Preservation of boar semen at 18 °C induces lipid peroxidation and apoptosis like changes in spermatozoa

Boar sperm functions, lipid peroxidation status, mitochondrial membrane potential (ΔΨm) and membrane permeability (apoptosis like features) were assessed during liquid preservation. Four ejaculates each from four Hampshire boars were extended with Beltsville Thawing Solution and preserved at 18 °C. At 0, 24, 48, 72 and 96 h of storage, each ejaculate was examined for sperm functions, lipid peroxidation, ΔΨm, and membrane permeability. The lipid peroxidation status of the sperm was assessed based on the malonaldehyde (MDA) levels. Detection of ΔΨm was done using 3,3′-dihexyloxacarbocyanine iodide [DiOC6(3)]/propidium iodide (PI) assay and Yo-pro-1/PI assay was used to detect change in plasma membrane permeability. The sperm motility, viability and acrosomal integrity declined significantly (p < 0.05) from 0 to 96 h of preservation. At the start of the preservation, the MDA levels (nM/109 sperm) were low in sperm (99.83 ± 2.69) and seminal plasma (191.98 ± 11.58), which gradually increased up to the 96 h of storage. Highest negative correlation (r value) was observed between MDA levels and sperm motility (−0.97), live percent (−0.97), acrosomal integrity (−0.97) and hypo-osmotic sperm swelling test (HOSST) positive sperm percentage (−0.98). Strong positive correlation was observed between HOSST positive sperm percentage and intact acrosome percentage (r = 0.98). There was a significant (p < 0.05) increase in the sperm cells with low ΔΨm from 0 to 96 h of preservation. Before preservation, 14.85 ± 4.66% of sperm cells of the ejaculate showed low mitochondrial membrane potential, whereas after 96 h of preservation, this proposition of cells increased up to 32.00 ± 6.25%. The apoptotic sperm population was 8.33 ± 2.31% in fresh semen, while this population was 25.19 ± 4.25% at 96 h of preservation and the difference was significant (p < 0.05). The findings of the present study revealed that liquid preservation of boar semen at 18 °C induces lipid peroxidation, decrease mitochondrial membrane potential and increase the plasma membrane permeability.