Endogenous reactive oxygen species content and modulation of tyrosine phosphorylation during sperm capacitation.
Generation of controlled amounts of reactive oxygen species (ROS) and phosphorylation of protein tyrosine (Tyr) residues are two main cellular changes involved in sperm capacitation. This study examined the relationship between tyrosine-phosphorylation (Tyr-P) and endogenous ROS production during sperm capacitation, and correlated them with both sperm motility and functionality expressed as acrosome-reacted cells. Immediate ROS generation was observed to peak after a 45-min incubation, followed by a rapid decrease in ROS content and successive regeneration of the ROS peak in 3 h and later. These two peaks were directly correlated with both the Tyr-P process involving sperm heads and tails, and the acrosome reaction (69 ± 8% and 65 ± 4%, respectively). The period of low-ROS content resulted in low Tyr-P patterns, located exclusively in the cell midpiece, and drastic reduction in acrosome-reacted cells. Ascorbic acid addition inhibited both Tyr-P patterns and acrosome reactions, whereas NADPH induced high ROS generation, with Tyr-P patterns located only on sperm tails, and prevented the acrosome reaction. Sperm hyperactivation was insensitive to ROS content. This is an important parameter for evaluation of sperm capacitation, which is achieved only when both ROS generation reaches a peak and Tyr-P involves the sperm head.
Donà
G
G
Department of Biological Chemistry Endocrinology Unit, University of Padova, Padova, Italy.
Fiore
C
C
Tibaldi
E
E
Frezzato
F
F
Andrisani
A
A
Ambrosini
G
G
Fiorentin
D
D
Armanini
D
D
Bordin
L
L
Clari
G
G
D000328:Adult; D015153:Blotting, Western; D006801:Humans; D007150:Immunohistochemistry; D049449:Luminescence; D008297:Male; D009249:NADP / Q000378:metabolism; D010766:Phosphorylation; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013075:Sperm Capacitation; D014443:Tyrosine / Q000378:metabolism*
D017382:Reactive Oxygen Species; D014443:Tyrosine; D009249:NADP