Sperm DNA Fragmentation and Apoptosis Levels: A Comparison of the Swim up and the Density Gradient Centrifugation Methods for Sperm Preparation

Abstract

This study investigated the efficiency of two sperm preparation methods, namely, density gradient centrifugation and the swim-up method, in selecting spermatozoa based on the DNA fragmentation index and the apoptosis levels. Fifteen semen samples from infertile couples underwent sperm preparation in this study. The Makler ® counting chamber and Eosin Y staining were used to analyse the sperm concentration, motility and viability. The sperm chromatin dispersion assay was used to determine the sperm DNA fragmentation index, while western blotting was used to determine the apoptosis levels. This study showed that Swim Up (SU) and Density Gradient Centrifugation (DGC) methods were able to select sperm with lower DFIs compared to sperm from whole semen. The SU method selected for sperm in both the "moderate" and "severe" DFI categories that have low DFIs better than the DGC method. Western blotting for the expression of caspase 3 in sperm showed a band of ∼35 kDa and this band's intensity was lower in post-SU sperm compared to post-DGC sperm. Both methods selected for sperm with low caspase 3 expression, although the SU method selected for sperm with low activity of caspase 3 better than the DGC method. In addition to selecting for sperm with better progressive motility and viability, the SU and DGC methods also selected for sperm with low sperm DFIs and apoptosis levels.

Sperm Na+, K+-ATPase and Ca2+-ATPase activity: A preliminary study of comparison of swim up and density gradient centrifugation methods for sperm preparation

“…It means too that this study confirms the result of Almeida et al [28]. In addition, Lestari SW et al also demonstrated that the sperm DNA fragmentation level was lower in the SU method compared to the DGC method, which was in line with the result of this study [29]. Furthermore, the results of this study were in agreement with the previous study that showed the lower level of caspase 3 expression as a representative of the apoptotic status in prepared sperm compared to the whole semen, even though the SU method was showed the lower level compared to the DGC method [29].…”

“…In addition, Lestari SW et al also demonstrated that the sperm DNA fragmentation level was lower in the SU method compared to the DGC method, which was in line with the result of this study [29]. Furthermore, the results of this study were in agreement with the previous study that showed the lower level of caspase 3 expression as a representative of the apoptotic status in prepared sperm compared to the whole semen, even though the SU method was showed the lower level compared to the DGC method [29]. Studies have shown that the amount of caspase 3 expression of sperms in fertile and infertile men was decreased after DGC methods, related to the deficit of immature sperm with higher expression of caspase 3.…”

Sperm quality after swim up and density gradient centrifugation sperm preparation with supplementation of alpha lipoic acid (ALA): A preliminary study

“…(Fig.2) On the contrary, in the 'moderate' and 'severe' category, the DFI of prepared sperm was lower compared to the whole semen. In contrast to the 'mild' category, this finding did not support the result by Jayaraman et al [3], but promoted the result by Lestari et al [6]. This could be explained by the existence of more sperms with DNA damage in the 'moderate' and 'severe' category of sperm sample, than the 'mild' category.…”

“…In addition, studies also reported that ALA enhances sperm ability to tolerate higher volumes of free radical attack, thus protecting the DNA integrity of the sperm [5]. Moreover, ALA also has proven in decreasing the sperm DNA fragmentation, related to the production of ROS [6].…”

Sperm DNA Fragmentation Index: A Comparison Study of Success Rates among Natural, Intrauterine Insemination (IUI) and In Vitro Fertilization (IVF)-Intra Cytoplasmic Sperm Injection (ICSI) Pregnancy Programs

“…In In addition, sperm in the semen that underwent washing before ART in IUI and IVF-ICSI programs underwent a selection process to obtain sperm with more intact DNA or less fragmented DNA than before washing, so sperm of higher quality was used for IUI and IVF-ICSI programs. 15…”