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Acrosome Reaction Induced by Recombinant Human Zona Pellucida 3 Peptides rhuZP3a²²¹⁷⁶ and rhuZP3b¹⁷⁷³⁴⁸ and Their Mechanism

KUN LI^*,,¶

WANXIANG XU

LIWEN SONG

KANGSHOU YAO

XINZONG ZHANG

From the ^* Department of Reproductive Physiology, Zhejiang Academy of Medical Sciences, Hangzhou, China; Department of Laboratory Medicine, Wenzhou Medical College, Wenzhou, China; Shanghai Institute of Planned Parenthood Research, Shanghai, China; Zhejiang Provincial Institute of Planned Parenthood Research, Hangzhou, China; and the ^|| Women's Hospital, School of Medicine, Zhejiang University, Hangzhou, China.

Correspondence to: Qi-xian Shi, Department of Reproductive Physiology, Zhejiang Academy of Medical Sciences, 182 Tian Mu Shan Road, Hangzhou, Zhejiang 310013, China (e-mail: qxshi{at}mail.hz.zj.cn).

To explore the biological characteristics of the recombinant zona pellucida 3 (ZP3) peptides of rhuZP3a²²¹⁷⁶ and rhuZP3b¹⁷⁷³⁴⁸, we examined whether rhuZP3a²²¹⁷⁶ or rhuZP3b¹⁷⁷³⁴⁸ trigger the acrosome reaction (AR) of human spermatozoa and we investigated the underlying mechanism. The assessment of AR was performed using chlortetracycline staining. The intracellular free calcium concentration ([Ca²⁺]i) in Fura-2/AM-loaded human sperm was monitored with a spectrofluorophotometer. We found that rhuZP3a²²¹⁷⁶ and rhuZP3b¹⁷⁷³⁴⁸ were capable of eliciting AR at different concentrations. With the addition of either peptide, the [Ca²⁺]i level was raised to a peak with or without a plateau. The AR could be inhibited by pertussis toxin (PTX), EGTA, and pimozide (a T-type calcium channel blocker), whereas verapamil was less effective in this regard. The results of the present study suggest that peptides rhuZP3a²²¹⁷⁶ and rhuZP3b¹⁷⁷³⁴⁸ have a role similar to human ZP3, and that the mechanism of the response to the peptides involves influx of calcium, the G protein pathway, and a T-type calcium channel.

     Key words: Human sperm, pertussis toxin, intracellular calcium, G protein, calcium channel, chlortetracycline