The goal of this study was to examine the functionality of the sperm isolated from mice subjected to resveratrol exposure in vivo. Sperm are not fully functional until after capacitation has taken place. Using a proximal ligation assay we recently identified a complex involving Protein-Kinase A (PKA) as being associated with the capacitation event. The complex contains a novel protein which undergoes tyrosine phosphorylation upon capacitation. TCP-11 is also part of the complex and becomes associated with PKA upon capacitation.
Mice were injected intraperitoneally with resveratrol (10mg/kg body weight) on a weekly basis for one month and then examined two or five months after injection. Freshly isolated sperm were immediately fixed or incubated for three hours in capacitation media prior to fixing. Cells were probed with an anti-phosphotyrosine antibody. The staining apparent after three hours capacitation of the sperm from untreated mice was similar in intensity to that seen immediately upon isolation in mice exposed to resveratrol in vivo.
Sperm proteins were isolated and subjected to Western blot analysis. Tyrosine phosphorylation occurred immediately and on different proteins to those seen after incubation with capacitation media.
We also found the reorganisation of the complex containing PKA occurred prematurely. In addition our novel protein was tyrosine phosphorylated. This indicates that the sperm from exposed mice are already capacitated upon collection.
These findings suggest that in vivo exposure to resveratrol leads to premature sperm capacitation. As well as being an antioxidant, resveratrol has also been found to be an inhibitor of a number of enzymes including phosphodiesterase. It is noteworthy that resveratrol can’t induce premature capacitation in vitro. This suggests the effect of resveratrol is not a direct effect on the level of sperm reactive oxygen species or PKA activation. Rather the effect is mediated by other mechanisms in vivo.