Kromatinstabilitet som grund för kvalitetsbedömning av hingstsperma

Abstract: This study is part of a project to evaluate and develop metods to control the semen quality in stallions. The study was carried out in cooperation between SLU and Flyinge AB. The objective of the study was to analyse -if there are individual differences in sperm chromatin integrety between different stallions -if there is a correlation between subjectively evaluated sperm motility and sperm chromatin integrity -if sperm chromatin integrity is changed after selection of the semen with a centrifugation method through the silica-spheres, gradient or a single-layer, i.e to use the chromatin integrity analysis as a method to evaluate the selection method. The use of chilled transported semen has increased a lot during the last 10 years, especially among the Swedish warmblood breed. Because the foaling percentage has decreased with the increased use of chilled transported semen, there is a need to develop methods to control the sperm quality. Evaluation of the chromatin integrity could be one way to evaluate the quality of the semen. Chromatin is the DNA in the nucleus of the sperm. If the chromatin is denatured it may lead to a failed fertilization or a disturbed development of the embryo or the placenta. Ten stallions of Swedish Warmblood breed at the Flyinge National Stud with the age 4-23 years were used in the study. The stallion semen was collected at seven times during a period of 2 ½ weeks. From every ejaculate samples were taken both before and after centrifugation through either of two different silica-spheres, single-layer or gradient for chromatin analysis and evaluation of sperm motility. The samples were immediatly frozen in TNE buffer and transported to Uppsala where they were analyzed with help of flow-cytometry. The results was presented as DFI (DNA Fragmentation Index) which measures the percentage of the sperms fragmentated DNA, and also the main DFI (X-DFI) and the standard deviation of DFI (SD-DFI). The result showed that the method of sperm centrifugation was effective in the selection of sperms with damage chromatin integrity. There was no difference in effectiveness between the two different metods of centrifugation, single–layer and gradient. The DNA Fragmentation Index (DFI) and the subjectively evaluated sperm motility was negative correlated. This means that the lower the DFI, the higher the motility. The result also showed that there was a statistical significant difference in DFI between stallions.