Porcine follicular fluid as chemoattractant improves sperm selection and in vitro fertilization.

VIERA, LA; AGUILERA A; DIANA, A; MATAS, C

Nantes

Congreso; 34th Scientific meeting of the AETE; 2018

Association of Embryo Technology in Europe

Porcine follicular fluid as chemoattractant improves sperm selection and in vitro fertilizationVieira LA1,2*, Aguilera A3, Diana A1 and Matás C1,21Department of Physiology, International Excellence Campus for Higher Education and Research (Campus Mare Nostrum), University of Murcia, Murcia 30100, Spain, 2Institute for Biomedical Research of Murcia (IMIB), Murcia, Spain.3Faculty ofNatural Sciences(FCEN), National University of Cuyo, Mendoza, Argentina. luisalberto.vieira@um.esIntroduction: Under physiologic conditions, different biofluids (follicular fluid (FF), oviductal fluid (OF), and secretion of cumulus-oocyte complex (COCs)) participate in the spermatozoa selection previous to fertilization. Despite the progesterone (P4) being part of the composition of these biofluids, it?s also considered as the main chemoattractant (Blengini et al., Asian Journal of Andrology, 13, 769-773, 2011). However, there are others components not defined in these media that could select spermatozoa more efficiently. Thus, the aim of this study was to investigate the ability of biofluids for sperm selection and the effect on in vitro fertilization (IVF) parameters.Materials and Methods: The OF and FF were collected in the periovulatory phase according to Coy et al., 2008, Canovas et al., 2017 respectively, and the conditionated medium was obtained after 48 h of in vitro maturation.To perform the assays, a chemotaxis system was designed using a Petri cell culture dishes (35 x 10 mm) with four wells separated by 3 mm (GN627170, Sigma). Six wells were filled with fresh spermatozoa (20x106/mL) from fertile boars (N=6) that were capacitated for 45 min in 180 µL of the capacitation media (TALP), previously equilibrated for 3h at 38.5ºC and 5% CO2. The opposite wells (six) were filled with TALP (control group) and TALP supplemented with the chemoattractants (experimental groups, see below). Posteriorly, capillars of 3-4 mm long were placed for 20 min between the wells containing capacitated spermatozoa and the opposite ones. After that, the capillars were removed and 22 (per replicate) denuded in vitro maturated oocytes were deposited with spermatozoa (20x103) with corresponding chemoattractans and control group (opposite wells). According to the chemoattractants in the opposite wells, the experimental groups were: 1) TALP (control), 2) FF (1%), 3) OF (1%), 4) CM (2%) 5) P4 (28.3 pM), and 6) FF, OF, CM, and P4: Σ. After 18 h the oocytes were fixed and IVF parameters were evaluated. Data were analysed by ANOVA followed by Tukeypost hoc to compare means and standard error (p