Correlation between tyrosine phosphorylation intensity of a 107 kDa protein band and A23187-induced acrosome reaction in human spermatozoa.
This study, performed using semen samples from 10 men, investigated the relationship between sperm protein tyrosine phosphorylation and acrosomal status in conditions supporting in vitro capacitation. Percoll-selected spermatozoa (cells from the 95% fraction) were incubated for 3 h at 37 degrees C under an atmosphere of 5% CO2 in air, in a polyvinyl alcohol (1 mg ml(-1)) containing Biggers-Whitten-Whittingham's medium, nonsupplemented or supplemented with either bovine serum albumin (BSA; fatty acid free, 3 mg ml(-1)) or 2-hydroxy-propyl-beta-cyclodextrin (2-OH-p-beta-CD; 0.5, 1, 2 mmol l(-1)). Sperm suspension in each medium was split into two aliquots. The first was used to evaluate the acrosomal status by staining with the fluorescein isothiocyanate Pisum sativum agglutinin after induction of the acrosome reaction (AR) for 45 min with 10 micromol l(-1) of A23187 calcium ionophore. The second aliquot was used for sodium dodecyl sulphate polyacrylamide gel electrophoresis and immunoblotting, followed by a densitometric analysis. Compared with the nonsupplemented medium, BSA- or 2-OH-p-beta-CD-supplementation induced an increase in both the percentage of live acrosome-reacted sperm and the tyrosine phosphorylation intensity of the main phosphorylated 107 kDa protein. A correlation between the percentage of live acrosome-reacted sperm and the 107-kDa protein phosphotyrosine intensity was observed. Therefore, the 107 kDa protein-phosphotyrosine level measurement would bring additional information to conventional semen parameters in the assessment of the human sperm functionality.
Picherit-Marchenay
C
C
Laboratoire de Biologie de la Reproduction-CECOS, Hôtel-Dieu, Clermont-Ferrand, France. christel.picherit@laposte.net
Bréchard
S
S
Boucher
D
D
Grizard
G
G
D000073738:2-Hydroxypropyl-beta-cyclodextrin; D020101:Acrosome Reaction / Q000187:drug effects; D000001:Calcimycin / Q000494:pharmacology*; D006801:Humans; D007476:Ionophores / Q000494:pharmacology*; D008297:Male; D008970:Molecular Weight; D010766:Phosphorylation / Q000187:drug effects; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D013094:Spermatozoa / Q000187:drug effects* / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D047392:beta-Cyclodextrins / Q000494:pharmacology
D007476:Ionophores; D011506:Proteins; D047392:beta-Cyclodextrins; D000073738:2-Hydroxypropyl-beta-cyclodextrin; D012710:Serum Albumin, Bovine; D000001:Calcimycin; D014443:Tyrosine