Localization of single-stranded DNA in human sperm nuclei

Zhang, Xiaoyang; Gabriel, Maria San; Libman, Jamie; Phillips, Simon; Courchesne, Annick; Zini, Armand

doi:10.1016/j.fertnstert.2007.01.025

Cited by 9 publications

(3 citation statements)

References 29 publications

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“…The presence of single-stranded DNA breaks in the periphery of the spermatozoa may act as putative DNA nicks that facilitate the production of further single-stranded DNA motifs and ultimately manifest as double-stranded breaks in highly affected spermatozoa, such as those presenting in this study as massive haloes of chromatin dispersion of the microgel following the SCD test. While this concept has not been thoroughly investigated, there are some observations where the sperm nuclei may exhibit discrete and preferable peripheral distribution of foci containing single-stranded DNA motifs and this fact has been linked to infertility in men (Zhang et al 2007). A highly ordered form of DNA damage has also been reported in short-beaked echidna (Johnston et al 2009).…”

Section: Discussionmentioning

confidence: 99%

Osmotic stress and cryoinjury of koala sperm: an integrative study of the plasma membrane, chromatin stability and mitochondrial function

Johnston¹,

Satake²,

Zee³

et al. 2012

Reproduction

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This study investigated whether cryopreservation-induced injury to koala spermatozoa could be explained using an experimental model that mimics the structural and physiological effects of osmotic flux. DNA labelling after in situ nick translation of thawed cryopreserved spermatozoa revealed a positive correlation (rZ0.573; P!0.001; nZ50) between the area of relaxed chromatin in the nucleus and the degree of nucleotide labelling. While the chromatin of some spermatozoa increased more than eight times its normal size, not all sperm nuclei with relaxed chromatin showed evidence of nucleotide incorporation. Preferential staining associated with sperm DNA fragmentation (SDF) was typically located in the peri-acrosomal and peripheral regions of the sperm head and at the base of the spermatozoa where it appear to be 'hot spots' of DNA damage following cryopreservation. Results of the comparative effects of anisotonic media and cryopreservation on the integrity of koala spermatozoa revealed that injury induced by exposure to osmotic flux, essentially imitated the results found following cryopreservation. Plasma membrane integrity, chromatin relaxation and SDF appeared particularly susceptible to extreme hypotonic environments. Mitochondrial membrane potential (MMP), while susceptible to extreme hypo-and hypertonic environments, showed an ability to rebound from hypertonic stress when returned to isotonic conditions. Koala spermatozoa exposed to 64 mOsm/kg media showed an equivalent, or more severe, degree of structural and physiological injury to that of frozen-thawed spermatozoa, supporting the hypothesis that cryoinjury is principally associated with a hypo-osmotic effect. A direct comparison of SDF of thawed cryopreserved spermatozoa and those exposed to a 64 mOsm/kg excursion showed a significant correlation (rZ0.878; P!0.05; nZ5); however, no correlation was found when the percentage of sperm with relaxed chromatin was compared. While a cryo-induced osmotic injury model appears to explain post-thaw changes in koala SDF, the mechanisms resulting in relaxed chromatin require further study. A lack of correlation between the percentage of sperm with relaxed chromatin and SDF suggests that the timing of these pathologies are asynchronous. We propose an integrative model of cryo-induced osmotic injury that involves a combination of structural damage (rupture of membrane) and oxidative stress that first leads to the reduction of MMP and the relaxation of chromatin, which is then ultimately followed by an increase in DNA fragmentation.

