DNA fragmentation in human sperm after magnetic-activated cell sorting
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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.
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.
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.
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.
KeywordsApoptosis DNA fragmentation Sperm TUNEL MACS
We would like to acknowledge Ângela Alves, Technical assistant for teaching and research (ICBAS-UP) in laboratorial assistance, Claúdia Osório, Bsc, Biologist (CGR-ABarros) in spermiogram analysis assistance and Gabriela Rodrigues, PhD, Assistant Professor (University of Lisbon) for reviewing the manuscript.
Multidisciplinary Unit for Biomedical Research (UMIB) is funded by National Funds through FCT-Foundation for Science and Technology, under the Pest-OE/SAU/UI0215/2014.
Conflict of interest
The authors declare that they have no conflict of interest.
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