113 The effects of bisphenols on cryopreserved bovine spermatozoa in vitro

Environmental agents with the capability to disrupt reproductive function, such as endocrine-disrupting compounds (EDCs), can contribute to negative reproductive outcomes both in farm animals and humans. Although the observed decline in cattle fertility is not solely due to increased exposure to EDCs—as a large number of other factors, such as genetic improvement for milk production at the expense of reproductive performance, certainly play a key role—an association between exposure of bulls and dairy cattle to EDC-contaminated drinking water/feed and reduced reproductive performance has been proven. The endocrine disruptor bisphenol A (BPA) is of particular interest because of its abundance in plastic and its known oestrogenic, antiandrogenic, and antithyroid effects. Furthermore, the growing use of “BPA-free” products has introduced other bisphenols, such as BPS and BPF as substitutes for BPA; however, their impact on fertility is even less known. Here, we hypothesise that bisphenols affect sperm parameters and survival, diminishing bull fertility overall. To test our hypothesis, we exposed cryopreserved bovine sperm from three different bulls (3 samples from each bull), suspended in 1 mL HEPES buffer, to 0.05 mg mL⁻¹ of BPA, BPS, or BPF in vitro to elucidate the effects of bisphenols on sperm motility, morphology, and apoptosis. Sperm motility (progressive motility in particular) is a major predictor of bull breeding soundness, providing an indication of a sperm’s potential to reach an ovum. Motility was measured via a computer-aided sperm analysis (CASA) program, while sperm morphology (percentage of morphologically normal head, midpiece and tail structures) was determined by staining and light microscopy. Furthermore, an annexin V/ propidium iodide assay and flow cytometry technique were used to measure the percentage of healthy, apoptotic, or necrotic sperm after bisphenol exposure. Our results showed that BPA, but not BPS or BPF, decreased the percentage of progressively motile sperm (900 sperm counted per bull per treatment group) after 4 h of exposure (P < 0.0001; Kruskal–Wallis test). As for morphology, there were no significant changes between bisphenol-treated and nontreated sperm (600 sperm counted per bull per treatment group). Last, preliminary results suggest a 30% increase in apoptosis in the BPA-treated group, but no increase in the BPS or BPF groups (10 000 sperm counted per bull per treatment group). Thus, we speculate that BPA is more harmful to sperm than BPS and BPF because it may be decreasing sperm motility through initiating apoptotic pathways. Future experiments will include assessments of the effects of bisphenols on fresh bull semen, which will allow us to compare the impacts of these EDCs on sperm before and after cryopreservation, a technique routinely used in cattle breeding. This study will ultimately further our knowledge on the effects of bisphenols on bull fertility and aid in determining whether BPS and BPF are safer alternatives to BPA.

This research was funded by NSERC and OVC.