Sperm chromatin condensation and single‐ and double‐stranded DNA damage as important parameters to define male factor related recurrent miscarriage

2020 ◽  
Vol 87 (11) ◽  
pp. 1126-1132
Author(s):  
Jordi Ribas‐Maynou ◽  
Carlos Abad ◽  
Sergio García‐Segura ◽  
Maria Oliver‐Bonet ◽  
Elena Prada ◽  
...  
Keyword(s):  
Dna Damage ◽  
Recurrent Miscarriage ◽  
Chromatin Condensation ◽  
Sperm Chromatin ◽  
Male Factor ◽  
Double Stranded Dna ◽  
Sperm Chromatin Condensation

308 GUANACO SPERM CHROMATIN EVALUATION USING TOLUIDINE BLUE

2010 ◽  
Vol 22 (1) ◽  
pp. 310 ◽  
Author(s):  
M. I. Carretero ◽  
S. Giuliano ◽  
A. Agüero ◽  
M. Pinto ◽  
M. Miragaya ◽  
...  
Keyword(s):  
Toluidine Blue ◽  
Chromatin Condensation ◽  
Positive Control ◽  
Sperm Chromatin ◽  
Chromatin Decondensation ◽  
South American Camelids ◽  
Splitting Factor ◽  
Dark Violet ◽  
High Degree ◽  
Sperm Chromatin Condensation

Guanacos, a wild species of South American camelids, have a high-quality fiber with great economic potential. To evaluate reproductive aptitude in guanacos, our laboratory has developed a reliable semen collection technique using electroejaculation and has applied various methods for evaluating semen characteristics. Studies for evaluating the state of sperm chromatin have also been initiated. Toluidine blue (TB) is a cationic stain that unites with the phosphate groups in the DNA, thus permitting differentiation between sperm heads according to the degree of chromatin decondensation. The objectives of this study were to determine the TB staining patterns of guanaco sperm chromatin, establish a positive control for the stain, and evaluate the effect of collagenase on sperm chromatin condensation. Semen was collected from 4 guanacos, between 6 and 9 years old, using electroejaculation. In Experiment 1, to establish a positive control for the stain, equal quantities of 1% dithiothreitol (DTT) and raw semen were incubated at room temperature for 30 s, 1.5 min, and 3 min. After incubation, smears were made and then dried, to avoid continuing the reaction, and finally were stained with 0.02% TB. A split-plot design was used with time as the splitting factor and considering the males as a block. In Experiment 2, raw semen was divided into 2 aliquots, one diluted 4 : 1 in 0.1% collagenase in HEPES-TALP-BSA medium and the other left without enzyme. Both aliquots were incubated 4 min at 37°C and, after centrifugation to remove the enzyme, smears were made and stained with TB. Spermatozoa were classified according to the degree of chromatin decondensation. Analysis of variance was performed using the males as a blocking factor and the treatment as a fixed factor. According to the degree of chromatin decondensation, three patterns of staining with TB were observed: light blue (negative, without alteration of chromatin condensation), light violet (intermediate, some degree of decondensation), and dark violet (positive, high degree of decondensation). A significant increase (P < 0.05) of sperm with highly decondensed chromatin was observed in semen incubated for 3 min with DTT when compared to 30 s of incubation. Therefore, 3 min of incubation with DTT was chosen as the positive control for Experiment 2. No significant differences in any of the 3 patterns of TB staining were observed between semen incubated with or without 0.1% collagenase. In conclusion, it is possible to use TB to evaluate the degree of chromatin decondensation in guanaco spermatozoa and to use DTT as a positive control for the stain. Treatment of guanaco semen with 0.1% collagenase did not affect sperm chromatin condensation; therefore, this enzyme can be used to decrease semen viscosity and aid handling in the laboratory.

scholarly journals The Cannabinoid Receptor CB1 Stabilizes Sperm Chromatin Condensation Status During Epididymal Transit by Promoting Disulphide Bond Formation

2020 ◽  
Vol 21 (9) ◽  
pp. 3117 ◽  
Author(s):  
Teresa Chioccarelli ◽  
Francesco Manfrevola ◽  
Veronica Porreca ◽  
Silvia Fasano ◽  
Lucia Altucci ◽  
...  
Keyword(s):  
Gene Deletion ◽  
Cannabinoid Receptor ◽  
Chromatin Condensation ◽  
Sperm Chromatin ◽  
Histone H4 ◽  
Nuclear Condensation ◽  
Heterozygous Condition ◽  
Knock Out ◽  
The Impact ◽  
Sperm Chromatin Condensation

