Mass Spectrometric Detection of Cholesterol Oxidation in Bovine Sperm1

Abstract

We report on the presence and formation of cholesterol oxidation products (oxysterols) in bovine sperm. Although cholesterol is the most abundant molecule in the membrane of mammalian cells and is easily oxidized, this is the first report on cholesterol oxidation in sperm membranes as investigated by state-of-the-art liquid chromatographic and mass spectrometric methods. First, oxysterols are already present in fresh semen samples, showing that lipid peroxidation is part of normal sperm physiology. After chromatographic separation (by high-performance liquid chromatography), the detected oxysterol species were identified with atmospheric pressure chemical ionization mass spectrometry in multiple-reaction-monitoring mode that enabled detection in a broad and linear concentration range (0.05-100 pmol for each oxysterol species detected). Second, exposure of living sperm cells to oxidative stress does not result in the same level and composition of oxysterol species compared with oxidative stress imposed on reconstituted vesicles from protein-free sperm lipid extracts. This suggests that living sperm cells protect themselves against elevated oxysterol formation. Third, sperm capacitation induces the formation of oxysterols, and these formed oxysterols are almost completely depleted from the sperm surface by albumin. Fourth, and most importantly, capacitation after freezing/thawing of sperm fails to induce both the formation of oxysterols and the subsequent albumin-dependent depletion of oxysterols from the sperm surface. The possible physiological relevance of capacitation-dependent oxysterol formation and depletion at the sperm surface as well as the omission of this after freezing/thawing semen is discussed.

Bivalent response to long‐term storage in liquid‐preserved boar semen: A flow cytometric analysis

“…Spermatozoa contain high amounts of polyunsaturated phospholipids and cholesterol which make them particularly vulnerable to lipid peroxidation (24,25). However, lipid peroxidation during 5 days liquid storage of boar semen has been found to be low (26), as has the incidence of intracellular reactive oxygen species (ROS) in boar spermatozoa (27).…”

A Hypomorphic Mutation in Lpin1 Induces Progressively Improving Neuropathy and Lipodystrophy in the Rat

“…A mixture of authentic PI and PS standards was used to correct for ionization and fragmentation efficiencies (23). Cholesterol was quantified as described previously, using deuterated cholesterol (Avanti Polar Lipids) as an internal standard (24).…”

Potential Risk of Senescence on Male fertility and Sperm DNA damage on Progeny

“…A study of the impact of oxidative stress on sperm-oocyte fusion has shown a biphasic response depending on the levels of oxidants [73] . Some authors demonstrated that at low level of oxidative stress spermoocyte fusion rates were increased supporting the hypothesis which suggests the role ROS play in activation of the tyrosine phosphorylation events that occur during sperm capacitation [74] and the importance of sterol oxidation in driving the efflux of cholesterol from sperm plasma membrane [75]. Conversely, at higher levels of oxidative stress, lipid peroxidation was induced in the plasma membrane and sperm-oocyte fusion was impaired, probably as a result of damage to acrosome which is involved in the fusion process between spermatozoa and oocytes [76].…”