Assessment of sperm motility, viability and apoptosis in human spermatozoa after hydrogen peroxide exposure

Abstract

ObjectiveMale infertility has been linked to excessive production of ROS in semen. ROS is involved in sperm cell apoptosis signaling pathway. Apoptosis is distinguished from necrosis, or accidental cell death, by characteristic morphological and biochemical changes. Our objective was to investigate sperm motility, viability, and incidence of apoptosis after exposure to hydrogen peroxide (H2O2).DesignProspective study.Materials and methodsSemen samples were collected from 12 semen donors. The neat samples were subjected to double density gradient centrifugation to separate mature and immature sperm subpopulation. Each was resuspended in buffer and divided into 2 aliquots. One aliquot was incubated with 50 uL of freshly prepared H2O2 added to 1 mL of sperm suspension at a final concentration of 20 uM for 15 min at 37°C, and the other aliquot served as control. The incidence of apoptosis and necrosis was assessed by flowcytometry using the YoPro-1 dye which enters apoptotic cells giving green fluorescence, and propidium iodide (PI), which stains necrotic sperm cells giving red fluorescence.ResultsTableComparing different sperm parameters after exposure to H2O2Values are mean (±SD). Paired Student t-test was utilized to compare different variables in different spem fractions. P<0.05 was considered significant. Open table in a new tab ConclusionsExposure of human spermatozoa to H2O2 is associated with a decrease in sperm viability. The increase in the proportion of apoptotic sperm reached significance only in the immature fractions. There was no apparent effect on sperm motility. Thus sperm motility may not serve as a marker for early oxidative sperm damage. ObjectiveMale infertility has been linked to excessive production of ROS in semen. ROS is involved in sperm cell apoptosis signaling pathway. Apoptosis is distinguished from necrosis, or accidental cell death, by characteristic morphological and biochemical changes. Our objective was to investigate sperm motility, viability, and incidence of apoptosis after exposure to hydrogen peroxide (H2O2). Male infertility has been linked to excessive production of ROS in semen. ROS is involved in sperm cell apoptosis signaling pathway. Apoptosis is distinguished from necrosis, or accidental cell death, by characteristic morphological and biochemical changes. Our objective was to investigate sperm motility, viability, and incidence of apoptosis after exposure to hydrogen peroxide (H2O2). DesignProspective study. Prospective study. Materials and methodsSemen samples were collected from 12 semen donors. The neat samples were subjected to double density gradient centrifugation to separate mature and immature sperm subpopulation. Each was resuspended in buffer and divided into 2 aliquots. One aliquot was incubated with 50 uL of freshly prepared H2O2 added to 1 mL of sperm suspension at a final concentration of 20 uM for 15 min at 37°C, and the other aliquot served as control. The incidence of apoptosis and necrosis was assessed by flowcytometry using the YoPro-1 dye which enters apoptotic cells giving green fluorescence, and propidium iodide (PI), which stains necrotic sperm cells giving red fluorescence. Semen samples were collected from 12 semen donors. The neat samples were subjected to double density gradient centrifugation to separate mature and immature sperm subpopulation. Each was resuspended in buffer and divided into 2 aliquots. One aliquot was incubated with 50 uL of freshly prepared H2O2 added to 1 mL of sperm suspension at a final concentration of 20 uM for 15 min at 37°C, and the other aliquot served as control. The incidence of apoptosis and necrosis was assessed by flowcytometry using the YoPro-1 dye which enters apoptotic cells giving green fluorescence, and propidium iodide (PI), which stains necrotic sperm cells giving red fluorescence. ResultsTableComparing different sperm parameters after exposure to H2O2Values are mean (±SD). Paired Student t-test was utilized to compare different variables in different spem fractions. P<0.05 was considered significant. Open table in a new tab Values are mean (±SD). Paired Student t-test was utilized to compare different variables in different spem fractions. P<0.05 was considered significant. ConclusionsExposure of human spermatozoa to H2O2 is associated with a decrease in sperm viability. The increase in the proportion of apoptotic sperm reached significance only in the immature fractions. There was no apparent effect on sperm motility. Thus sperm motility may not serve as a marker for early oxidative sperm damage. Exposure of human spermatozoa to H2O2 is associated with a decrease in sperm viability. The increase in the proportion of apoptotic sperm reached significance only in the immature fractions. There was no apparent effect on sperm motility. Thus sperm motility may not serve as a marker for early oxidative sperm damage.