Sperm DNA fragmentation induced by cryopreservation: new insights and effect of a natural extract from Opuntia ficus-indica

Meamar, Mehrdad; Zribi, Nassira; Cambi, Marta; Tamburrino, Lara; Marchiani, Sara; Filimberti, E.; Fino, Maria Grazia; Biggeri, Annibale; Ménézo, Yves; Forti, Gianni; Baldi, Elisabetta; Muratori, Monica

doi:10.1016/j.fertnstert.2012.05.001

Cited by 53 publications

(45 citation statements)

References 35 publications

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“…The occurrence of 8-OHdG was detected in spermatozoa counterstained with PI (Figs 1, 2 and 3) in order to eliminate anucleated interferents present in semen (Marchiani et al 2007, Muratori et al 2008. Besides allowing a more accurate evaluation of sperm parameters (Muratori et al 2008, Marchiani et al 2011, PI staining reveals the occurrence of two different sperm populations, named PI br and PI dim , as recently reported by our group (Muratori et al 2008, Marchiani et al 2011, Meamar et al 2012. Interestingly, in all tested samples, only PI br spermatozoa showed signs of oxidative DNA damage, whereas virtually no cell in PI dim population presented 8-OHdG residues (Figs 2 and 3).…”

Section: Relationship Between 8-ohdg Levels and Semen Qualitymentioning

confidence: 71%

“…In our study, the labelling of 8-OHdG was coupled to flow cytometry detection that allowed us to reveal 8-OHdG separately in the PI br and PI dim sperm populations, showing a variable incidence among subfertile subjects (Muratori et al 2008, Marchiani et al 2011, Meamar et al 2012. These populations occur in human semen and were revealed because of a difference in PI staining (Muratori et al 2008), detectable in flow cytometry but not in fluorescence microscopy.…”

Section: Discussionmentioning

confidence: 93%

“…In our hands, OxyDNA kit produced measures (37.2G30.7%) comparable to those previously reported in similar sperm preparations (Meseguer et al 2008, Aitken et al 2010, Bellver et al 2010, albeit, on average, fourfold greater than those yielded by the immunofluorescence method (11.4G6.9%). It is anticipated that such a discrepancy is not due to a hampered access of the antibody molecules into sperm nuclei as no difference was observed when the 8-OHdG labelling was obtained after a step of (Muratori et al 2008, Marchiani et al 2011, Meamar et al 2012 and that only PI br spermatozoa show 8-OHdG fluorescence. (Muratori et al 2003) as well as a reduction of progressive motility (this study) without increasing the amount of sperm with severe cell damage that could undergo degradation during the treatment and thus provoke a cell selection (Muratori et al 2010).…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, in all tested samples, only PI br spermatozoa showed signs of oxidative DNA damage, whereas virtually no cell in PI dim population presented 8-OHdG residues (Figs 2 and 3). (Muratori et al 2008, Marchiani et al 2011, Meamar et al 2012. Effect of exceeding concentrations of 8-OHdG on the antibody 15A3 (A, right panels) and the BP-F (B, right panels) ability to detect 8-OHdG.…”

Section: Relationship Between 8-ohdg Levels and Semen Qualitymentioning

confidence: 99%

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Development of a specific method to evaluate 8-hydroxy,2-deoxyguanosine in sperm nuclei: relationship with semen quality in a cohort of 94 subjects

Cambi¹,

Tamburrino²,

Marchiani³

et al. 2013

Reproduction

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Oxidative stress (OS) is involved in many disoders including male infertility. Human spermatozoa are very sensitive targets of reactive oxygen species (ROS) and most sperm functions are impaired in the case of OS. In addition unbalanced production of ROS is considered one of the most important causes of sperm DNA fragmentation, a semen trait of infertile men. The relationship between oxidative damage and semen quality is partially controversial, probably due to the different methods and/or targets used to reveal the OS. In this study, by fluorescence microscopy and flow cytometry, we compared two methods to reveal 8-hydroxy,2-deoxyguanosine (8-OHdG), the hallmark of oxidative DNA damage: an immunofluorescence method and the commercial OxyDNA kit. We found that although both methods localized the labelling in sperm nuclei they yielded different measures, and only with the immunofluorescence method was the labelling specific for sperm 8-OHdG. The immunofluorescence method, coupled to flow cytometry, was thus selected to analyse the 8-OHdG content in semen samples from 94 subfertile patients and to investigate the relationship with semen quality. We found that the percentages of spermatozoa with 8-OHdG (meanGS.D., 11.4G6.9%) were related to sperm count (Pearson's correlation coefficient (r)ZK0.27, PZ0.04 (ANOVA and student's t-test)), motility (progressive: rZK0.22, PZ0.04; non-progressive: rZ0.25, PZ0.01), and normal morphology (rZK0.27, PZ0.01).In conclusion, we demonstrate that immunofluorescence/flow cytometry is a reliable and specific method to detect 8-OHdG at single-cell level and show that oxidative damage only partially overlaps poor semen quality, suggesting that it could provide additional information on male fertility with respect to routine semen analysis.

