Low levels of mouse sperm chromatin fragmentation delay embryo development

Numerous reports show that the epididymis plays a key role in the acquisition of sperm fertilizing ability but less information exists on its contribution to embryo development. Evidence from our laboratory showed that mammalian CRISP (Cysteine-Rich Secretory Proteins), known to be expressed in the epididymis, to regulate calcium (Ca2+) channels and to participate in fertilization, may also be relevant for embryo development. More specifically, we found that males with simultaneous mutations inCrisp1andCrisp3genes exhibited normalin vivofertilization but impaired embryo development. In the present work, aimed to investigate the mechanisms underlying this reproductive phenotype, we observed that embryo development failure was not due to delayed fertilization as no differences in sperm transport within the female tract nor inin vivofertilization were found shortly after mating. The observation that impaired embryo development was also found in eggs fertilized by epididymal sperm either after uterine insemination orin vitrofertilization, revealed that the defects were already present at epididymal level. Of note, eggs fertilizedin vitroby mutant sperm exhibited impaired meiotic resumption not due to defects in Ca2+oscillations during egg activation, prompting us to examine potential sperm DNA defects. Interestingly, higher levels of both DNA fragmentation and intracellular Ca2+ were observed for mutant than for control epididymal sperm, supporting sperm DNA damage, likely linked to a Ca2+ dysregulation, as the main responsible for the early development failure of mutant males. Together, our results support the contribution of the epididymis beyond fertilization, identifying CRISP1 and CRISP3 as novel male factors relevant for DNA integrity and early embryo development. Given the existence of human functional homologues of CRISP and the incidence of DNA fragmentation in infertile men, we believe these findings not only provide relevant information on the impact of epididymal factors on embryonic development but will also contribute to a better understanding, diagnosis and treatment of human infertility.

show abstract

Cryopreservation of bovine sperm causes single-strand DNA breaks that are localized in the toroidal regions of chromatin

Ribas-Maynou,

Muiño,

Tamargo

et al. 2024

J Animal Sci Biotechnol

View full text Add to dashboard Cite

Background Sperm cryopreservation is widely used in the cattle industry, as it allows for disassociating the localization of sires and the collection of semen from the timing of artificial insemination. While freeze-thawing is known to impair sperm DNA integrity, whether the damage induced consists of single- (SSB) or double-strand breaks (DSB) has not been determined. In addition, no previous study has addressed if DNA breaks preferentially reside in specific genome regions such as those forming the toroid linker regions, or are rather spread throughout the regions linked to protamines. The main aim of the present work, therefore, was to elucidate the type and localization of the DNA damage generated by cryopreservation and to evaluate its impact on artificial insemination outcomes in cattle. Results The incidence of SSB and DSB was evaluated in 12 ejaculates before and after cryopreservation with the Comet assay, and the localization of the DNA breaks was assessed using pulsed-field gel electrophoresis (PFGE). Before cryopreservation, the incidence of SSB was 10.99% ± 4.62% and involved 20.56% ± 3.04% of sperm cells, whereas these figures significantly (P < 0.0001) increased up to 34.11% ± 3.48% and 53.36% ± 11.00% in frozen-thawed sperm. In contrast, no significant differences in the incidence of DSB were observed (P > 0.990) before and after cryopreservation (before: incidence of 13.91% ± 1.75% of sperm DNA affecting 56.04% ± 12.49% of sperm cells; after: incidence of 13.55% ± 1.55% of sperm DNA involving 53.36% ± 11.00% of sperm cells). Moreover, PFGE revealed that the percentage of sperm DNA fragments whose length was shorter than a toroid (< 31.5 kb) was greater (P < 0.0001) after (27.00% ± 4.26%) than before freeze-thawing (15.57% ± 4.53%). These differences indicated that the DNA breaks induced by cryopreservation affect the regions condensed in protamines, which are structured in toroids. On the other hand, in vivo fertility rates were associated to the incidence of SSB and DSB in frozen-thawed sperm (P = 0.032 and P = 0.005), but not with the size of the DNA fragments resulting from these breaks (P > 0.05). Conclusion Cryopreservation of bovine sperm generates single-strand DNA breaks, which are mainly located in protamine-condensed toroidal regions. The incidence of DNA breaks in cryopreserved sperm has an impact on cattle fertility, regardless of the size of generated fragments.

show abstract

Low levels of mouse sperm chromatin fragmentation delay embryo development

Cited by 5 publications

References 37 publications

Paternally Mediated Developmental Toxicity

Paternally Mediated Developmental Toxicity

Contribution of the epididymis beyond fertilization: relevance of CRISP1 and CRISP3 for sperm DNA integrity and early embryo development

Cryopreservation of bovine sperm causes single-strand DNA breaks that are localized in the toroidal regions of chromatin

Contact Info

Product

Resources

About