Impact of Particle Size of Nanoliposomes on the Biological Response of Frozen-Thawed Sperm in Holstein Bulls.

Abstract

Nanoliposomes could improve the delivering profile of protecting substances in sperm-freezing extenders. This study aimed to assess the impact of nanoliposomes with varying particle sizes and containing 2.5 mM glutathione (GSH) on the post-thaw quality of bull sperm. Ejaculates were obtained weekly from six mature Holstein bulls for six weeks using an artificial vagina. Semen was pooled and mixed with the lecithin-based extender. Semen was pooled and diluted with lecithin-based extenders containing 2.5 mM encapsulated GSH in nanoliposomes with particles in the size range of <50, 50-100, or 100-200 nm and Andromed (control). Intracellular GSH was 4.8±0.14 nmol/10⁸ sperm pre-freezing, decreasing by 58% to 2.8±0.14 nmol/10⁸ sperm post-thawing in control (no differences among treatments). Sperm motility, viability, membrane functionality, apoptotic status, mitochondrial activity, lipid peroxidation, and DNA fragmentation were assessed after cryopreservation. Compared to the control, the 50-100 nm group showed significantly higher viability (72.7%±2.7 vs. 61.2%±2.7) and plasma membrane integrity (82.0%±1.6 vs. 73.3%±1.6). Conversely, the 100-200 nm group showed lower acrosomal integrity (P<0.05 vs. others) and increased DNA fragmentation (P<0.05 vs. 50-100 nm and control). Supplementing the lecithin-based freezing extender with 50-100 nm nanoparticles loaded with 2.5 mM GSH could enhance bull semen cryopreservation.