated to infertility and DNA damage disorders. On the other hand, different methods evaluating DNA integrity, such as Sperm Chromatin Dispersion (SCD) and Comet tests, need a previous complete DNA decondensation to properly assess DNA breaks. Related with this, the present study aims to analyze the resilience of sperm DNA to decodensation in different eutherian mammals. Sperm samples from humans, horses, cattle, pigs and donkeys were used. Samples were embedded in low melting point agarose and treated with lysis solutions to induce DNA decondensation and formation of sperm haloes. The treatment consisted of three steps: (1) incubation in SDS + DTT for 30 min; (2) incubation in DTT + NaCl for 30 min; and (3) incubation in DTT + NaCl with or without proteinase K for a variable time of 0, 30, or 180 min. How incubation with the third lysis solution (with or without proteinase K) for 0, 30, and 180 min affected DNA decondensation was tested through analyzing core and halo diameters in 50 sperm per sample. Halo/core length ratio was used as an indicator of complete chromatin decondensation. While incubation time with the third lysis solution had no impact on halo/core length ratios in species having P1 and P2 (human, equine and donkey), DNA decondensation of pig and cattle sperm, which only present P1, significantly (P < 0.05) increased following incubation with the third lysis solution for 180 min. In addition, the inclusion of proteinase K was found to accelerate DNA decondensation. In conclusion, longer incubations in lysis solution including proteinase K lead to higher DNA decondensation in porcine and bovine sperm. This suggests that tests intended to analyze DNA damage, such as halo or Comet assays, require complete chromatin deprotamination to achieve high sensitivity in the detection of DNA breaks.

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Impacts administration of Rifampicin on sperm DNA integrity and Male Reproductive System parameters in rats

Research Journal of Pharmacy and Technology ◽

10.52711/0974-360x.2021.00851 ◽

2021 ◽

pp. 4897-4902

Author(s):

Furqan Mohammed Al-Asady ◽

Dalia Abdulzahra Al-Saray

Keyword(s):

Sperm Motility ◽

Sperm Concentration ◽

Sperm Chromatin ◽

Dna Integrity ◽

Sperm Function ◽

High Dose ◽

Adult Rats ◽

Male Adult ◽

Sperm Dna Integrity ◽

Sperm Dna

Objective: Evaluate the impacts of rifampicin on certain sperm function parameters and to determine whether rifampicin has an impact on chromatin quality or sperm DNA integrity. Materials and Methods: Forty two male adult rats were subjected to this study. The entire rats were subjected to random division into six groups; four rifampicin- treated groups and two control groups. Rifampicin- treated groups were treated with a dose of either (27mg/kg/day) or (54mg/kg/day) and for each treatment dose, the treatment persists for either 14 days or 28 days. Certain parameters of sperm function including sperm concentration and sperm motility were assessed. Furthermore, analysis of sperm DNA integrity and chromatin quality were also studied. Results: No significant changes related to sperm concentration were observed when rifampicin was given in different doses and different durations. A significant change in sperm motility were recorded only when rifampicin was given in high dose for 28 days and there was a significant reduction in sperm progressive and total motility. Rifampicin showed a significant increase in sperm DNA staining capability when the dose and duration was increased. Administration of rifampicin in high dosage for 28 days represented in larger adverse impact on structure of sperm chromatin. Conclusion: Rifampicin could negatively affect male fertility potential in rats mainly through affecting the quality of sperm chromatin structure.

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Sperm DNA fragmentation: etiology, pathogenesis, the influence on reproductive function

Urologicheskie vedomosti ◽

10.17816/uroved44804 ◽

2020 ◽

Vol 10 (4) ◽

pp. 337-345

Author(s):

Maxim N. Korshunov ◽

Ekaterina S. Korshunova ◽

Sergey P. Darenkov

Keyword(s):

Dna Fragmentation ◽

Reproductive Function ◽

Dna Integrity ◽

Sperm Dna Fragmentation ◽

Dna Breaks ◽

Sperm Dna Damage ◽

Sperm Dna ◽

Recurrent Pregnancy Losses ◽

Clinical Interest ◽

The Relationship

The literature review of sperm DNA fragmentation is evaluated. Russian and foreign literary data over the past 10 years, including fundamental researching of the evaluation of the gametes genome integrity, are analyzed. The main etiological factors and the possible reasons of the DNA breaks formation on the different stages of spermatogenesis are described. The influence of the sperm oxidative stress reaction to the DNA integrity is analyzed. The relationship between DNA fragmentation of spermatozoa pregnancy and live birth rates in the assisted reproductive techniqueare noted. Risk of the recurrent pregnancy losses in male infertility cases with the sperm DNA damage is presented. Review confirms the significant prognostic value of sperm DNA fragmentation detection in infertility cases. Further studies in evaluation of pathogenesis of sperm DNA have a clinical interest to reproductive health physicians.

