403. Differential effect of hexoses on sperm metabolism and function in culture

Abstract

Currently there is lack information regarding how human spermatozoa regulate their energy metabolism. This is surprising considering that carbohydrate metabolism is a vital point for the understanding of sperm function. This coupled with the increased use of assisted reproductive technology and the importance of a well balanced culture media has led us to hypothesise that an imbalance of carbohydrate presence in the media may alter sperm function, particularly in relation to oxidative stress, DNA damage and lipid peroxidation. Sperm samples were obtained from three healthy normospermic donors for this study. Motile sperm were separated from semen samples using density gradient separation. Samples were incubated at different media conditions with varying glucose or fructose concentrations (0, 2.5, 25 mM) for 6–24hrs. Reactive oxygen species (ROS) were measured using 5-(and-6)-carboxy-2', 7'-dichlorofluorescein diacetate (DCFDA). Sperm DNA damage was determined using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-nick end labelling (TUNEL). Lipid peroxidation was assessed using the probe BODIPY (581/591) C_11. Carbohydrate uptake from the media was measured using a fluorometric procedure. Statistical differences between treatments were assessed by ANOVA and Bonferroni post-hoc test. No significant motility differences were found following treatments. Results showed an increased level of ROS production as glucose concentration increased (P < 0.05). This was accompanied by an increased number of TUNEL positive cells (P < 0.05). Furthermore, lipid peroxidation of spermatozoa was significantly increased when incubated under high glucose concentrations (P < 0.01). In contrast, increases in fructose concentrations did not alter ROS levels or the number of TUNEL positive cells. Sperm metabolised both glucose and fructose in vitro and the removal of one carbohydrate resulted in a compensatory increase in the metabolism of the other. To our knowledge, this is the first report providing evidence that altered carbohydrate metabolism may induce ROS production, lipid peroxidation and increase the number of sperm exhibiting DNA damage.