Involvement of cannabinoid receptor‐1 activation in mitochondrial depolarizing effect of lipopolysaccharide in human spermatozoa

Abstract

Gram-negative bacteria frequently involved in urogenital tract infections release the endotoxin lipopolysaccharide (LPS); its receptor, toll-like receptor-4 (TLR4), has been recently identified in human spermatozoa, and its direct activation has been suggested in mediating adverse effects of LPS on human spermatozoa. However, the underlying signal transduction remains to be clarified. In other cell types, LPS induces the generation of endocannabinoids, which are involved in mediating endotoxin effects. In human spermatozoa, which exhibit a completely functional endocannabinoid system, the activation of cannabinoid receptor-1 (CB1) inhibited sperm mitochondrial membrane potential (ΔΨm). In this study, we tested the hypothesis of a contribution of CB1 activation by sperm-generated endocannabinoids in the adverse effects exerted by LPS on human spermatozoa. The exposure of motile sperm suspensions to E. coli LPS produced a significant decrease in sperm ΔΨm, assessed at flow cytometry with JC-1, similar to that induced by Metanandamide (Met-AEA), a non-hydrolyzable analogue of the endocannabinoid AEA. The LPS-induced inhibition of ΔΨm was prevented by the selective CB1 cannabinoid receptor antagonist, SR141716. However, the inhibition of ΔΨm induced by either LPS or Met-AEA did not affect sperm motility. Consistent with this finding, the CB1-mediated inhibition of ΔΨm was neither associated to mitochondrial generation of reactive oxygen species as evaluated by flow cytometry with MytoSox Red nor to apoptosis pathway activation as evaluated with cytoflorimetric assay for activated caspase-9 and caspase-3. Any oxidative genomic damage was also ruled out with the cytoflorimetric quantification of the oxidized base adduct 8-hydroxy-2'-deoxyguanosine. In conclusion, E. coli LPS inhibited sperm ΔΨm through the activation of CB1, but this effect was not accompanied to the activation of mitochondrial dysfunction-related apoptotic/oxidative mechanisms, which could affect sperm motility and genomic integrity.

Coenzyme Q10 improves the quality of sheep sperm stored at room temperature by mitigating oxidative stress

“…Studies have indicated that the sperm MMP is positively correlated with ATP content and sperm motility (Barbonetti et al, 2014). The results of this experiment indicated that 50 μmol/L CoQ 10 significantly improved sperm ATP content, MMP, and motility, which is consistent with the above results.…”

“…This may be related to the scavenging of free radicals by CoQ 10 resulting in inhibition of LPO, which is quite evident from the lower MDA content and ROS content in this group. Studies have indicated that the sperm MMP is positively correlated with ATP content and sperm motility (Barbonetti et al, 2014). The results of this experiment indicated that 50 μmol/L CoQ 10 significantly improved sperm ATP content, MMP, and motility, which is consistent with the above results.…”

Characterisation and localisation of the endocannabinoid system components in the adult human testis

“…In humans the data are scarce, and largely limited to expression at the RNA level in whole testis tissue extracts, including the testis-dominant expression of CB2A variant 24,25 . At the protein level previous human studies only investigated ejaculated spermatozoa; CNR1 alone was detected in one study 17 , while both CNR1 and CNR2 were found by other groups 19,20,34 .…”

Semen leukocytes and oxidative-dependent DNA damage of spermatozoa in male partners of subfertile couples with no symptoms of genital tract infection

“…3). Positive controls were prepared by incubating samples for 1 h at RT with H 2 O 2 (2 mM) and FeCl2Á4H 2 O (1 mM) in a final volume of 200 lL of PBS, as previously described (Barbonetti et al, 2014), before fixation procedure.…”