Comparison of semen preservation methods and in vitro fertilisation embryo quality in Simmental cattle.

Abstract

Objective: Semen quality plays a crucial role in embryo formation in cattle. This study aims to compare the effects of 0°C refrigerated semen and liquid nitrogen-frozen semen on the quality of in vitro fertilized embryos in Simmental cattle.

Methods: Semen from six bulls was devidedt equally into two groups. One was diluted with cold storage solution and stored at 0°C for 72 h; the other with cryopreservation solution and preserved in liquid nitrogen for 72 h. Post-storage, sperm quality (motility, progressive motility via computer-assisted sperm analysis, and acrosome/plasma membrane integrity) was assessed, along with malondialdehyde, reactive oxygen species, adenosine triphosphate, and mitochondrial membrane potential levels. For in vitro fertilisation (IVF), oocytes from Simmental cows (>50 per group) were inseminated with the two semen groups. At 45 h post-IVF, embryo cleavage rate was recorded; on day 8, blastocyst rate was determined. Embryos then underwent BrdU staining (proliferation) and TUNEL staining (apoptosis), with proliferation and apoptosis levels quantified via marker genes.

Results: The results showed that compared with liquid nitrogen-frozen semen, 0°C refrigerated semen exhibited significantly higher sperm motility, forward motility rate, acrosome integrity, and plasma membrane integrity (p<0.05). in vitro fertilized embryos also exhibited significant increases in quality and quantity (cleavage rate and blastocyst rate) (p<0.05), and embryos derived from 0°C refrigerated semen contained significantly more proliferating cells (p<0.05) and fewer apoptotic cells (p<0.05) than those from frozen semen.

Conclusion: These findings suggest that 0°C refrigerated semen is superior to liquid nitrogen frozen semen in terms of sperm quality parameters and in vitro embryo development.