Functional Significance of the Pentose Phosphate Pathway and Glutathione Reductase in the Antioxidant Defenses of Human Sperm1

Abstract

Glutathione peroxidase is one of the principal antioxidant defense enzymes in human spermatozoa, but it requires oxidized glutathione to be reduced by glutathione reductase using NADPH generated in the pentose phosphate pathway. We investigated whether flux through the pentose phosphate pathway would increase in response to oxidative stress and whether glutathione reductase was required to protect sperm from oxidative damage. Isotopic measurements of the pentose phosphate pathway and glycolytic flux, thiobarbituric acid assay of malondialdehyde for lipid peroxidation, and computer-assisted sperm analysis for sperm motility were assessed in a group of normal, healthy semen donors. Applying moderate oxidative stress to human spermatozoa by adding cumene hydroperoxide, H(2)O(2), or xanthine plus xanthine oxidase or by promoting lipid peroxidation with ascorbate increased flux through the pentose phosphate pathway without changing the glycolytic rate. However, adding higher concentrations of oxidants inhibited both the pentose phosphate pathway and glycolytic flux. At concentrations of 50 microg/ml or greater, the glutathione reductase-inhibitor 1,3-bis-(2-chloroethyl) 1-nitrosourea decreased flux through the pentose phosphate pathway and blocked the response to cumene hydroperoxide. It also increased lipid peroxidation and impaired the survival of motility in sperm incubated under 95% O(2). These data show that the pentose phosphate pathway in human spermatozoa can respond dynamically to oxidative stress and that inhibiting glutathione reductase impairs the ability of sperm to resist lipid peroxidation. We conclude that the glutathione peroxidase-glutathione reductase-pentose phosphate pathway system is functional and provides an effective antioxidant defense in normal human spermatozoa.

Reactive Oxygen Species and Sperm Cells

“…On the contrary, in morphologically normal spermatozoa, cytoplasm deposits in the midpiece are extruded to allow cell elongation and condensation to occur during spermiogenesis, and the cytoplasm deposits contain large amounts of the glucose-6-phosphate dehydrogenase enzyme that produces nicotinamide adenine dinucleotide phosphate (NADPH). As a result, ROS is generated from NADPH via an intramembrane-located NADPH oxidase (NOX) [31]. So, immature spermatozoa are characterized by large amounts of cytoplasm that are expected to produce higher levels of ROS.…”

L’AMPK, régulateur de l’énergie et des fonctions des spermatozoïdes

“…Elle stimule en effet la dégradation des mitochondries défectueuses par l'intermédiaire d'ULK1/2 (Unc51-like kinase), une kinase associée au processus d'autophagie, et en activant la biogenèse de nouvelles mitochondries, via PGC1α (peroxisome proliferatoractivated receptor γ coactivator 1a) impliqué dans leur régulation, et donc, indirectement, les systèmes antioxydants [42,43]. Mais la stimulation de la régénération du NADPH par l'AMPK [44], par la voie des pentoses phosphates, mise en évidence dans les spermatozoïdes de mammifères, et qui contribue au système antioxydant par la stimulation de la GPx (NADPH-dependent glutathione peroxidase-like protein), pourrait également avoir un rôle dans cette fonction. Ces données pourraient être d'une grande utilité pour développer et améliorer les méthodes utilisées pour la manipulation et le stockage du sperme.…”

Mammalian sperm metabolism: oxygen and sugar, friend and foe

“…Production of the NADPH needed for regeneration of GSH has been shown to occur through the combined actions of hexokinase and glucose-6-phosphate dehydrogenase, with the activity of the dehydrogenase being rate-limiting (Storey et al, 1998;Williams and Ford, 2004). Mouse sperm are more heavily dependent on the GPX system for inhibition of lipid peroxidation Storey, 1984a, 1989).…”