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Protective effects of exogenous gangliosides on ROS-induced changes in human spermatozoa.

ABSTRACT: This article summarizes the available evidence on the efficacy of gangliosides to reduce the degree of reactive oxygen species (ROS)-mediated damage. The antioxidative efficacy of exogenous gangliosides in protecting different cells encouraged us to examine their ability to protect human spermatozoa. Gangliosides are sialic acid-containing glycosphingolipids with strong amphiphilic character due to the bulky headgroup made of several sugar rings with sialic acid residues and the double-tailed hydrophobic lipid moiety. The amphiphilicity of gangliosides allows them to exist as micelles in aqueous media when they are present at a concentration above their critical micellar concentration. The protective effect of ganglioside micelles on spermatozoa is believed to stem from their ability to scavenge free radicals and prevent their damaging effects. In our study, we particularly focused our attention on the protective effect of ganglioside micelles on DNA in human spermatozoa exposed to cryopreservation. The results indicate that ganglioside micelles can modulate the hydrophobic properties of the sperm membrane to increase tolerance to DNA fragmentation, thus protecting the DNA from cryopreservation-induced damage. Further actions of ganglioside micelles, which were documented by biochemical and biophysical studies, included (i) the modulation of superoxide anion generation by increasing the diffusion barrier for membrane events responsible for signal translocation to the interior of the cell; (ii) the inhibition of iron-catalysed hydroxyl radical formation due to the iron chelation potential of gangliosides; and (iii) inhibition of hydrogen peroxide diffusion across the sperm membrane.

SUBMITTER: Gavella M

PROVIDER: S-EPMC3739653 | biostudies-other | 2013 May

REPOSITORIES: biostudies-other

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Project description:BACKGROUND: There has been an ongoing debate in the reproductive field about whether mammalian spermatozoa rely on glycolysis, oxidative phosphorylation or both for their energy production. Recent studies have proposed that human spermatozoa depend mainly on glucose for motility and fertilization but the mechanism behind an efficient glycolysis in human spermatozoa is not well understood. Here, we demonstrate how human spermatozoa utilize exogenous pyruvate to enhance glycolytic ATP production, motility, hyperactivation and capacitation, events that are crucial for male fertility. METHODS: Purified human spermatozoa from healthy donors were incubated under capacitating conditions (including albumin, bicarbonate and glucose) and tested for changes in ATP levels, motility, hyperactivation and tyrosine phosphorylation after treatment with pyruvate. The experiments were repeated in the presence of sodium cyanide in order to assess the contribution from mitochondrial respiration. The metabolism of (13)C labeled glucose and pyruvate was traced by a combination of liquid chromatography and mass spectrometry. RESULTS: The treatment of human spermatozoa with exogenous pyruvate increased intracellular ATP levels, progressive motility and hyperactivation by 56, 21 and 130%, respectively. In addition, added pyruvate induced a significant increase in tyrosine phosphorylation levels. Blocking of the electron transport chain did not markedly affect the results, indicating that the mechanism is independent of oxidative phosphorylation. However, the observed effects could be counteracted by oxamate, an inhibitor of lactate dehydrogenase (LDH). Metabolic tracing experiments revealed that the observed rise in ATP concentration resulted from an enhanced glycolytic flux, which was increased by more than 50% in the presence of exogenous pyruvate. Moreover, all consumed (13)C labeled pyruvate added was converted to lactate rather than oxidized in the tricarboxylic acid cycle. CONCLUSIONS: Human spermatozoa seem to rely mainly, if not entirely, on glycolysis as the source of ATP fueling the energy-demanding processes of motility and capacitation. The efficient glycolysis is dependent on exogenous pyruvate, which indirectly feeds the accelerated glycolysis with NAD(+) through the LDH-mediated conversion of pyruvate to lactate. Pyruvate is present in the human female reproductive tract at concentrations in accordance with our results. As seen in other mammals, the motility and fertility of human spermatozoa seem to be dictated by the available energy substrates present in the conspecific female.

Project description:BackgroundTo investigate the protective effects of exogenous spermine on renal ischemia-reperfusion injury in rats.Methods(I) Different doses of spermine were injected into rats to determine the safe dose on the kidneys. Kidney toxicity was assessed by hematoxylin and eosin (HE) staining of kidney tissue and enzyme-linked immunosorbent assay (ELISA) detection of neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule 1 (KIM-1) in the venous blood. (II) A rat model of renal ischemia-reperfusion injury was established. Different doses of spermine were injected into the rats through the tail vein 30 minutes before and 3 days after the establishment of the model. Blood samples and kidney tissues were collected and renal injury was assessed via HE staining of the renal tissue, detection of apoptosis using the TUNEL assay, and detection of NGAL and KIM-1 in blood samples using ELISA. (III) Human HK-2 renal tubular epithelial cells were cultured under hypoxia/reoxygenation conditions. To evaluate the protective effects of spermine, apoptosis was assessed by flow cytometry and TUNEL assay. The mechanisms underlying the effects of spermine were studied using Western blot analyses.ResultsAt spermine concentrations below 200 µM (2 mL/kg body weight), no significant damage to the kidney was observed by HE staining, and there was no significant difference in NGAL and KIM-1 levels between rats treated with spermine and control rats (P<0.05). At spermine doses below 200 µM, HE staining showed that the degree of renal ischemia-reperfusion injury was gradually alleviated with increasing doses of spermine. TUNEL assays demonstrated that spermine reduced the apoptosis of renal tissue, and increasing doses of spermine gradually decreased the levels of NGAL and KIM-1 in the blood compared with the control group (P<0.05). Western blot analysis revealed that spermine increased the expression of pro-caspase9, phosphorylated protein kinase B (p-Akt), hypoxia-inducible factor 1 alpha (HIF-1α), B cell lymphoma 2 (Bcl-2), and Bcl2 interacting protein 3 (BNIP3), and decreased the expression of cleaved caspase-3, Bax and cytochrome C compared to control cells.ConclusionsExogenous spermine exerted a protective effect on renal ischemia-reperfusion injury in rats by inhibiting the apoptosis of renal tubular epithelial cells.

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Protective effects of exogenous gangliosides on ROS-induced changes in human spermatozoa.

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