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

Semen quality plays a crucial role in embryo production 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 fertilised embryos in Simmental cattle.

Semen from six bulls was split 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 CASA, acrosome/plasma membrane integrity) was assessed, along with MDA, ROS, ATP, and MMP levels. For 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.

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 fertilised 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.

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 production.