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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.

, more cells with inactive mitochondria (class III, P = 0.001), fewer cells with active mitochondria (class I, P = 0.005) and fewer spermatozoa with intact acrosomes ( P < 0.001). Finally, no significant differences were observed in lipid peroxidation levels. CONCLUSION• Men with varicocele showed an increase in sperm DNA fragmentation and a reduction in mitochondrial activity and acrosome integrity. However, lipid peroxidation levels remained unchanged. KEYWORDSvaricocele, spermatozoa, oxidative stress, acrosome, DNA damage, mitochondria, metabolism Study Type -Aetiology (case control) Level of Evidence 3bWhat's known on the subject? and What does the study add? Varicocele leads to alterations in sperm DNA integrity even when alterations in semen quality are not yet observed in adolescents. In adults, alterations to sperm DNA are associated to altered sperm morphology, indicating that altered spermatogenesis may be an important cause for the increased sperm DNA fragmentation observed in these men. One other important cause of increased DNA fragmentation is oxidative stress, and we wished to verify if this was the case.The study adds the information that, in the adult varicocele, it is most likely that an altered testicular environment is leading to increased DNA fragmentation and decreased mitochondrial activity and acrosome integrity, because no increase in oxidative stress was observed. OBJECTIVE• To assess the effect of varicocele on sperm DNA integrity, mitochondrial activity, lipid peroxidation and acrosome integrity. PATIENTS AND METHODS• In all, 30 patients with a clinically diagnosed varicocele of grade II or III and 32 men without a varicocele were evaluated for sperm DNA fragmentation (comet assay), mitochondrial activity (3,3 ′ -diaminobenzidine assay), lipid peroxidation (malondialdehyde) and acrosome integrity (fluorescent probe labelled peanut agglutinin).

Higher temperatures lead to an increase of testicular metabolism that results in spermatic damage. Oxidative stress is the main factor responsible for testicular damage caused by heat stress. The aim of this study was to evaluate lasting effects of heat stress on ejaculated sperm and immediate or long-term effects of heat stress on epididymal sperm. We observed decrease in motility and mass motility of ejaculated sperm, as well as an increase in the percentages of sperm showing major and minor defects, damaged plasma and acrosome membranes, and a decrease in the percentage of sperm with high mitochondrial membrane potential in the treated group until one spermatic cycle. An increased enzymatic activity of glutathione peroxidase and an increase of stressed cells were observed in ejaculated sperm of the treated group. A decrease in the percentage of epididymal sperm with high mitochondrial membrane potential was observed in the treated group. However, when comparing immediate and long-term effects, we observed an increase in the percentage of sperm with low mitochondrial membrane potential. In conclusion, testicular heat stress induced oxidative stress that led to rescuable alterations after one spermatic cycle in ejaculated sperm and also after 30 days in epididymal sperm.

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BackgroundIn order to improve the efficiency of bovine sperm cryopreservation process, it is important to understand how spermatozoa respond to differences in temperature as well as the ability to recover its own metabolism. The combination between flow cytometry approach and antioxidant enzymes activity allows a more sensible evaluation of sperm cell during cryopreservation. The aim of this study was to evaluate sperm attributes and antioxidant enzymes activity during different stages of cryopreservation process. Semen samples from Holstein bulls (n = 4) were separated in 3 treatments: fresh (37 °C); cooled (5 °C); and thawed. Evaluation occurred at 0 h and 2 h after incubation. Membrane integrity, mitochondrial membrane potential (MMP) and DNA damages were evaluated by flow cytometry; activities of antioxidant enzymes such as catalase, superoxide dismutase and gluthatione peroxidase were measured by spectrofotometry.ResultsThere was an increase in the percentage of sperm with DNA damage in the thawed group, compared to fresh and cooled, and for 2 hs of incubation when compared to 0 h. Considering MMP, there was an increase in the percentage of cells with medium potential in thawed group when compared to fresh and cooled groups. Opposingly, a decrease was observed in the thawed group considering high mitochondrial potential. Also in the thawed group, there was an increase on cells with damaged acrosome and membrane when compared to fresh and cooled groups. Significant correlations were found between antioxidant enzymes activity and membrane or mitochondrial parameters.ConclusionBased on our results, we conclude that cryopreservation affects cellular and DNA integrity and that the critical moment is when sperm cells are exposed to freezing temperature. Also, our study indicates that intracellular antioxidant machinery (SOD and GPX enzymes) is not enough to control cryodamage.