The improvement of the embryo culture media is of high relevance due to its influence on successful implantation rates, pregnancy, neonatal outcomes, and potential effects in adult life. The ideal conditions for embryo development are those naturally occurring in the female reproductive tract, i.e., the oviductal and uterine fluids. To shed light on the differences between chemical and natural media, we performed the first comparative study of the low abundance proteins in plasma, uterine, and oviductal fluid collected, simultaneously, from healthy and fertile women that underwent a salpingectomy. The rationale for this design derives from the fact that high-abundant proteins in these fluids are usually those coming from blood serum and frequently mask the detection of low abundant proteins with a potentially significant role in specific processes related to the embryo–maternal interaction. The proteomic analysis by 1D-nano LC ESI-MSMS detected several proteins in higher amounts in oviductal fluid when compared to uterine and plasma samples (RL3, GSTA1, EZRI, DPYSL3, GARS, HSP90A). Such oviductal fluid proteins could be a target to improve fertilization rates and early embryo development if used in the culture media. In conclusion, this study presents a high-throughput analysis of female reproductive tract fluids and contributes to the knowledge of oviductal and uterine secretome.
The egg is a spherical cell encapsulated by the zona pellucida (ZP) which forms a filamentous matrix composed of several glycoproteins that mediate gamete recognition at fertilization. Studies on molecular mechanisms of sperm-egg binding are limited in many mammalian species by the scarcity of eggs, by ethical concerns in harvesting eggs, and by the high cost of producing genetically modified animals. To address these limitations, we have reproduced a three-dimensional (3D) model mimicking the oocyte’s shape, by means of magnetic sepharose beads coated with recombinant ZP glycoproteins (BZP) and cumulus cells. Three preparations composed of either ZP2 (C and N-termini; BZP2), ZP3 (BZP3) or ZP4 (BZP4) were obtained and characterized by protein SDS-PAGE, immunoblot and imaging with confocal and electron microscopy. The functionality of the model was validated by adhesion of cumulus cells, the ability of the glycoprotein-beads to support spermatozoa binding and induce acrosome exocytosis. Thus, our findings document that ZP-beads provide a novel 3D tool to investigate the role of specific proteins on egg-sperm interactions becoming a relevant tool as a diagnostic predictor of mammalian sperm function once transferred to the industry.
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