Canine spermatozoa DNA damage in a milk based cooling extender: a preliminary report.

VALIENTE C; DIAZ JD; PICCO S; CORRADA Y; ABEYA M; GOBELLO C.

Buenos Aires

Jornada; II Jornadas Internacionales del Instituto de Investigación y Tecnología en Reproducción Animal; 2010

Fac. Cs. Veterinarias. UBA

Skimmed milk is a low-cost, user- friendly extender for cooling canine semen (Valiente et al., 2010). Semen chilling facilitates destabilization in the chromatin structure of spermatozoa which is associated with poor pregnancy outcome. The objective of this study was to investigate the effect of cooling (4 C), in a milk-based extender, on canine spermatozoa DNA. The relationship between motility and DNA damage was also described thoughout the study period. The second fraction of ejaculates was collected by manual stimulation from 6 male dogs at weekly intervals during a month. The fractions were pooled, diluted to 75-100 106 sperm/ml in non-fat dry skimmed milk [Ilolay®, Argentina] with antibiotics [benzyl penicillin 1mg/ml and dihydrostreptomycin sulphate 1 mg/ml] and then cooled and stored at 4º C during 4 days. Percentages of motility and DNA integrity (modified Comet assay; Singh et al., 1988) were assessed at four time points: immediately after dilution and then 24, 60 and 96 hours later. Data were expressed as mean±SEM and cell damage and motility were correlated by the Spearman test. Percentages of DNA damaged spermatozoa in the four progressive observations were 2.2±0.3, 5.2±3.3, 7.4±2.2 and 10.8±0.3. Percentage motility decreased from 91.8±0.9 to 53.7±4.4 throughout the study period. Correlation between DNA damage and motility was r -0.97 (P < 0.05). To conclude, canine spermatozoa DNA damage during storage at 4º C in a milk-based extender seems to be mild and slowly progressive. Further work using this extender is warranted.