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Influence of bovine sperm DNA fragmentation and oxidative stress on early embryo in vitro development outcome

Reproduction ◽

10.1530/rep-13-0123 ◽

2013 ◽

Vol 146 (5) ◽

pp. 433-441 ◽

Cited By ~ 48

Author(s):

Renata Simões ◽

Weber Beringui Feitosa ◽

Adriano Felipe Perez Siqueira ◽

Marcilio Nichi ◽

Fabíola Freitas Paula-Lopes ◽

...

Keyword(s):

Oxidative Stress ◽

Dna Fragmentation ◽

Sperm Chromatin ◽

Dna Integrity ◽

Sperm Dna Fragmentation ◽

Cleavage Rate ◽

Sperm Dna Integrity ◽

Sperm Dna ◽

Significant Difference

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.

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Effect of Alcohol Consumption on the Sperm DNA Integrity: A Systematic Review

Journal of Agricultural Science ◽

10.5539/jas.v9n13p136 ◽

2017 ◽

Vol 9 (13) ◽

pp. 136

Author(s):

Farah Hanan Fathihah Jaafar ◽

Khairul Osman ◽

Jaya Kumar ◽

Siti Fatimah Ibrahim

Keyword(s):

Dna Damage ◽

Alcohol Consumption ◽

Alcohol Withdrawal ◽

Semen Parameters ◽

Sperm Chromatin ◽

Dna Integrity ◽

Sperm Dna Damage ◽

Sperm Dna Integrity ◽

Sperm Dna ◽

Keywords Alcohol

There is no solid conclusion on the conventional sperm parameters in association with alcohol consumption, evaluation of sperm DNA integrity thus become a more reliable parameter. Hereby, this literature search was performed to summarize alcohol consumption on the sperm DNA integrity. A computerized database search was done through MEDLINE via Ovid (since 1946 until August 2017) and Cochrane was used. The following set of keywords: ‘alcohol consumption OR alcohol intake OR alcohol diet OR drinking alcohol OR ethanol diet’ AND ‘sperm DNA OR sperm chromatin OR sperm genome OR sperm histone OR sperm protamine’ were utilised. 24 articles were retrieved where only five studies conform to the inclusion criteria All studies demonstrated a negative effect of alcohol consumption on sperm DNA integrity, regardless of various range of alcohol doses and duration of alcohol consumption. Out of five studies reviewed, four studies were using a different approach to measure the sperm DNA damage. Hereby, this review identified a need to use a single approach of DNA damage test by having various method of alcohol administration and/or vice versa so that the extension of sperm DNA damage to alcohol consumption will have a better conclusion. On the same note, a few studies have reported the reversibility on conventional semen parameters, none has been done on the sperm DNA damage upon alcohol withdrawal. Therefore, the role of alcohol withdrawal on the reversibility of sperm DNA damage needs to be as well investigated further.

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Sperm DNA fragmentation induced by DNAse I and hydrogen peroxide: an in vitro comparative study among different mammalian species

Reproduction ◽

10.1530/rep-10-0176 ◽

2010 ◽

Vol 140 (3) ◽

pp. 445-452 ◽

Cited By ~ 37

Author(s):

Paola Villani ◽

Patrizia Eleuteri ◽

Maria Giuseppa Grollino ◽

Michele Rescia ◽

Pierluigi Altavista ◽

...

Keyword(s):

Hydrogen Peroxide ◽

Dna Damage ◽

Comet Assay ◽

Chromatin Structure ◽

Dnase I ◽

Sperm Chromatin ◽

Sperm Dna Fragmentation ◽

Dna Breaks ◽

Sperm Dna

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.

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Sperm DNA Integrity Assessment: A New Tool in Diagnosis and Treatment of Fertility

Obstetrics and Gynecology International ◽

10.1155/2012/531042 ◽

2012 ◽

Vol 2012 ◽

pp. 1-6 ◽

Cited By ~ 36

Author(s):

Mona Bungum

Keyword(s):