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Section: Relationship Between 8-ohdg Levels and Semen Qualitymentioning

confidence: 71%

Section: Discussionmentioning

confidence: 93%

Section: Discussionmentioning

confidence: 99%

Section: Relationship Between 8-ohdg Levels and Semen Qualitymentioning

confidence: 99%

See 2 more Smart Citations

Development of a specific method to evaluate 8-hydroxy,2-deoxyguanosine in sperm nuclei: relationship with semen quality in a cohort of 94 subjects

Cambi¹,

Tamburrino²,

Marchiani³

et al. 2013

Reproduction

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show abstract

“…Freezing and thawing of sperm was performed as described previously (Meamar et al 2012). Briefly, each semen sample (nZ6) was divided into four aliquots.…”

Section: Evaluation Of Sdf and Sumo1 After Freezing And Thawing And Hmentioning

confidence: 99%

SUMO1 in human sperm: new targets, role in motility and morphology and relationship with DNA damage

et al. 2014

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In studies carried out previously, we demonstrated that small ubiquitin-like modifier 1 (SUMO1) is associated with poor sperm motility when evaluated with a protocol that reveals mostly SUMO1-ylated live sperm. Recently, with another protocol, it has been demonstrated that SUMO is expressed in most sperm and is related to poor morphology and motility, suggesting that sumoylation may have multiple roles depending on its localisation and targets. We show herein, by confocal microscopy and co-immunoprecipitation, that dynaminrelated protein 1 (DRP1), Ran GTPase-activating protein 1 (RanGAP1) and Topoisomerase IIa, SUMO1 targets in somatic and/or germ cells, are SUMO1-ylated in mature human spermatozoa. DRP1 co-localises with SUMO1 in the mid-piece, whereas RanGAP1 and Topoisomerase IIa in the post-acrosomal region of the head. Both SUMO1 expression and co-localisation with the three proteins were significantly higher in morphologically abnormal sperm, suggesting that sumoylation represents a marker of defective sperm. DRP1 sumoylation at the mid-piece level was higher in the sperm of asthenospermic men. As in somatic cells, DRP1 sumoylation is associated with mitochondrial alterations, this protein may represent the link between SUMO and poor motility. As SUMO pathways are involved in responses to DNA damage, another aim of our study was to investigate the relationship between sumoylation and sperm DNA fragmentation (SDF). By flow cytometry, we demonstrated that SUMO1-ylation and SDF are correlated (rZ0.4, P!0.02, nZ37) and most sumoylated sperm shows DNA damage in co-localisation analysis. When SDF was induced by stressful conditions (freezing and thawing and oxidative stress), SUMO1-ylation increased. Following freezing and thawing, SUMO1-Topoisomerase IIa co-localisation and co-immunoprecipitation increased, suggesting an involvement in the formation/repair of DNA breakage.

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Interventions to Prevent Sperm DNA Damage Effects on Reproduction

Esteves

2019

Genetic Damage in Human Spermatozoa

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Sperm DNA fragmentation induced by cryopreservation: new insights and effect of a natural extract from Opuntia ficus-indica

Cited by 53 publications

References 35 publications

Development of a specific method to evaluate 8-hydroxy,2-deoxyguanosine in sperm nuclei: relationship with semen quality in a cohort of 94 subjects

Development of a specific method to evaluate 8-hydroxy,2-deoxyguanosine in sperm nuclei: relationship with semen quality in a cohort of 94 subjects

SUMO1 in human sperm: new targets, role in motility and morphology and relationship with DNA damage

Interventions to Prevent Sperm DNA Damage Effects on Reproduction

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