title: Purification of binder of sperm protein 1 (BSP1) and its effects on bovine in vitro embryo development after fertilization with ejaculated and epididymal sperm.
abstract: The present study evaluated functional aspects of binder of sperm 1 (BSP1) in the bovine species. In a first experiment, cumulus-oocyte complexes (n = 1274) were incubated with frozen-thawed ejaculated sperm (18 hours) in Fert-TALP medium containing: heparin, 10, 20, or 40 μg/mL BSP1. Heparin followed by gelatin affinity chromatography was used for purification of BSP1 from bovine seminal vesicle fluid. With ejaculated sperm, cleavage rates were similar when Fert-TALP medium was incubated with heparin (74.1 ± 2.7%), 10 μg/mL BSP1 (77.8 ± 3.1%), or 20 μg/mL BSP1 (74 ± 2.0%). Day-7 blastocyst rates were equivalent after incubations with heparin (40.8 ± 5.0%) and 10 μg/mL BSP1 (34.1 ± 4.4%), but reduced after 20 μg/mL BSP1 (22.4 ± 2.9%) and 40 μg/mL BSP1 (19.3 ± 4.1%; P < 0.05). In the second experiment, cumulus-oocyte complexes (n = 1213) were incubated with frozen-thawed cauda epididymal sperm (18 hours) in Fert-TALP medium containing: no heparin, heparin, 10, 20, or 40 μg/mL. Cleavage and blastocyst rates were similar after treatments with heparin (68.5 ± 1.3% and 24.7 ± 3.2%, respectively) or without heparin (65.5 ± 1.8% and 27.3 ± 1.6%, respectively). Cleavage was higher after treatment with any BSP1 concentrations (74.2 ± 2.7%-79.0 ± 1.1%) than without heparin (P < 0.05). Also, cleavage was better after Fert-TALP medium incubation with 40 μg/mL BSP1 (79.0 ± 1.1%) than with heparin (68.5 ± 1.3%; P < 0.05). Embryo development was higher (P < 0.05) after treatment with 20 μg/mL BSP1 (35.6 ± 2.5%) and 40 μg/mL (41.1 ± 2%) than after incubations with heparin (24.7 ± 3.2%) or without heparin (27.3 ± 1.6%). Interestingly, BSP1 did not cause reductions in blastocyst rates after fertilization with epididymal sperm, as observed with ejaculated sperm. On the basis of immunocytochemistry, there was BSP1 binding to frozen-thawed ejaculated but not to epididymal sperm. Also, anti-BSP1 reaction remained on ejaculated sperm (as expected) and appeared on epididymal sperm after incubation with purified BSP1. Acrosome reaction of ejaculated and epididymal sperm was induced after incubation with purified BSP1 as well, indicating an effect of BSP1 on capacitation. In conclusion, purified BSP1 from bull seminal vesicles was able to bind to and induce capacitation of ejaculated and epididymal sperm. Also, BSP1 added to fertilization media and allowed proper cleavage and embryo development, with the effects being modulated by previous exposure or not of spermatozoa to seminal plasma.
authors.affiliation: Animal Physiology Laboratory, Department of Animal Science, Federal University of Ceara, Fortaleza, Brazil.
authors.forename: P
authors.initials: P
authors.lastname: Rodríguez-Villamil
authors.affiliation: Animal Physiology Laboratory, Department of Animal Science, Federal University of Ceara, Fortaleza, Brazil.
authors.forename: V
authors.initials: V
authors.lastname: Hoyos-Marulanda
authors.affiliation: Animal Physiology Laboratory, Department of Animal Science, Federal University of Ceara, Fortaleza, Brazil.
authors.forename: J A M
authors.initials: JA
authors.lastname: Martins
authors.affiliation: Animal Physiology Laboratory, Department of Animal Science, Federal University of Ceara, Fortaleza, Brazil.
authors.forename: A N
authors.initials: AN
authors.lastname: Oliveira
authors.affiliation: Molecular and Developmental Biology Laboratory, University of Fortaleza, Fortaleza, Brazil.
authors.forename: L H
authors.initials: LH
authors.lastname: Aguiar
authors.affiliation: School of Pharmacy, University of Fortaleza, Fortaleza, Brazil.
authors.forename: F B
authors.initials: FB
authors.lastname: Moreno
authors.affiliation: Animal Physiology Laboratory, Department of Animal Science, Federal University of Ceara, Fortaleza, Brazil.
authors.forename: A L M C S
authors.initials: AL
authors.lastname: Velho
authors.affiliation: School of Pharmacy, University of Fortaleza, Fortaleza, Brazil.
authors.forename: A C
authors.initials: AC
authors.lastname: Monteiro-Moreira
authors.affiliation: School of Pharmacy, University of Fortaleza, Fortaleza, Brazil.
authors.forename: R A
authors.initials: RA
authors.lastname: Moreira
authors.affiliation: Department of Biochemistry and Molecular Biology, Federal University of Ceara, Fortaleza, Brazil.
authors.forename: I M
authors.initials: IM
authors.lastname: Vasconcelos
authors.affiliation: Molecular and Developmental Biology Laboratory, University of Fortaleza, Fortaleza, Brazil.
authors.forename: M
authors.initials: M
authors.lastname: Bertolini
authors.affiliation: Animal Physiology Laboratory, Department of Animal Science, Federal University of Ceara, Fortaleza, Brazil. Electronic address: amoura@ufc.br.
authors.forename: A A
authors.initials: AA
authors.lastname: Moura
mesh_terms: D020101:Acrosome Reaction / Q000187:drug effects; D000818:Animals; D001755:Blastocyst / Q000502:physiology; D002417:Cattle; D015925:Cryopreservation / Q000662:veterinary; D003470:Culture Media; D054885:Cumulus Cells / Q000502:physiology; D004542:Ejaculation; D047108:Embryonic Development / Q000187:drug effects*; D004822:Epididymis / Q000166:cytology; D005260:Female; D005307:Fertilization in Vitro / Q000379:methods / Q000662:veterinary*; D006493:Heparin / Q000494:pharmacology; D059471:In Vitro Oocyte Maturation Techniques / Q000662:veterinary; D008297:Male; D009865:Oocytes / Q000502:physiology; D012662:Semen Preservation / Q000662:veterinary; D029607:Seminal Vesicle Secretory Proteins / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology*; D013075:Sperm Capacitation / Q000187:drug effects; D013094:Spermatozoa / Q000378:metabolism / Q000502:physiology*
chemical_list: C519889:BSPH1 protein, human; D003470:Culture Media; D029607:Seminal Vesicle Secretory Proteins; D006493:Heparin
keywords: BSP1; Embryo; Fertilization; Seminal plasma; Sperm