Comparative study of the biological activity of spermatozoa–zona pellucida binding inhibitory factors from human follicular fluid on various sperm function parameters*

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Abstract
Previous studies showed that human follicular fluid (hFF) from gonadotrophin stimulated cycles contained two glycoproteins, named as ZIF‐1 and ZIF‐2, that reduced the zona binding capacity of spermatozoa. The present study showed that the spermatozoa–zona pellucida binding inhibitory activity was also present in hFF from natural cycle. Using the hemizona binding assay, the inhibitory effect of ZIF‐1 on the zona binding capacity of spermatozoa was dose‐dependent. The effect of ZIF‐2 was also dose‐dependent, in the range of 10–100 ng/ml. The inhibitory effects of both ZIF‐1 and ‐2 increased with the duration of the spermatozoa–ZIF interaction. The effect of the former was present up to 120 min incubation, whilst that of latter occurred for the first 90 min. The zona binding inhibitory effect of ZIF‐1 and ‐2 was additive when they were used together to treat the spermatozoa. The biological activity of ZIFs on other sperm parameters that might affect spermatozoa–zona pellucida binding was also investigated. ZIF‐1 did not affect the acrosomal status of human spermatozoa while ZIF‐2 significantly increased the number of acrosome reacted spermatozoa in the range of 0.1–10 μg. However, the increase in the incidence of acrosome‐reacted spermatozoa after ZIF‐2 treatment could not totally account the inhibitory effect of ZIF‐2 on zona binding. Both glycoproteins did not affect the motility of human spermatozoa. Radioactively‐labelled ZIFs bound to human spermatozoa. Unlabelled ZIF displaced the bound radioactivity of spermatozoa treated with the corresponding labelled ZIF. These suggested the presence of specific binding sites of ZIFs on human spermatozoa. Mol. Reprod. Dev. 61: 205–212, 2002.