ABSTRACT
Objective: A recent innovative approach, based on induction of sublethal oxidative stress to enhance sperm cryosurvival, has been applied before sperm cryopreservation. The purpose of this study was to investigate the effects of different induction times of sublethal oxidative stress before cryopreservation on human post-thawed sperm quality
Materials and Methods: In this experimental study, we selected semen samples [n=20] from normozoospermic men according to 2010 World Health Organization [WHO] guidelines. After processing the samples by the density gradient method, we divided each sample into 5 experimental groups: fresh, control freezing, and 3 groups exposed to 0.01M sodium nitroprusside [SNP] [nitric oxide [NO] donor] for 30 [T30], 60 [T60], or 90 minutes [T90] at 37C and 5% CO2 before cryopreservation. Motion characteristics [computer-assisted sperm analyser], viability, apoptosis [annexin V/propidium iodide [PI] assay], DNA fragmentation [sperm chromatin structure assay [SCSA]], and caspase 3 activity [FLICA Caspase Detection Kit] were assessed after thawing. The results were analysed by using one-way ANOVA and Tukey's test. The means were significantly different at P<0.05
Results: Cryopreservation significantly decreased sperm viability and motility parameters, and increased the percentage of apoptosis, caspase 3 activity, and DNA fragmentation [P<0.01] compared to the fresh group. The T60 group had a higher significant percentage of total motility [TM] and progressive motility compared with other cryopreserved groups [P<0.05]. We observed a significantly lower percentage of apoptotic rate and caspase 3 activity in the T60 group compared to the other cryopreserved groups [P<0.05]. DNA integrity was not significantly affected by this time of sublethal stress induction [P>0.05]
Conclusion: Our results have demonstrated that the application of sublethal oxidative stress by using 0.01 MuM NO for 60 minutes before the freezing process can be a beneficial approach to improve post-thawed human sperm quality
ABSTRACT
Objective: Pluripotent stem cells [PSCs], with the capacity to self-renew and differentiate into all other cell types, are of benefit in regenerative medicine applications. Tightly controlled gene expression networks and epigenetic factors regulate these properties. In this study, we aim to evaluate the metabolic signature of pluripotency under 2i and R2i culture conditions versus serum condition
Materials and Methods: In this experimental study, we investigated bioinformatics analysis of the shotgun proteomics data for cells grown under 2i, R2i, and serum culture conditions. The findings were validated by cell cycle analysis and gene expressions of the cells with flow cytometry and quantitative reverse transcription-polymerase chain reaction [qRT-PCR], respectively
Results: Expressions of 163 proteins increased in 2i-grown cells and 181 proteins increased in R2i-grown cells versus serum, which were mostly involved in glycolysis signaling pathway, oxidation-reduction, metabolic processes, amino acid and lipid metabolism. Flow cytometry analysis showed significant accumulation of cells in S phase for 2i [70%] and R2i [61%] grown cells
Conclusion: This study showed that under 2i and R2i conditions, glycolysis was highlighted for energy production and used to maintain high levels of glycolytic intermediates to support cell proliferation. Cells grown under 2i and R2i conditions showed rapid cell cycling in comparison with the cells grown under serum conditions