REGULATION OF PORCINE SPERM FUNCTION BY PROSTASOMES

FISCHMAN, ML; PIEHL, LL; MOLEJON MI; ZAMPINI, MC; CISALE, H; MIRANDA PV

Budapest

Congreso; 16th International Congress on Animal Reproduction - ICAR 2008 Budapest; 2008

The mammalian male reproductive tract has the ability to secrete membrane vesicles usually called prostasomes. These vesicles can interact with sperm but the function and relevance of this interaction is controversial. In this report, the effect of vesicles isolated from pig seminal plasma in sperm function was analyzed. Samples were obtained by the gloved hand method. Prostasome-like vesicles were isolated by ultracentrifugation of seminal plasma at 100000 g and molecular filtration on a Sephadex G-200 column. Sperm were diluted in Tyrode medium supplemented with 3 mg/ml BSA and incubated for 3 hs at 39C under 5% CO2. After incubation, sperm were treated with Lysophosphatidylcholine (LPC, 100 g/ml) for 30 min to induce acrosome reaction (AR). To analyze the effect of prostasomes on different sperm functions, vesicles (3-fold seminal plasma concentration) were included during the whole capacitation period. Tyrosine phosphorylation (TyrP) was analyzed by Western blot. Apical plasma membranes were obtained by nitrogen cavitation (600 psi) and sequential centrifugation. Cholesterol (Cho) content was determined with the Colestat® kit (Wiener Lab) and membrane fluidity by the order parameter (S) using electron spin resonance (a decrease in S represents an increase in membrane fluidity). Sperm incubation under capacitating conditions increased TyrP in a subset of proteins and made sperm sensitive to LPC (AR= 286 vs 92, p