Inhibition of the cholesterol transporter ABCA1 by probucol decreases capacitation and tyrosine phosphorylation of dog spermatozoa, and is dose dependent

• Published in: Theriogenology 2023-02, Vol.197, p.159-166
• Main Authors: Schäfer-Somi, S., Claaßen, S., Lechner, D.
• Format: Article
• Language: English
• Subjects: ABCA1; Acrosome Reaction; Animals; Capacitation; Cholesterol; Cholesterol - metabolism; Dogs; Male; Phosphorylation; Probucol - metabolism; Probucol - pharmacology; Semen - metabolism; Sperm Capacitation; Sperm membrane; Sperm Motility - physiology; Spermatozoa - physiology; Tyrosine - metabolism; Tyrosine - pharmacology
• ISSN: 0093-691X; 1879-3231
• DOI: 10.1016/j.theriogenology.2022.11.046

The ATP binding cassette (ABC) transporter molecule ABCA1 participates in the cholesterol transport within and through cell membranes. We recently demonstrated that in dog spermatozoa, capacitation could be decreased with probucol (PRO), an ABCA1 specific antagonist. In this study, a dose-effect relationship of PRO on dog sperm capacitation, tyrosine phosphorylation and cholesterol efflux from the sperm plasma membrane was investigated. A total of 16 ejaculates from dogs of different breeds, aged 2–4 years were used. Sperm motility and membrane integrity in the main fraction was determined by CASA. Samples were stained with a boron dipyrromethene difluoride (BODIPY) fluorophore (P9672, Sigma- Aldrich, A) diluted in DMSO at a final concentration of 0.4 μM. All samples were divided into 5 aliquots, with 0, 100, 250, 500 and 1000 μM of PRO. After incubation at 37 °C for 2 h, PI was added and flow cytometry performed. All aliquots were examined for capacitation and acrosome reaction by using the CTC assay and tyrosine phosphorylation (TP). Membrane integrity was measured in all aliquots to investigate the effect of PRO on cell membranes. Membrane integrity did not differ between controls (0 μM), and 100, 250 and 500 μM PRO, but decreased with 1000 μM PRO (p