title: Desalted and lyophilized bovine seminal plasma delays induction of the acrosome reaction in frozen-thawed bovine spermatozoa in response to calcium ionophore.
abstract: Cryopreservation is partially damaging and induces capacitation-like changes in spermatozoa. Seminal plasma (SP) contains a variety of biochemical components, such as protein and lipids, which are specific for the regulation of sperm cell function including those effective for decapacitation of spermatozoa. Therefore, this study tested the hypothesis that desalted and lyophilized SP could prevent premature capacitation (cryocapacitation) of Japanese Black bull spermatozoa. Seminal plasma was desalted by using Sephadex G-25 desalting column and lyophilized before added to semen extender at final concentrations 0, 2.5, 12.5, and 25 mg/mL. Frozen-thawed sperm progressive motility, acrosomal integrity, abnormal morphology, and the calcium ionophore A23187-induced acrosome reaction were assessed. Protein and lipid compositions in SP were analyzed by SDS-PAGE and thin-layer chromatography, respectively. The results revealed that progressive motility, intact acrosome, and abnormal morphology were not substantially modified by addition of SP. Stimulation of spermatozoa with calcium ionophore A23187 resulted in a time-dependent induction of the acrosome reaction, which was delayed by the desalted and lyophilized SP. There was no difference in the protein profile of SP before and after gel filtration. In total, 19 protein bands with molecular masses ranging from 5.2 to 185.8 kDa were detected and those of 185.8, 80, 34, 20.8, 18.8, 17.5, and 10 kDa were considered as novel proteins. Neutral lipids and phospholipids before and after gel filtration were the same, and the detected neutral lipid spots were monoacylglycerol, cholesterol, 1,2- and 1,3-disaturated diacylglycerol, 1,2- and 1,3-saturated, unsaturated diacylglycerol, whereas the detected phospholipid spots were sphingomyelin, phosphatidylcholine, phosphatidylserine, and three species of phosphatidylinositol, phosphatidylethanolamine, cerebroside, and polyglycerol phosphatide. The results suggest that premature capacitation during freeze-thaw processes could be reduced by adding desalted and lyophilized SP.
authors.affiliation: Laboratory of Theriogenology, Faculty of Applied Biological Sciences, Department of Veterinary Medicine, Gifu University, Gifu, Japan; Faculty of Veterinary Medicine, Department of Theriogenology, Kafrelsheikh University, Kafrelsheikh, Egypt.
authors.forename: Essam
authors.initials: E
authors.lastname: Almadaly
authors.affiliation: Hida Beef Cattle Research Department, Gifu Prefectural Livestock Research Institute, Takayama, Japan.
authors.forename: Youichirou
authors.initials: Y
authors.lastname: Hoshino
authors.affiliation: Hida Beef Cattle Research Department, Gifu Prefectural Livestock Research Institute, Takayama, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Ueta
authors.affiliation: Hida Beef Cattle Research Department, Gifu Prefectural Livestock Research Institute, Takayama, Japan.
authors.forename: Koushi
authors.initials: K
authors.lastname: Mukoujima
authors.affiliation: Faculty of Veterinary Medicine, Kafrelsheikh University, Kafrelsheikh, Egypt.
authors.forename: Mostafa
authors.initials: M
authors.lastname: Shukry
authors.affiliation: Faculty of Veterinary Medicine, Kafrelsheikh University, Kafrelsheikh, Egypt.
authors.forename: Foad
authors.initials: F
authors.lastname: Farrag
authors.affiliation: Faculty of Veterinary Medicine, Department of Theriogenology, Kafrelsheikh University, Kafrelsheikh, Egypt.
authors.forename: Ismail
authors.initials: I
authors.lastname: El-Kon
authors.affiliation: Hida Beef Cattle Research Department, Gifu Prefectural Livestock Research Institute, Takayama, Japan.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Kita
authors.affiliation: Laboratory of Theriogenology, Faculty of Applied Biological Sciences, Department of Veterinary Medicine, Gifu University, Gifu, Japan. Electronic address: muraset@gifu-u.ac.jp.
authors.forename: Tetsuma
authors.initials: T
authors.lastname: Murase
mesh_terms: D020101:Acrosome Reaction / Q000187:drug effects*; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D061207:Calcium Ionophores / Q000494:pharmacology*; D002417:Cattle / Q000502:physiology*; D002850:Chromatography, Gel; D015925:Cryopreservation / Q000662:veterinary; D003911:Dextrans; D005612:Freeze Drying / Q000662:veterinary; D006358:Hot Temperature; D008055:Lipids / Q000032:analysis; D008297:Male; D010743:Phospholipids / Q000032:analysis; D012661:Semen / Q000737:chemistry* / Q000502:physiology*; D012662:Semen Preservation / Q000662:veterinary; D029589:Seminal Plasma Proteins / Q000032:analysis; D013048:Specimen Handling / Q000379:methods / Q000662:veterinary*; D013075:Sperm Capacitation / Q000502:physiology; D013081:Sperm Motility
chemical_list: D061207:Calcium Ionophores; D003911:Dextrans; D008055:Lipids; D010743:Phospholipids; D029589:Seminal Plasma Proteins; D000001:Calcimycin; C025614:sephadex; D002118:Calcium
keywords: Acrosome reaction; Desalted and lyophilized; HPTLC; SDS-PAGE; Seminal plasma; TLC