The effects of progesterone and cholesterol on sperm capacitation, acrosome reaction, and in vitro embryo production in cattle

Afsaneh Motamed Khorasani
Progesterone (P₄) is known to induce mammalian sperm acrosome reaction (AR) in vitro, whereas cholesterol (CH) one of the major components in seminal plasma, inhibits AR. The objectives of this study were: 1) to elucidate the in vitro effects of P₄, 17 β-estradiol (E₂), CH, and their combination on cryopreserved bovine sperm A R , 2) to study the mechanisms o f CH action on P₄-induced A R , and 3) to determine the effects of exogenous P₄ and CH on zonabinding ability of bovine sperm and in
more » ... e sperm and in vitro fertilization (IVF). For the first two objectives, swim-up separated sperm from six bulls were incubated in modified Tyrodes* medium for 0-6 h (38.5°C, 5% CO₂). Acrosome reaction was determined following addition of P4 (10 μg/m1), E₂ (10 μg/ml), db-cAMP (1 mM), forskolin (10 μM), and Ca²⁺-ionophore (10 μm), either in the presence or absence of CH (1 (μg/ml). The effects of β-sitosterol (1 μg/ml), a CH plant analog, and CH on P₄-induced AR were also compared. For zona-binding assay, thawed sperm were washed in modified Tyrodes' medium. Sperm (0.2 x 106 sperm/ml) were pre-treated with P₄ (10 pig/ml), either in the presence or absence o f CH (1μg/mi) at 38.5°C, 5% CO₂. Sperm were co-incubated with cumulus-free oocytes for 18 nr. Following washing, fixing, and staining, the number of sperm attached to oocytes was counted under a fluorescent microscope. For IVF, 30 min prior to the addition of matured oocytes, sperm (0.2 x 10⁶ sperm/ml) were pre-treated with P₄ (10 μg/ml), CH (1 μg/ml), combination of P₄ (10 μg/ml) and CH (1 μg/ml), and cyclodextrin (10 μg/ml). Sperm exposed oocytes were then incubated in modified Tyrodes' medium (supplemented with 10% serum) at 38.5°C and 5% CO₂. Cleavage rates, as an indicator of fertilization, were recorded after 72 nr. Acrosome reaction was induced by P₄ (34.4 ± 2.5%), E₂ (4.5 ± 0.7%), db-cAMP (49.3 ± 2.0%), forskolin (81.8 ± 1.2%), and Ca²⁺-ionophore (95.7 ± 1.0%) immediately after-thawing (p < 0.05 for all). Cholesterol and β-sitosterol inhibited P₄- induced A [...]
doi:10.14288/1.0089077 fatcat:tlbqc4s3snaoherdugtdkcyxce