The Effects of Supplemented Coenzyme Q10 to Extender on the Endoplasmic Reticulum Stress-Related Genes and Sperm Quality Parameters in Cryopreservation of Mouse Spermatozoa.

Abstract

The endoplasmic reticulum (ER) is the organelle responsible for protein folding in the cell. The damage that may occur during the freezing process of the sperm can exceed the protein loading capacity in the ER. Antioxidants, such as coenzyme Q₁₀ (CoQ₁₀), are added to freezing media to protect sperm cells. In this study, the aim was to investigate the expression levels of ER stress-related genes (protein kinase-like ER kinase [PERK], activating transcription factor 4 [ATF4], CCAAT-enhancer-binding-protein homologous protein [CHOP], and nuclear factor erythroid 2-related factor 2 [NRF2]) and quality parameters (viability, motility, acrosome status, and plasma membrane integrity) of mice sperm after freezing with an extender containing CoQ₁₀. Male BALB/c mouse spermatozoa were cryopreserved using a combination of 18% raffinose + 3% skimmed milk and 50 µM CoQ₁₀. The combination of 18% raffinose + 3% skimmed milk without CoQ₁₀ was used as the control group. The results showed that post-thaw sperm motility, viability, plasma membrane integrity, and intact acrosome rates were significantly higher in the CoQ₁₀-supplemented group compared with the control (untreated) group (p < 0.05). The expression of ER stress-related genes was then analyzed to investigate whether CoQ₁₀ attenuates ER stress in frozen-thawed sperm. The results significantly revealed that the addition of 50 µM CoQ₁₀ to the extender increased PERK, ATF4, and CHOP mRNA levels compared with the control group (p < 0.001). Next, NRF2 gene expression was analyzed to investigate whether CoQ₁₀ affects the antioxidant mechanism of post-thaw sperm. It was revealed that the expression of the NRF2 gene significantly increased in the CoQ₁₀ group compared with the control group (p < 0.001). Collectively, these results suggest that the freeze-thaw process induces ER stress in mouse sperm, and the supplementation of CoQ₁₀ to the cryoprotectant agent reduces ER stress-related genes, activates the gene related to the antioxidant defense system, and improves post-thaw sperm quality parameters.