Effect of oviductal fluid proteins on buffalo sperm characteristics during cryopreservation.
The objective was to determine the effects of oviductal proteins on sperm function. Abbatoir-derived buffalo oviducts were flushed with PBS; the fluid recovered (protein concentration, 2.3 mg/mL; average of 3.5 mg protein/oviduct) was centrifuged, dialyzed, and clarified, and the supernatant applied to a Heparin-Sepharose affinity column. Unbound fractions were collected and bound proteins were separately eluted (with elution buffer). Eight distinct protein bands (from 12 to 177 kDa) in the H-unbound fraction and 15 distinct protein bands (from 12 to 165 kDa) in the H-bound fraction were detected in SDS-PAGE. Semen from four buffalo bulls was divided into three parts: Parts 1 and 2 were treated with the heparin binding (H-bound) and non-heparin binding (H-unbound) oviductal proteins, respectively, whereas Part 3 remained as an untreated control. Equilibrated and frozen-thawed semen was assessed for motility, viability, intact acrosome percentage, mucus penetration distance, and hypo-osmotic swelling test. The H-bound oviductal fluid proteins enhanced (P<0.05) the proportion of sperm that were progressively motile, alive, had an intact acrosome and functional plasma membrane (hypo-osmotic swelling test), as well as the distance covered in the cervical mucus sperm penetration test during cryopreservation. Addition of the H-unbound oviductal protein fraction did not increase sperm motility and penetration distance but increased (P<0.05) the proportion of sperm that were live, had an intact acrosome, and functional plasma membrane (hypo-osmotic swelling test). We concluded that the H-bound fraction of buffalo oviductal fluid protein(s) maintained sperm motility, viability and membrane integrity during cryopreservation, whereas the H-unbound proteins maintained sperm viability and membrane integrity.
Imam
S
S
Artificial Insemination Laboratory, Division of Animal Reproduction, Indian Veterinary Research Institute, Izatnagar 243 122, U.P., India.
Ansari
M R
MR
Ahmed
N
N
Kumaresan
A
A
D000177:Acrosome / Q000187:drug effects / Q000502:physiology; D000818:Animals; D002020:Buffaloes; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002582:Cervix Mucus / Q000502:physiology; D002846:Chromatography, Affinity / Q000662:veterinary; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D005187:Fallopian Tubes / Q000737:chemistry*; D005260:Female; D006493:Heparin / Q000737:chemistry / Q000494:pharmacology; D008297:Male; D009995:Osmosis / Q000187:drug effects; D011506:Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility / Q000187:drug effects / Q000502:physiology; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology
D011506:Proteins; D006493:Heparin