CHANGES IN N-ACETYLGLUCOSAMINE CONTENT ASSOCIATED TO CAPACITACION AND CHEMOTAXIS IN PORCINE SPERMATOZOA

BUTAZZONI A; AGUILERA AC; ALVAREZ P; DIANA A; SOSA MA

Congreso; XXXVI Reunión Anual de la Sociedad de Biología de Cuyo; 2020

Sociedad de Biología de Cuyo

In mammals only a small number of ejaculated spermatozoa (SPZ) reach the region of the oviduct (ampulla) after the copula, where they encounter and fertilize the egg . It has been suggested that a sperm subpopulation is selected during their transit trought the female genital tract, so that only those with high fertilizing capability and the best skills for supporting embryo development can fertilize the egg. Fluids of the female genital tract; such as the follicular (FF), and the oviductal fluid (OF), and the secretion of cumulus-oocyte complex (COCs), could promote the SPZ chemotaxis to the fertilization site. . Progesterone (P4) is considered as an effective chemoattractant in most mammalian species, though other components of the fluids could attract SPZ even more efficiently.In the present study, we evaluated possible changes in carbohydrate composition of sperm surface after capacitation and chemotaxis. For this, we detrmined the content of N-acetyl-glucosamine (NAG) in porcine ejaculated SPZ after the mentioned processesby using WGA-FITC lectin and flow cytometry. We observed that NAG content was significantly higher in the capacitated SPZ (30 min in capacitation media TALP, at 38.5ºC and with 5% CO2) compared to fresh ejaculated SPZ or SPZ stored in BTS (diluent media). For the chemotaxis assays, OF and FF collected from prepubertal (OF0 and FF0) and periovulatory (OF2 and FF2) were used as chemoatracttants for SPZ. Six wells were filled with fresh spermatozoa (20x106/mL) from fertile boars (N = 3) selected in a discontinuous percoll gradient and immediately transferred to TALP, previously equilibrated at 38.5ºC and 5% CO2. The opposite wells of the chemotaxis chamber (six) were filled with TALP (control group) or TALP supplemented with the chemoattractants as indicated: 1) TALP (control), 2) FF0 (1,25%), 3) FF2 (1,25%), 4) OF0 (1,25%), 5) OF2 (1,25%) 6) P4 (28.3 pM). After 20 min at 38.5ºC and 5% CO2, the SPZ from the opposite wells were rescued, processed for NAG detection and analyzed by flow cytometry. We observed that NAG content was significantly lower in the SPZ obtained from the groups 3 and 6 compared to the control group or the original SPZ (P < 0.05).. These preliminar results suggest that FF2 and P4 can attract selectively a SPZ subpopulation with low content of NAG in the plasma membrane under the in-vitro conditions.