Related Experiment Videos
Measuring apoptosis in human spermatozoa: a biological assay for semen quality?
G J Oosterhuis1, A B Mulder, E Kalsbeek-Batenburg
1Department of Obstetrics and Gynecology, Medisch Spectrum Twente Hospital Group, Enschade, The Netherlands.
Objective:
[1] To determine whether apoptosis can be measured in ejaculated spermatozoa by flow cytometry using the Annexin V assay, which measures expression of phosphatidylserine on the outer leaflet of the cell membrane, or the TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP [deoxy-uridine triphosphate] nick end labeling) assay, which measures occurrence of DNA strand breaks and [2] to correlate the outcome with routine semen variables and the hypoosmotic swelling (HOS) test.
Design:
Pilot study and clinical trial.
Setting:
Large teaching hospital and fertility center.
Patient(S):
Men whose semen was studied for various reasons.
Main Outcome Measure(S):
Percentage of apoptotic spermatozoa by two different assays, percentage of necrotic spermatozoa, concentration and motility of spermatozoa, and outcome of the HOS test.
Result(S):
Apoptosis can be measured in spermatozoa by flow cytometry using the Annexin V assay and the TUNEL assay. Twenty percent of spermatozoa were apoptotic according to both assays. A significant inverse correlation was seen between phosphatidylserine expression (Annexin V assay) and sperm concentration (r = -0.389; P<.05) and motility (r = -0.289; P<.05). A highly significant inverse correlation was seen between DNA double-strand breaks (TUNEL assay) and sperm concentration (r = -0.629; P<.0001).
Conclusion(S):
Flow cytometry can easily and reliably detect phosphatidylserine expression on the outer leaflet of the cell membrane and DNA strand breaks, both of which are hallmarks of apoptosis. About 20% of ejaculated spermatozoa are apoptotic, and the concentration of spermatozoa is lower in men with more apoptotic spermatozoa.