Sephadex filtration as successful alternative to density-gradient centrifugation procedures for ram sperm selection with improved kinetics

D.A Galarza,A López-Sebastián,H Woelders,E Blesbois,J Santiago-Moreno

doi:10.1016/j.anireprosci.2018.03.022

Abstract

Density-gradients centrifugation (DGC) and filtration columns (FC) are used to separate deformed or dead sperm, debris, and other cells that may negatively affect the fertilizing capacity of sperm in fresh, chilled and frozen/thawed semen. The present study was conducted to evaluate the suitability of DGC (BoviPure®, Percoll® and Accudenz®) and FC (Sephadex G-15®) sperm selection procedures for fresh-extended and cold-stored ram semen by assessment of post-treatment sperm quality variables. Twenty normospermic ejaculates from ten adult Merino rams were used. Sperm concentration of recovered cells was greater (P < 0.001) after BoviPure treatment than other procedures in both fresh and cold semen. With the Sephadex method, there were more desirable values than with use of DGC procedures in several sperm motility variables measured by using the CASA system. In non-refrigerated semen samples, the percentage of progressive sperm motility (%PSM) after Sephadex filtration was greater (P < 0.05) than after BoviPure treatment; the straightline velocity (VSL) value after Sephadex filtration was greater (P < 0.01) than after Accudenz treatment; the amplitude of lateral head displacement (ALH) after Sephadex and Accudenz treatment was less than non-filtered semen (P < 0.001) and after Percoll (P < 0.01) and BoviPure (P < 0.05) treatments. In cold-stored semen samples, the %PSM after Sephadex filtration was greater than non-filtered (P < 0.05) semen and after BoviPure (P < 0.05), Percoll (P < 0.05) and Accudenz (P < 0.001) treatments. It is concluded that Sephadex column filtration can be used to select ram sperm in non-refrigerated and cooled semen, because percentage progressively motile sperm and some other sperm motility characteristics are greater with use of this techniques as compared with use of DGC methods.

Highlights

Artificial insemination has an important role in sheep breeding but its use is limited because the poor fertility achieved when stored semen is used for vaginal insemination (Gil et al, 2003)
In non-refrigerated semen, the use of all sperm selection treatments appeared to reduce the percentage abnormalities and percentage of sperm cells with a damaged plasma membrane and/or acrosome membrane, the effects were small and not all pairwise comparisons with non-filtered sperm were significant, and the use of Accudenz had a negative effect on the % abnormal sperm tails
The percentage progressively motile sperm was only greater with use of Sephadex filtration and not with use of the Density-gradients centrifugation (DGC) methods

Summary

Introduction

Artificial insemination has an important role in sheep breeding but its use is limited because the poor fertility achieved when stored semen is used for vaginal insemination (Gil et al, 2003). The success of this procedure in sheep is limited by the anatomic characteristics of the ewe’s cervix and the short time that ram sperm can be stored as a liquid. The sperm characteristics and fertilizing capacity of extended ram semen may be improved by applying methods for sperm selection prior to artificial insemination. May have adverse "iatrogenic" effects related to mechanical damage due to centrifugation-resuspension procedures, and the promotion of lipid peroxidation during sperm pelleting due to the close vicinity of leukocytes and damaged sperm that may generate reactive oxygen species (ROS)(Sbracia et al, 1996)

Methods

Results

Conclusion