Nucleons, I: A Model for Studying the Mechanism of Sperm Nucleus Swelling in Vitro

Delgado, N. M.; Sánchez-Vázquez, M L; Reyes, R.; Merchant‐Larios, Horacio

doi:10.1080/014850199262779

Cited by 8 publications

(11 citation statements)

References 53 publications

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“…Under physiological conditions, GAGs bind to specific receptors in the sperm plasma membrane [Peluso et al 1992;Sánchez-Prieto et al 1996], induce capacitation [Medeiros and Parrish 1996;Miller et al 1990;Reyes et al 1989] and the acrosome reaction Delgado et al 1988]. Moreover, they induce the release of basic nucleoproteins [Reyes et al 1991] and facilitate nuclear swelling [Delgado et al 1999, Romanato et al 2003, 2005. The observations reported in this study are consistent with these known roles of GAGs in fertilization.…”

Section: Discussionsupporting

confidence: 93%

“…To isolate nucleons from sperm nuclei [Delgado et al 1999], aliquots of 10 8 cells/0.6 ml sperm suspension were treated with 0.2 ml of 9.0 mM freshly prepared DL-dithiothreitol (DTT, Merck, Darmstadt, Germany) in 50 mM Tris-HCl, pH 8.0, and allowed to stand for 18 min at room temperature. At the end of this period, 0.2 ml of 1% N-cetyl-N,N,N-trimethyl ammonium bromide (CTAB, Merck, Darmstadt, Germany) was added and the mixture was allowed to stand for 15 min [Delgado et al 2001].…”

Section: Bovine Sperm Nuclei Preparationmentioning

confidence: 99%

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Presence and Release of Bovine Sperm Histone H1 During Chromatin Decondensation by Heparin-Glutathione

Sánchez-Vázquez

Flores-Alonso

Merchant‐Larios

et al. 2008

Systems Biology in Reproductive Medicine

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During spermatogenesis, changes in sperm nuclear morphology are associated with the replacement of core somatic histones by protamines. Although protamines are the major nucleoproteins of mature sperm, not all species totally replace the histones. Histone H1, along with protamines, mediates chromatin condensation into an insoluble complex that is transcriptionally inactive. In vitro, heparin-reduced glutathione causes sperm decondensation, and the structures formed are morphologically similar to the in vivo male pronucleus. To study the participation of histone H1 in bovine sperm chromatin remodelling, we measured the presence and release of histone H1 by immunofluorescence, acetic acid-urea-triton-polyacrylamide gel electrophoresis, and immunoblotting. Nuclear decondensation was induced by 80 microM heparin and 15.0 mM reduced glutathione (GSH) for 7, 14, and 21 h at 37 degrees C. Additionally, nucleons, composed of nuclei isolated from the sperm, were decondensed with 20.0 microM heparin and 5.0 mM GSH for 4.0 h at 37 degrees C. Controls were incubated in buffer for similar periods of time. Immunofluorescent localization of histone H1 was carried out with mouse monoclonal antibody, and DNA localization was visualized by 0.001% quinacrine staining. Chromatin decondensation was accompanied by increased sperm nuclei and nucleon surface area. We observed that histone H1 was localized exclusively in the nuclei of intact sperm and nucleons. Histone H1 immunofluorescent intensity did not change in control samples but decreased over time in samples treated with heparin-GSH. There was a negative correlation between the surface area of sperm nuclei and immunohistochemical intensity of histone H1 (P < 0.05). Nucleon decondensation showed a similar relationship. By electrophoresis and immunoblotting, we verified the loss of histone H1 from the sperm and nucleons and its release into the incubation media. Based on these results, we propose that histone H1 is present in the bovine sperm nuclei. H1 depletion may participate in chromatin decondensation and nuclear swelling induced by heparin-GSH.

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Section: Discussionsupporting

confidence: 93%

Section: Bovine Sperm Nuclei Preparationmentioning

confidence: 99%

Presence and Release of Bovine Sperm Histone H1 During Chromatin Decondensation by Heparin-Glutathione

Sánchez-Vázquez

Flores-Alonso

Merchant‐Larios

et al. 2008

Systems Biology in Reproductive Medicine

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“…It also modulates sperm capacitation and acrosome reaction through binding to its receptors on the sperm plasma membrane (Delgado et al, 1988;Sanchez-Prieto et al, 1996). Importantly, heparin binds tightly to intact nucleosomes and induces the release of histone H1 and protamine, which is a critical step in important for sperm chromatin decondensation (Delgado et al, 1999;Romanato et al, 2003Romanato et al, , 2005Villeponteau, 1992).…”

Section: Introductionmentioning

confidence: 99%

Improved Bovine ICSI Outcomes by Sperm Selected after Combined Heparin-Glutathione Treatment

Sekhavati

Shadanloo

Hosseini

et al. 2012

Cellular Reprogramming

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Despite widespread application of intracytoplasmic sperm injection (ICSI) in human-assisted reproductive techniques (ART), the efficiency of this method is still far from satisfactory in livestock, particularly in the bovine species with its unique sperm condensation. On the basis of the natural chemical structure of chromatin in condensed sperm, we developed a novel combined heparin-reduced glutathione (GSH) sperm pretreatment that improves the efficiency of bovine ICSI via selection of the most appropriate sperm at the time of ICSI. Assessment of sperm DNA integrity revealed that this pretreatment can be considered as a safe and efficient approach for in vitro sperm decondensation when compared to conventional sperm pretreatments with dithiothreitol (DTT). Injection of completely decondensed bull sperm derived from this pretreatment significantly improved fertilization and blastocyst formation rates compared to untreated or intact sperm injection (34.8 ± 2.7 and 29.1 ± 1.5 vs. 12.0 ± 3.2 and 15.9 ± 1.2%, respectively; p<0.05). Real-time PCR analysis revealed that expression of pluripotent and anti-apoptosis markers in blastocysts derived by injection of completely decondensed sperm from heparin-GSH pretreatment were comparable to IVF when compared to the DTT pretreatment and control ICSI groups (p<0.05). The results of this study suggested that the degree of sperm decondensation derived from heparin-GSH pretreatment may affect ICSI efficiency in bovine.

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“…Nucleons have been able to induce their chromatin decondensation by the action of heparin/GSH [8]. Results that joined with the one obtained from metabolic activity allow us to use nucleons for ICSI fertilization with the possibility to obtain suitable fertilization and pregnancies rates.…”

Section: Resultsmentioning

confidence: 99%

“…Nucleons (human, pig, hamster, mouse, rat, and bull) have been able to induce their chromatin decondensation by the action of heparin/GSH [8], displaying the same classic pattern of chromatin swelling of the sperm nuclei of other animals. Human nucleons have also shown a metabolic activity represented by oxygen uptake, which increases ATP levels, suggesting the presence of aerobic synthesis of ATP [22].…”

Section: R Reyes Et Almentioning

confidence: 99%

Nucleons Ii: Cryopreservation and Metabolic Activity

Reyes

Flores-Alonso²,

Rodríguez-Hernández³

et al. 2001

Archives of Andrology

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Nucleons, I: A Model for Studying the Mechanism of Sperm Nucleus Swelling in Vitro

Cited by 8 publications

References 53 publications

Presence and Release of Bovine Sperm Histone H1 During Chromatin Decondensation by Heparin-Glutathione

Presence and Release of Bovine Sperm Histone H1 During Chromatin Decondensation by Heparin-Glutathione

Improved Bovine ICSI Outcomes by Sperm Selected after Combined Heparin-Glutathione Treatment

Nucleons Ii: Cryopreservation and Metabolic Activity

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