Spermatozoa generate small amounts of O2°- and NO°. Under physiological condition these compounds exist at very low concentration and the formation of peroxynitrite (ONOO−) appears likely. Peroxynitrite reacts rapidly with proteins, lipids and DNA. Moreover, tyrosine nitration is a widely used marker of peroxynitrite. The nitration of protein residues gives rise to 3-nitrotyrosine which represents a protein modification specific for ONOO− formation in vivo. We determined NO and ONOO− production in semen and their potential role with sperm motility in infertile men, and we set out to determine whether protein tyrosine nitration takes place in sperm cells motility. Semen samples from 25 normal fertile donors and 40 infertile patients were analyzed. NO was measured by Griess Reaction while peroxynitrite concentration through the fluorescence of the DCFDA probe. Protein tyrosine nitration was determined by Western Immuno Blot as well as the presence of eNOS and iNOS. The controls exhibited both NO and ONOO− productions that was significantly lower than asthenozoospermic patients and were inverse correlated with the motility parameters. Moreover, the western immuno blots showed an increase in the nitration of the tyrosine residues in the asthenozoospermic samples compared to controls and the presence of both iNOS and eNOS proteins. The same pattern was confirmed by means of immunohistochemistry. The present data suggest a critical negative effect of NO and peroxynitrite on sperm motility when spermatozoa concentration is normal. Thus, a possible pathogenic role in infertile men when asthenozoospermia is the main critical problem may be suggested.
Declaration of interest: The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project.
Funding: This research did not receive any specific grant from any funding agency in the public, commercial or not-for-profit sector.
05 - 09 May 2012
European Society of Endocrinology