DNA fragmentation in human sperm after magnetic-activated cell sorting.
OBJECTIVE
As fertilization with unselected apoptotic spermatozoa may contribute to failures in assisted reproductive techniques, it has become essential to remove this type of sperm in order to increase the success rates. Magnetic-activated cell sorting (MACS) is a sperm preparation technique that isolates non-apoptotic spermatozoa based on the expression of phosphatidylserine in the membrane of apoptotic sperm. Therefore, we aimed to evaluate whether there was a significant decrease in sperm DNA fragmentation (sDNAfrag) and verify which protocol was the most efficient.


METHODS
Hundred semen samples were allocated into five distinct groups and processed according to a combination of MACS with density gradient centrifugation (DGC) and swim-up (SU) techniques. Sperm DNA fragmentation was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay.


RESULTS
Groups DGC-SU (73.4 %), DGC-MACS-SU (78.9 %), DGC-SU-MACS (53.8 %) and MACS-SU (73.5 %) presented a significant decrease in sDNAfrag but the highest reduction rate was obtained with MACS-DGC-SU (83.3 %). The later was also negatively correlated with sperm vitality, membrane integrity and progressive motility. Additionally, teratozoospermic patients presented a tendency to have lower sDNAfrag reduction rates than asthenozoospermic and asthenoteratozoospermic patients.


CONCLUSIONS
Based on the results, MACS showed potential to optimize the sDNAfrag reduction rate, when applied to raw semen, before DGC and SU, especially in samples with low values of progressive motility, vitality and hypoosmotic swelling test.
Department of Microscopy, Laboratory of Cell Biology, Institute of Biomedical Sciences Abel Salazar (ICBAS), University of Porto (UP), Multidisciplinary Unit for Biomedical Research (UMIB), Rua Jorge Viterbo Ferreira, 228, 4050-313, Porto, Portugal.
Sara
S
Bucar
Ana
A
Gonçalves
Eduardo
E
Rocha
Alberto
A
Barros
Mário
M
Sousa
Rosália
R
Sá
D000328:Adult; D053627:Asthenozoospermia / Q000235:genetics* / Q000473:pathology; D002469:Cell Separation; D053938:DNA Fragmentation; D006801:Humans; D008280:Magnetics; D008297:Male; D013081:Sperm Motility / Q000502:physiology; D013094:Spermatozoa