title: Kinetic characterization of the changes in protein tyrosine phosphorylation of membranes, cytosolic Ca2+ concentration and viability in boar sperm populations selected by binding to oviductal epithelial cells.
abstract: On reaching the oviduct, spermatozoa are retained in the isthmic region of the oviduct until ovulation occurs. The essential steps of capacitation are co-ordinated in this region. In this study, a primary cell culture system of oviductal epithelial cells was established to investigate sperm binding to oviductal epithelium and modulation of sperm function during incubation under capacitating conditions in co-culture with oviductal epithelial cells. Epithelial cells were stripped from the oviducts of sows and cultivated for 5-7 days on Lab-Tek Chamber slides on Matrigel. The preparations on chamber slides and suspensions of control spermatozoa were incubated for 3 h in Tyrode's albumin lactate pyruvate (TALP) medium. At 3, 30, 60, 90 and 180 min the free-swimming spermatozoa were collected by washing, and membrane integrity, tyrosine phosphorylation patterns and [Ca(2+)](i) of bound, unbound and control spermatozoa were assessed with fluorescent probes (propidium iodide, Cy-3 and fluo-3-AM). The cells bound to oviductal epithelial cells showed reduced cytosolic Ca(2+) concentration, reduced and almost absent tyrosine phosphorylation of membrane proteins and higher viability at the time of the first sampling. Increases in Ca(2+) concentration and cell death occurred much more slowly during incubation in cells bound to oviductal epithelial cells compared with free-swimming spermatozoa, and no changes in tyrosine phosphorylation were observed. The preferential binding of viable, low-Ca(2+) cells with suppressed tyrosine phosphorylation and slower functional modulation of boar spermatozoa attached to oviductal epithelial cells might represent a mechanism for selecting functionally competent spermatozoa and prolonging their lifespan by delaying capacitation in the oviductal reservoir.
authors.affiliation: Institute for Reproductive Medicine, School of Veterinary Medicine of Hannover, Bünteweg 15, 30559 Hannover, Germany.
authors.forename: A M
authors.initials: AM
authors.lastname: Petrunkina
authors.forename: J
authors.initials: J
authors.lastname: Friedrich
authors.forename: W
authors.initials: W
authors.lastname: Drommer
authors.forename: G
authors.initials: G
authors.lastname: Bicker
authors.forename: D
authors.initials: D
authors.lastname: Waberski
authors.forename: E
authors.initials: E
authors.lastname: Töpfer-Petersen
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D018929:Cell Culture Techniques; D002462:Cell Membrane / Q000648:ultrastructure; D002470:Cell Survival; D018920:Coculture Techniques; D003600:Cytosol / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism; D005187:Fallopian Tubes / Q000378:metabolism*; D005260:Female; D005456:Fluorescent Dyes; D007150:Immunohistochemistry; D007700:Kinetics; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D008854:Microscopy, Electron; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D013075:Sperm Capacitation; D013094:Spermatozoa / Q000502:physiology* / Q000648:ultrastructure; D013552:Swine
chemical_list: D005456:Fluorescent Dyes; D008565:Membrane Proteins; D019000:Phosphotyrosine; D002118:Calcium