The cannabinoid receptor CB1 regulates differentiation of spermatids. We recently characterized spermatozoa from caput epididymis of CB1-knock-out mice and identified a considerable number of sperm cells with chromatin abnormality such as elevated histone content and poorly condensed chromatin. In this paper, we extended our findings and studied the role of CB1 in the epididymal phase of chromatin condensation of spermatozoa by analysis of spermatozoa from caput and cauda epididymis of wild-type and CB1-knock-out mouse in both a homozygous or heterozygous condition. Furthermore, we studied the impact of CB1-gene deletion on histone displacement mechanism by taking into account the hyperacetylation of histone H4 and players of displacement such as Chromodomain Y Like protein (CDYL) and Bromodomain testis-specific protein (BRDT). Our results show that CB1, via local and/or endocrine cell-to-cell signaling, modulates chromatin remodeling mechanisms that orchestrate a nuclear condensation extent of mature spermatozoa. We show that CB1-gene deletion affects the epididymal phase of chromatin condensation by interfering with inter-/intra-protamine disulphide bridges formation, and deranges the efficiency of histone removal by reducing the hyper-acetylation of histone H4. This effect is independent by gene expression of Cdyl and Brdt mRNA. Our results reveal a novel and important role for CB1 in sperm chromatin condensation mechanisms.

Effects of testicular sperm chromatin condensation assay using aniline blue-eosin staining in IVF-ET cycle

2011 ◽  
Vol 96 (3) ◽  
pp. S173-S174
Author(s):  
Y.-S. Park ◽  
M.K. Kim ◽  
S.-H. Lee ◽  
J.W. Cho ◽  
I.O. Song ◽  
...  
Keyword(s):  
Chromatin Condensation ◽  
Aniline Blue ◽  
Sperm Chromatin ◽  
Testicular Sperm ◽  
Sperm Chromatin Condensation ◽  
Ivf Et

scholarly journals PREDICTIVE VALUE OF SPERM CHROMATIN CONDENSATION (ANILINE BLUE STAINING) IN THE ASSESSMENT OF MALE FERTILITY

2001 ◽  
Vol 46 (2) ◽  
pp. 99-104 ◽  
Author(s):  
M. E. Hammadeh ◽  
T. Zeginiadov ◽  
P. Rosenbaum ◽  
T. Georg ◽  
W. Schmidt ◽  
...  
Keyword(s):  
Predictive Value ◽  
Male Fertility ◽  
Chromatin Condensation ◽  
Aniline Blue ◽  
Blue Staining ◽  
Sperm Chromatin ◽  
Aniline Blue Staining ◽  
Sperm Chromatin Condensation

Low lead environmental exposure alters semen quality and sperm chromatin condensation in northern Mexico

2005 ◽  
Vol 20 (2) ◽  
pp. 221-228 ◽  
Author(s):  
Isabel Hernández-Ochoa ◽  
Gonzalo García-Vargas ◽  
Lizbeth López-Carrillo ◽  
Marisela Rubio-Andrade ◽  
Javier Morán-Martínez ◽  
...  
Keyword(s):  
Environmental Exposure ◽  
Semen Quality ◽  
Chromatin Condensation ◽  
Sperm Chromatin ◽  
Northern Mexico ◽  
Sperm Chromatin Condensation

scholarly journals Evidence that single-stranded DNA breaks are a normal feature of koala sperm chromatin, while double-stranded DNA breaks are indicative of DNA damage

Reproduction ◽  
2009 ◽  
Vol 138 (2) ◽  
pp. 267-278 ◽  
Author(s):  
Yeng Peng Zee ◽  
Carmen López-Fernández ◽  
F Arroyo ◽  
Stephen D Johnston ◽  
William V Holt ◽  
...  
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Dna Fragmentation ◽  
Sperm Nucleus ◽  
Severe Form ◽  
Sperm Chromatin ◽  
Dna Breaks ◽  
Single Stranded Dna ◽  
Double Stranded Dna ◽  
Dispersion Test

In this study, we have used single and double comet assays to differentiate between single- and double-stranded DNA damage in an effort to refine the interpretation of DNA damage in mature koala spermatozoa. We have also investigated the likelihood that single-stranded DNA breakage is part of the natural spermiogenic process in koalas, where its function would be the generation of structural bends in the DNA molecule so that appropriate packaging and compaction can occur. Koala spermatozoa were examined using the sperm chromatin dispersion test (SCDt) and comet assays to investigate non-orthodox double-stranded DNA. Comet assays were conducted under 1) neutral conditions; and 2) neutral followed by alkaline conditions (double comet assay); the latter technique enabled simultaneous visualisation of both single-stranded and double-stranded DNA breaks. Following the SCDt, there was a continuum of nuclear morphotypes, ranging from no apparent DNA fragmentation to those with highly dispersed and degraded chromatin. Dispersion morphotypes were mirrored by a similar diversity of comet morphologies that could be further differentiated using the double comet assay. The majority of koala spermatozoa had nuclei with DNA abasic-like residues that produced single-tailed comets following the double comet assay. The ubiquity of these residues suggests that constitutive alkali-labile sites are part of the structural configuration of the koala sperm nucleus. Spermatozoa with ‘true’ DNA fragmentation exhibited a continuum of comet morphologies, ranging from a more severe form of alkaline-susceptible DNA with a diffuse single tail to nuclei that exhibited both single- and double-stranded breaks with two comet tails.

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