title: The activity of N-acetyl-β-hexosaminidase in boar seminal plasma is linked with semen quality and its suitability for cryopreservation.
abstract: The determination of sperm cryotolerance is an important step in the process of developing optimal techniques for the storage of boar semen. The objective of this study was to determine individual proteome variations in boar seminal plasma and spermatozoa and establish their influence on the cryotolerance of ejaculate. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed the presence of protein with estimated molecular weight of 90 kDa in sperm extracts from ejaculates of selected boars. In all cases, dialysis performed at the initial stage of cryopreservation effectively removed the protein from sperm cells. The protein had an affinity for Zn(2+) ions. Mass spectrometry revealed similarities between the discussed protein and the β subunit of N-acetyl-β-hexosaminidase (β-HEX). Seminal plasma β-HEX was purified 252-fold with approximately 27% recovery and specific activity of 1800 U/mg of protein. Enzyme activity in fresh seminal plasma was correlated with superoxide dismutase activity (r = -0.42, P < 0.05), glutathione peroxidase activity (r = -0.42, P < 0.05), mitochondrial function (r = 0.31, P < 0.05), glutathione content (r = 0.34, P < 0.05), total protein content (r = 0.42, P < 0.05), and total oxidant status of seminal plasma (r = 0.37, P < 0.05). After thawing, β-HEX activity in seminal plasma was negatively correlated with the total motile sperm count (r = -0.33, P < 0.05), plasma membrane integrity (r = -0.31, P < 0.05), and lipid peroxidation (r = 0.33, P < 0.05). The observed correlations indicate that lower levels of β-HEX activity in boar seminal plasma are linked with higher quality of sperm after thawing. Based on those observations, the ejaculates were divided into two groups characterized by low (<20,000 U/L) and high (>20,000 U/L) levels of β-HEX activity in seminal plasma. In plasma with high β-HEX activity, spermatozoa were characterized by lower plasma membrane integrity (84.7%, P < 0.05). Higher glutathione levels (1250.3 μM), higher total protein content (50 mg/mL), and higher total oxidant status (6.82-μmol H2O2 Equiv/L) were also observed (P < 0.05). After thawing, lower sperm motility (20.4%), lower plasma membrane integrity (41.7%), and higher lipid peroxidation (30.9-nM malondialdehyde/10(8) spermatozoa/h) were reported in ejaculates with high seminal plasma β-HEX activity. The results of this study indicate that β-HEX activity in seminal plasma is a useful indicator in preliminary evaluations of boar sperm cryotolerance.
authors: Department of Animal Biochemistry and Biotechnology, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland. Electronic address: pawel.wysocki@uwm.edu.pl.
authors: Paweł
authors: P
authors: Wysocki
authors: Department of Animal Biochemistry and Biotechnology, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland.
authors: Aleksandra
authors: A
authors: Orzołek
authors: Department of Animal Biochemistry and Biotechnology, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland.
authors: Jerzy
authors: J
authors: Strzeżek
authors: Department of Animal Biochemistry and Biotechnology, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland.
authors: Magdalena
authors: M
authors: Koziorowska-Gilun
authors: Department of Animal Biochemistry and Biotechnology, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland.
authors: Łukasz
authors: Ł
authors: Zasiadczyk
authors: Department of Animal Biochemistry and Biotechnology, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland.
authors: Władysław
authors: W
authors: Kordan
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000032:analysis; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D005978:Glutathione / Q000032:analysis; D015227:Lipid Peroxidation; D008297:Male; D011506:Proteins / Q000032:analysis; D012661:Semen / Q000201:enzymology*; D055101:Semen Analysis / Q000662:veterinary; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013076:Sperm Count; D013081:Sperm Motility; D013094:Spermatozoa / Q000201:enzymology*; D034421:Sus scrofa; D001619:beta-N-Acetylhexosaminidases / Q000032:analysis* / Q000378:metabolism
chemical_list: D000975:Antioxidants; D011506:Proteins; D001619:beta-N-Acetylhexosaminidases; D005978:Glutathione
keywords: Boar; Cryopreservation; Proteomics; Sperm cryotolerance