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Anim Reprod Sci. 2017 Dec;187:167-173. doi: 10.1016/j.anireprosci.2017.11.002. Epub 2017 Nov 4.

Effect of Single Layer Centrifugation Porcicoll (70%, 80% and 90%) or supplementation with reduced glutathione, seminal plasma and bovine serum albumin on frozen-thawed boar sperm.

Author information

1
Institute of Animal Health and Cattle Development (INDEGSAL), University of León, 24071, León, Spain.
2
Institute of Biomedicine (IBIOMED), University of León, 24071, León, Spain.
3
Department of Clinical and Experimental Medicine (IKE). BKH, Obstetrics and Gynecology, Linköping University, 58183, Linköping, Sweden. Electronic address: manuel.alvarez-rodriguez@liu.se.
4
Department of Animal Medicine, Surgery and Anatomy (Animal Medicine and Surgery), University of León, 24071, León, Spain.
5
Division of Reproduction, Swedish University of Agricultural Sciences, 75007, Uppsala, Sweden.
6
Institute of Animal Health and Cattle Development (INDEGSAL), University of León, 24071, León, Spain; Dept. of Molecular Biology (Cell Biology), University of León, León, Spain.

Abstract

Selecting the optimal sperm population is essential for success with reproductive techniques. Porcicoll (formerly Androcoll-P) is a colloid formulation for selection of high-quality boar spermatozoa by single layer centrifugation (SLC). To date, most studies have been carried out with fresh semen and large volumes. We carried out 2 experiments to test the use of Porcicoll for thawed boar semen in small volumes. In Experiment 1, cryopreserved semen doses were thawed, split in 200-μL aliquots and layered on 1mL of Porcicoll 70%, 80% or 90%, or buffer without colloid. We assessed sperm recovery (the proportion of the loading dose that appeared in the pellet, %), and the physiology of the selected spermatozoa (flow cytometry: Viability, apoptotic changes, capacitation, mitochondrial activity, intracellular reactive oxygen species). The most suitable proportion was Porcicoll 80%, allowing acceptable sperm recovery (16.9±4.2%, compared to 70% (35.4%±3.0, p<0.001) and 90% (8.2%±3.0, P=0.001), and improved quality (mitochondrial activity: Porcicoll 80%: 77.7±1% vs Control: 60.3±0.7%, P<0.05). In Experiment 2, we compared 3 supplements to Porcicoll 80%: 500mM reduced glutathione (GSH), 20% seminal plasma (SP) and 0.5% bovine serum albumin (BSA). Supplementation with GSH or BSA did not cause relevant changes relative to Control. In contrast, SP induced membrane and acrosomal changes resembling capacitation, which might preclude its use in some applications, and decreased recovery (5.5%±1.9 vs. 24.3%±1.2 Control; P<0.001). However, it could be useful prior to other applications such as in vitro fertilisation. Overall, Porcicoll is an effective colloid for isolating a high-quality population from thawed boar sperm, 80% being a balanced option for good recovery and high quality. Supplements could be useful depending on the proposed use of the spermatozoa.

KEYWORDS:

Bovine serum albumin; Pig; Reduced glutathione; Semen cryopreservation; Seminal plasma; Single-layer centrifugation

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