Semen Quality ◽

Significant Proportion ◽

Semen Parameters ◽

Sperm Chromatin ◽

Dna Integrity ◽

Sperm Dna Fragmentation ◽

Integrity Assessment ◽

Sperm Dna Integrity ◽

Sperm Dna

Infertility affects 15% of all couples. Although male infertility factors with reduced semen quality are contributing to about half of all involuntary childlessness, the value of standard semen parameters in prediction of fertilityin vivoand choice of proper method for assisted reproduction is limited. In the search for better markers of male fertility, during the last 10 years, assessment of sperm DNA integrity has emerged as a strong new biomarker of semen quality that may have the potential to discriminate between infertile and fertile men. Sperm DNA Fragmentation Index (DFI) as assessed by the flow cytometric Sperm Chromatin Structure Assay (SCSA) can be used for evaluation of sperm chromatin integrity. The biological background for abnormal DFI is not completely known, but clinical data show that DFI above 30% is associated with very low chance for achieving pregnancy in natural way or by insemination, but notin vitro. Already when the DFI is above 20%, the chance of natural pregnancy may be reduced, despite other sperm parameters being normal. Thus this method may explain a significant proportion of cases of unexplained infertility and can be beneficial in counselling involuntary childless couples need ofin vitrofertilisation.

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Single and Double Strand Sperm DNA Damage: Different Reproductive Effects on Male Fertility

Genes ◽

10.3390/genes10020105 ◽

2019 ◽

Vol 10 (2) ◽

pp. 105 ◽

Cited By ~ 14

Author(s):

Jordi Ribas-Maynou ◽

Jordi Benet

Keyword(s):

Dna Damage ◽

Male Fertility ◽

Sperm Chromatin ◽

Dna Breaks ◽

Sperm Dna Damage ◽

Reproductive Effects ◽

Double Strand Dna Breaks ◽

Sperm Dna ◽

Break Points ◽

Double Strand Dna

Reproductive diseases have become a growing worldwide problem and male factor plays an important role in the reproductive diagnosis, prognosis and design of assisted reproductive treatments. Sperm cell holds the mission of carrying the paternal genetic complement to the oocyte in order to contribute to an euploid zygote with proper DNA integrity. Sperm DNA fragmentation had been used for decades as a male fertility test, however, its usefulness have arisen multiple debates, especially around Intracytoplasmic Sperm Injection (ICSI) treatments. In the recent years, it has been described that different types of sperm DNA breaks (single and double strand DNA breaks) cause different clinical reproductive effects. On one hand, single-strand DNA breaks are present extensively as a multiple break points in all regions of the genome, are related to oxidative stress and cause a lack of clinical pregnancy or an increase of the conception time. On the other hand, double-strand DNA breaks are mainly localized and attached to the sperm nuclear matrix as a very few break points, are possibly related to a lack of DNA repair in meiosis and cause a higher risk of miscarriage, low embryo quality and higher risk of implantation failure in ICSI cycles. The present work also reviews different studies that may contribute in the understanding of sperm chromatin as well as treatments to prevent sperm DNA damage.

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A cost for high levels of sperm competition in rodents: increased sperm DNA fragmentation

Proceedings of The Royal Society B Biological Sciences ◽

10.1098/rspb.2015.2708 ◽

2016 ◽

Vol 283 (1826) ◽

pp. 20152708 ◽

Cited By ~ 5

Author(s):

Javier delBarco-Trillo ◽

Olga García-Álvarez ◽

Ana Josefa Soler ◽

Maximiliano Tourmente ◽

José Julián Garde ◽

...

Keyword(s):

Dna Damage ◽

Sperm Competition ◽

Dna Fragmentation ◽

Sperm Chromatin ◽

Dna Integrity ◽

Sperm Dna Fragmentation ◽

Sperm Dna Damage ◽

Sperm Dna Integrity ◽

Sperm Dna ◽

Physical And Chemical

Sperm competition, a prevalent evolutionary process in which the spermatozoa of two or more males compete for the fertilization of the same ovum, leads to morphological and physiological adaptations, including increases in energetic metabolism that may serve to propel sperm faster but that may have negative effects on DNA integrity. Sperm DNA damage is associated with reduced rates of fertilization, embryo and fetal loss, offspring mortality, and mutations leading to genetic disease. We tested whether high levels of sperm competition affect sperm DNA integrity. We evaluated sperm DNA integrity in 18 species of rodents that differ in their levels of sperm competition using the sperm chromatin structure assay. DNA integrity was assessed upon sperm collection, in response to incubation under capacitating or non-capacitating conditions, and after exposure to physical and chemical stressors. Sperm DNA was very resistant to physical and chemical stressors, whereas incubation in non-capacitating and capacitating conditions resulted in only a small increase in sperm DNA damage. Importantly, levels of sperm competition were positively associated with sperm DNA fragmentation across rodent species. This is the first evidence showing that high levels of sperm competition lead to an important cost in the form of increased sperm DNA damage.

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