Sperm Cholesterol Content Modifies Sperm Function and TRPV1-Mediated Sperm Migration

Abstract

Transient receptor potential channels-vanilloid receptor 1 (TRPV1) regulates thermotaxis in sperm-oriented motility. We investigated the role of membrane cholesterol (Chol) on TRPV1-mediated human sperm migration. Semen samples were obtained from five normozoospemic healthy volunteers. Sperm membrane Chol content, quantified by liquid chromatography-mass spectrometry, was modified by incubating cells with 2-hydroxypropyl-ß-cyclodextrin (CD) or the complex between CD and Chol (CD:Chol). The effect on sperm migration on a 10 μM capsaicin gradient (CPS), a TRPV1 agonist, was then investigated. Motility parameters were evaluated by Sperm Class Analyser. Intracellular calcium concentration and acrosome reaction were measured by staining with calcium orange and FITC-conjugated anti-CD46 antibody, respectively. TRPV1-Chol interaction was modelled by computational molecular-modelling (MM). CD and CD:Chol, respectively, reduced and increased membrane Chol content in a dose-dependent manner, resulting in a dose-dependent increase and reduction of sperm migration in a CPS gradient. MM confirmed a specific interaction of Chol with a TRPV1 domain that appeared precluded to the Chol epimer epicholesterol (Epi-Chol). Accordingly, CD:Epi-Chol was significantly less efficient than CD:Chol, in reducing sperm migration under CPS gradient. Chol inhibits TRPV1-mediated sperm function by directly interacting with a consensus sequence of the receptor.

Membrane Cholesterol Inhibits Progesterone-Mediated Sperm Function through the Possible Involvement of ABHD2

“…In this study, we provide evidence of a widened role of membrane cholesterol levels in human sperm cells function, being negatively associated with progesterone-mediated effects, along with the previously highlighted temperature-driven migration [13], in a concentration-dependent manner. In addition, a possible direct effect of Chol on the enzyme function of the P4 membrane receptor ABHD2 was suggested by a computational approach.…”

“…An initial series of experiments were performed in order to address the proper concentration of P4 to be used in subsequent migration assays, through the migration device that was built as previously described (Figure 2A) [13,15]. To this aim, a dose-response curve was created by relating the concentration of P4 as chemoattractant and the percentage of cells accumulated within the recovery chamber of the migration device after a migration period of 30 min at 37 • C, as detailed in the methods section.…”

“…For the purpose of subsequent migration tests, the highest concentration of P4 associated sperm migration was chosen, namely 160 nM. In order to modify the cholesterol content in the sperm plasma membrane, 2-hydroxypropyl-ß-clodextrin (CD) was used as a molecular carrier of hydrophobic species, as previously described [13,16]. Incubation with di CD saturated with Chol (CD:Chol), used as Chol donor, was performed to increase the membrane Chol content as previously described [17].…”

“…However, more specific mechanisms of protein activity regulation by Chol are increasingly being disclosed. In a recent study from our group, we showed that the membrane Chol content in sperm cells reduced the temperature gradient-oriented motility, or thermotaxis, through the likely interaction with a Chol recognition amino acid consensus sequence of the transient receptor potential vanilloid 1 (TRPV1) ion channel [13]. Even more recently, the human voltage-gated proton channel (hHv1), whose activity is involved in sperm hypermotility gain during capacitation, was shown to be functionally inhibited by Chol through the stabilization of the voltage-sensing S4 segment at resting conformation [14].…”

Deterioration of Frozen Semen of Bali Cattle after Cooling at 5°C

“…Cell membranes containing cholesterol become sensitive to changes in temperature (De Toni et al, 2021). The condition of the spermatozoa cells membrane is influenced by the fatty acid composition and lipid ratio (Mandal et al, 2014).…”