Simultaneous determination in situ of DNA fragmentation and 8-oxoguanine in human sperm

Santiso, Rebeca; Tamayo, María; Gosálvez, Jaime; Meseguer, Marcos; Garrido, Nicolás; Fernández, José Luís

doi:10.1016/j.fertnstert.2009.07.969

Cited by 44 publications

(29 citation statements)

References 13 publications

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“…Here we showed that the expression of 8-OHdG, a bio product specific for oxidant-induced DNA damage (Kasai, 1997) detected in spermatozoa at single-cell level using an immunofluorescence method coupled to flow cytometry (Cambi et al, 2013), is strongly associated with sperm DNA damage detected by TUNEL assay. This confirmed previous data obtained with fluorescence microscopy or flow cytometry to measure a binding protein to 8-OHdG (OxyDNA assay) (De Iuliis et al, 2009;Santiso et al, 2010;Zribi et al, 2011). All results are consistent with the contention that DNA damage in spermatozoa is mainly oxidatively induced (Aitken et al, 2014).…”

Section: (A) (B) (C) (D)supporting

confidence: 92%

“…Sperm DNA fragmentation may also be the result of a persistence of DNA strand breaks (Sakkas et al, 2002) and of abnormal chromatin condensation (De Iuliis et al, 2009) during spermiogenesis and it may be oxidatively induced (Aitken et al, 2014). This is substantially supported by the observation that sperm DNA fragmentation measured by the terminal deoxynucleotidyl transferase (TdT)-mediated fluorescein-dUTP nick end labeling (TUNEL) or sperm chromatin dispersion assays is highly associated with the expression of sperm 8-hydroxy-2 0 -deoxyguanosine (8-OHdG) (De Iuliis et al, 2009;Santiso et al, 2010). The latter is a bio product specific for an oxidant-induced DNA damage (Kasai, 1997) and can be measured on a single-cell basis in ejaculated spermatozoa (De Iuliis et al, 2009;Cambi et al, 2013), therefore its determination may explore the relationship between semen infiltration of leukocytes and sperm DNA damage.…”

Section: Introductionmentioning

confidence: 95%

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Semen leukocytes and oxidative-dependent DNA damage of spermatozoa in male partners of subfertile couples with no symptoms of genital tract infection

et al. 2016

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SUMMARYThe influence of seminal leukocytes on generation of oxidative damage to sperm DNA was here investigated on male partners of subfertile couples asymptomatic for a genital tract infection. The study included 111 ejaculates from men attending the Andrology Centre at University of L'Aquila. Semen leukocytes subset included round cells expressing pan-leukocyte CD45 antigen, monocyte/ macrophage lineage antigen CD14, and activated macrophages HLA-DR antigen. In conclusion, oxidative-dependent DNA damage in spermatozoa was associated to poor semen quality but not to different leukocyte subpopulations in ejaculates of men asymptomatic for a genital tract infection.

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Section: (A) (B) (C) (D)supporting

confidence: 92%

Section: Introductionmentioning

confidence: 95%

Semen leukocytes and oxidative-dependent DNA damage of spermatozoa in male partners of subfertile couples with no symptoms of genital tract infection

et al. 2016

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“…The OxyDNA kit is based on the use of a fluorescent binding protein that localizes the yielded fluorescence in sperm nuclei (Santiso et al (2010) and this study). 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%).…”

Section: Discussionmentioning

confidence: 99%

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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“…8-OH-2-deoxyguanosine (oxo8dG) type is one of the commonly nucleotide transformation derived from the aforesaid processes. This type of base mutation can be easily detected using specific immunoassaying on spermatozoa and its presence can be directly related to the presence of DNA fragmentation [142]. To explain the decreased reproductive potential of men with elevated levels of sperm DNA damage, it is possible that the remaining spermatozoa with no measurable DNA damage have some type of damage at varying extents [143].…”

Section: Clinical Utility Of Sperm Functional Tests In Unexplained Mamentioning

confidence: 99%

Role and Significance of Sperm Function in Men with Unexplained Infertility

Esteves¹,

Verza²,

Sharma

et al. 2015

Unexplained Infertility

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Simultaneous determination in situ of DNA fragmentation and 8-oxoguanine in human sperm

Cited by 44 publications

References 13 publications

Semen leukocytes and oxidative-dependent DNA damage of spermatozoa in male partners of subfertile couples with no symptoms of genital tract infection

Semen leukocytes and oxidative-dependent DNA damage of spermatozoa in male partners of subfertile couples with no symptoms of genital tract infection

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

Role and Significance of Sperm Function in Men with Unexplained Infertility

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