Expression of phospholipase PLC Zeta in human spermatozoa: impact of cryopreservation

BACKGROUND: Cryopreservation is used for infertility treatment and for fertility preservation. The results of the use of frozen spermatozoa for ART (Assisted Reproductive Technology) are lower than those of fresh spermatozoa. The phospholipase C Zeta (PLCζ) protein is involved in oocyte activation. OBJECTIVES: The aim of this study was to compare the percentage of spermatozoa expressing phospholipase C Zeta protein before and after a frozen-thawing cycle. MATERIALS AND METHODS: Samples were provided after at least 2 days of sexual abstinence. A part of the fresh ejaculate (200 μL) was recovered for the study. Fifty microliters was necessary to carry out the technique before freezing. The remaining 150 μl was frozen according to a slow manual freezing technique. The samples were treated based on the procedure described by Yelumalai et al. (Fertil. Steril., 104, 2015, 561-568.e4) and Grasa et al. (Hum. Reprod. Oxf. Engl. 23, 2008, 2513-2522). RESULTS: Freezing was associated with a decrease in the percentage of spermatozoa exhibiting PLCζ (44 ± 22% before vs 31 ± 19% after, p < 0.05). The percentage of spermatozoa exhibiting PLCζ at post-acrosomal position was significantly greater before freezing (8% vs 5%, p < 0.05). There was no significant difference for the percentage of spermatozoa exhibiting PLCζ at equatorial position (15% before freezing versus 12% after thawing, NS). DISCUSSION: The results of the present study show that the presence of PLCζ on spermatozoa is decreased after freezing-thawing procedures. PLCζ is a soluble cytosolic protein (Nomikos et al., ), so it can be lost during cryopreservation. These membrane alterations are probably multifactorial. CONCLUSION: Our results, in agreement with other studies, raise the hypothesis that cryopreservation reduces spermatic PLCζ expression.