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Section: Discussionmentioning

confidence: 99%

Osmotic stress and cryoinjury of koala sperm: an integrative study of the plasma membrane, chromatin stability and mitochondrial function

Johnston¹,

Satake²,

Zee³

et al. 2012

Reproduction

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“…For example, the consequences of single-or double-stranded sperm DNA breakage for fertilization or embryogenesis remain unknown, although recognized as potentially important (Genescá et al 1992, Khanna & Jackson 2001, Zhang et al 2007, and recognized in some recent studies (Gandini et al 2004, Payne et al 2005. In fish with external fertilization, artificial reproduction in hatcheries can result in embryo development problems and offspring malformations (Hauser & Sissenwine 1991, Lough et al 2008; we are now investigating relationships between DNA breakage and embryogenesis in fish.…”

Section: T0mentioning

confidence: 99%

Rapid rates of sperm DNA damage after activation in tench (Tinca tinca: Teleostei, Cyprinidae) measured using a sperm chromatin dispersion test

López‐Fernández¹,

Gage²,

Arroyo³

et al. 2009

Reproduction

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Spermatozoal haplotypic DNA is prone to damage, leading to male fertility problems. So far, the assessment of sperm DNA breakage has been challenging because protamines render the nuclear chromatin highly compacted. Here, we report the application of a new test to quantify DNA fragmentation in spermatozoa of an externally fertilizing teleost fish. The sperm chromatin dispersion (SCD) test uses a species-specific lysing solution to generate controlled protein depletion that, followed by DNA-specific fluorescent labelling, allows an easy morphological discrimination between nuclei affected by DNA damage. Using tench (Tinca tinca) as our model, we first trialled the test against established, but more technically demanding, assays employing in situ nick translation (ISNT) and the comet assay. The SCD test showed high concordance with ISNT, comet assay measures and a chromatin-swelling test, confirming the application of this straightforward SCD technique to various aspects of reproductive biology. Second, we examined between-male variation in DNA damage, and measured changes through time following spermatozoal activation. Between-male variation in the basal levels of average DNA damage ranged from 0 to 20% of sperm showing damage, and all showed increases in DNA fragmentation through time (0-60 min). The rates of DNA damage increase are the fastest so far recorded in sperm for a living organism, and may relate to the external fertilization mode. Our findings have relevance for broodstock selection and optimizing IVF protocols routinely used in modern aquaculture.

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“…The procedure gives rise to haloes of dispersed chromatin due to the spreading of nuclear DNA loops and/or fragments of DNA; when the sperm nucleus contains fragmented DNA, the size of the halo is directly proportional to the amount of sperm DNA damage. Other methodologies based on the use of specifi c antibodies against a modifi ed DNA molecule, such as presence of single strand DNA motifs (Zhang et al 2007 ), presence of 8-hydroxy-G (Santiso et al 2010 ), or indirect markers related to apoptotic processes, are also effective for targeting sperm DNA damage, although at this stage of their development they still have technical and cost constraints for routine use.…”

Section: Sperm Dna Fragmentation: What Is It?mentioning

confidence: 99%

Sperm DNA Fragmentation and Its Role in Wildlife Conservation

Gosálvez

Holt

Johnston

2014

Advances in Experimental Medicine and Biology

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Until about 20 years ago, sperm assessment in the laboratory was focused on motility, morphology and acrosomal integrity. Then came the gradual realisation that, because the main objective of a spermatozoon is to deliver an intact genetic payload of DNA to the egg, being able to check DNA quality of spermatozoa would be equally important, if not more so. Research over the last two decades has therefore led to the development of several techniques for reliably detecting DNA strand breaks, and the more recent focus has been directed towards understanding the fertility implications of DNA damage. It is now clear that evolutionary history has played an important role in determining the stability of sperm DNA under stressful conditions, and that the nature of the DNA-protein interactions also influence the extent to which fertility is affected by both technical procedures involved in sperm preservation and the basic biology of the species concerned. Here we present an overview of the principles involved in DNA assessment and also provide some cases studies that illustrate the influences of species diversity.

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Localization of single-stranded DNA in human sperm nuclei

Cited by 9 publications

References 29 publications

Osmotic stress and cryoinjury of koala sperm: an integrative study of the plasma membrane, chromatin stability and mitochondrial function

Osmotic stress and cryoinjury of koala sperm: an integrative study of the plasma membrane, chromatin stability and mitochondrial function

Rapid rates of sperm DNA damage after activation in tench (Tinca tinca: Teleostei, Cyprinidae) measured using a sperm chromatin dispersion test

Sperm DNA Fragmentation and Its Role in Wildlife Conservation

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