Effects of hydrogen peroxide on DNA and plasma membrane integrity of human spermatozoa.
OBJECTIVE
To evaluate the effects of oxidative stress on DNA and plasma membrane integrity of human spermatozoa.


METHODS
Prospective cohort study.


METHODS
University-based, tertiary-care infertility center.


METHODS
Men (n = 10) undergoing infertility investigation.


METHODS
Purified populations of sperm with high motility were separated using Percoll density gradients. Then, spermatozoa were incubated with 0, 10, 100, and 200 microM hydrogen peroxide (H(2)O(2)) under capacitating conditions.


METHODS
Motion parameters were assessed by computer analysis. Genomic integrity was examined by the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL) assay. Plasma membrane integrity was evaluated by the annexin V-binding assay, a measure of phosphatidylserine translocation.


RESULTS
Under basal conditions, there was a significant and negative relationship between sperm motility and the percentages of sperm with DNA fragmentation and membrane translocation of phosphatidylserine. After a 2-h incubation, there was a significant, dose-dependent effect of H(2)O(2) on motion parameters (decrease) and DNA fragmentation (increase). The percentage of annexin V(-) live (normal) cells declined significantly as the level of oxidative stress increased. Although the percentages of annexin V(+) live cells (sperm depicting translocation of phosphatidylserine) and necrotic cells increased at the highest H(2)O(2) levels, these changes were not significant.


CONCLUSIONS
In vitro sperm incubation with H(2)O(2) induces DNA fragmentation in a dose-dependent fashion. The sublethal effects of oxidative stress on motion parameters were not significantly associated with membrane translocation of phosphatidylserine.
Kemal Duru
N
N
Department of Obstetrics and Gynecology, GATA School of Medicine, Ankara, Turkey.
Morshedi
M
M
Oehninger
S
S
D017304:Annexin A5 / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D015331:Cohort Studies; D004247:DNA / Q000187:drug effects*; D053938:DNA Fragmentation; D016678:Genome; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008297:Male; D009336:Necrosis; D010718:Phosphatidylserines / Q000378:metabolism; D011446:Prospective Studies; D013081:Sperm Motility; D013094:Spermatozoa / Q000502:physiology*
D017304:Annexin A5; D010718:Phosphatidylserines; D004247:DNA; D006861:Hydrogen Peroxide