Cholesterol and phospholipids of bovine spermatozoa. II. Effect of liposomes prepared from egg phosphatidylcholine and cholesterol on sperm cholesterol, phospholipids, and viability at 4 degrees C and 37 degrees C.

Abstract

Diluents containing sonicated liposomes of egg phosphatidyl-choline (EPC), with cholesterol to phospholipid (C/P) molar ratios of 0:1, 1:2, and 1:1, were evaluated for their effect on resistance of sperm to cold shock and acrosomal maintenance at 4°C and 37°C. The effect of liposomes with C/P ratios of 0:1 and 1:1 on sperm cholesterol (C) and phospholipids (P) was also examined. Aliquots of pooled ejaculates from Holstein bulls were diluted to 1 × 108 sperm/ml in diluents containing 50 mM PIPES, 50 mM fructose, NaCl, and either no liposomes (control) or liposomes with C/P ratios of 0:1, 1:2, or 1:1 (290 mOsm, pH 7.0). Liposomes were reduced to microvesicles (20–200 nm) by sonication. Percent live sperm (PLS) were measured on untreated and cold-shocked samples (0°C/10 min) after 0 and 3 h at 37°C. Percent intact acrosomes (PIAs) were measured after 0, 3, and 6 h at 37°C immediately postdilution and after storage for 1 day at 4°C. PLS was higher after cold shock in diluents containing liposomes and was higher with EPC alone than in combination with cholesterol. Sperm resistance to cold shock declined for all diluents after 3 h at 37°C. No difference in PIAs was observed among diluents during incubation on the day of semen collection. After 4°C storage, PIAs were higher in diluents containing liposomes and highest with liposomes containing only EPC. Sperm C and P were analyzed on aliquots of 5 × 108 sperm after 0 and 3 h at 37°C and at 3 h after cooling at 4°C. No difference in the C/P ratio of sperm was observed among diluents initially or after cooling to 4°C. After 3 h at 37°C, a slightly increased C/P ratio for sperm incubated with EPC liposomes containing equimolar C and a decreased ratio for sperm incubated with liposomes of EPC alone were observed. It was concluded that liposomes of EPC protect sperm from cold shock and during cooling and storage at 4°C. Addition of cholesterol to liposomes reduces the effectiveness of the protection provided by EPC. Protection was provided to sperm immediately postdilution and therefore could not be the result of an altered sperm C/P ratio.