Effect of genistein on acrosome reaction and zona pellucida binding independent of protein tyrosine kinase inhibition in bull

AIM: To investigate if the phytoestrogen, genistein, affects essential functions of cryopreserved bovine spermatozoa. METHODS: The effect of genistein upon motility was assessed by computer-assisted motion analysis. Hemizona assay was performed to detect the ability of spermatozoa binding to the zona pellucida. The inducibility of the acrosome reaction using progesterone and ZP3-6 peptide was analysed by fluorescein-conjugated Pisum sativum agglutinin (FITC-PSA)/Hoechst 33258 double staining. Capacitation after incubation with genistein was assessed by the chlortetracycline (CTC) assay. Immunoblots showed the pattern of protein tyrosine phosphorylation of cryopreserved bovine spermatozoa. RESULTS: Immunodetection of tyrosine-phosphorylated proteins showed that genistein did not affect tyrosine phosphorylation in cryopreserved bovine spermatozoa. However, genistein significantly reduced the progesterone- and ZP3-6 peptide-mediated induction of the acrosome reaction and led to a dose-dependent inhibition of sperm-zona pellucida binding; while sperm motility and capacitation were not affected by this phytoestrogen, as indicated by computer-assisted sperm motion analysis and the CTC assay, respectively. CONCLUSION: Our results suggest that in cryopreserved bovine spermatozoa, genistein affects a protein tyrosine phosphorylation-independent signal transduction pathway that is involved in sperm capacitation, the acrosome reaction and sperm-zona pellucida binding.