Clin Exp Reprod Med.  2022 Jun;49(2):117-126. 10.5653/cerm.2021.05120.

The relationship between reactive oxygen species, DNA fragmentation, and sperm parameters in human sperm using simplified sucrose vitrification with or without triple antioxidant supplementation

• Juanpanich T
• Suttirojpattana T
• Parnpai R
• Vutyavanich T

• KMID: 2531713
• DOI: http://doi.org/10.5653/cerm.2021.05120

Abstract

Objective
This study examined whether the addition of triple antioxidants (3A)—10 μM acetyl-L-carnitine, 10 μM N-acetyl-L-cysteine, and 5 μM α-lipoic acid—in freezing-thawing medium during human sperm cryopreservation using the sucrose vitrification (SuV) and liquid nitrogen vapor (Vapor) techniques could improve post-thaw survival of spermatozoa.
Methods
We analyzed 30 samples from healthy human sperm donors. Each sample was allocated into one of five groups: fresh control, SuV, SuV+3A, Vapor, and Vapor+3A. The sperm motility, morphology, viability, intracellular and extracellular reactive oxygen species (ROS) levels, and sperm DNA fragmentation (SDF) were evaluated.
Results
The cryopreserved spermatozoa had significantly reduced percentages of motility (p<0.05) and viability (p<0.05). Antioxidant supplementation non-significantly improved these parameters (p>0.05). No significant differences were found in sperm morphology between the fresh and frozen-thawed groups (p>0.05). After freezing, the extracellular ROS levels in the frozen-thawed groups were significantly higher (p<0.05) than in the fresh group. However, we did not find any differences in intracellular ROS parameters among these groups (p>0.05). The SDF was higher in the SuV and Vapor groups than in the fresh group, but without statistical significance (p=0.075 and p=0.077, respectively).
Conclusion
Cryopreservation had detrimental effects on sperm motility, viability, and extracellular ROS levels, without changing the morphology or intracellular ROS levels. Antioxidant supplementation was slightly effective in preventing SDF in frozen-thawed spermatozoa.

Keyword

Human spermatozoa; Reactive oxygen species; Sperm chromatin structure assay; Sucrose vitrification; Triple antioxidant