Reactive oxygen species may be involved in the signaling of equine sperm chemotaxis

LOSINNO, LUIS; DOMÍNGUEZ, ESTEBAN; MORENO, AYELEN; GIOJALAS, LAURA CECILIA

Cambridge

Simposio; XIIth International Symposium on Equine Reproduction; 2018

Equine Science Society

To find out the way to the oocyte, spermatozoa may be oriented by chemical cues (e.g., progesterone), transport mechanism called chemotaxis (Eisenbach et al. Reviews Molecular Cell Biology; 2006. 4:276-285). Only capacitated spermatozoa can orient their movement by chemotaxis, where part of the signaling (at least in humans) is mediated by the AC-cAMP-PKA pathway (Teves et al. PLoS ONE; 2009. 12: e8211) and oxidative state (Sanchez et al. Fertility and Sterility; 2010. 1:150-153). The aim of this work was to verify whether reactive oxidative species (ROS) are also involved in the chemotaxis signaling and by which mechanism. Sperm samples from three stallions where frozen-thawed, removing the seminal plasma and cryoprotectant by a modified swim up. Spermatozoa were incubated in BWW media, with or without capacitating conditions (25 mM NaHCO3 and 0.3%BSA), at 38.5°C at an atmosphere of 5% CO2 on air, for 45 minutes. For sperm chemotaxis assays, the Sperm Selection Assay (SSA) device was used (Gatica et al. Molecular Human Reproduction;2013. 9: 559-569) which consists of two wells connected by a tube. Well 1 (W1) was filled with the sperm suspension and well 2 (W2) with the attractant solution, which diffused along the connecting tube as a gradient. After the SSA, chemotactic spermatozoa are accumulated in W2. The percentage of sperm accumulation in W2 was determined as the difference between with and without attractant. Since sperm capacitation and chemotaxis are tight associated1, the optimal concentration of H2O2 that induces capacitation was evaluated by protein tyrosine phosphorylation (PY). The level of capacitated spermatozoa was significantly increased with 0.2 µM H2O2 (p