The aim of this study was to evaluate the effect of different extenders on the quality of European red deer epididymal sperm stored at 5 °C. Epididymal spermatozoa were collected post mortem from 10 stags and diluted with three extenders (Bovidyl^®, BoviFree^®, and BioXcell^®) and stored at 5 °C. Sperm motility (TMOT), motility parameters (system CASA), plasma membrane integrity (SYBR-14⁺/PI⁻), acrosomal membrane integrity (FITC-PNA⁻/PI⁻), mitochondrial activity (JC-1/PI), viability, and apoptotic-like changes (YOPRO/PI) were evaluated. The analyses were conducted on the first and successive days of storage (D1–D7). The applied extender, storage time, and the interactions between these factors significantly (p < 0.001) affected most of the analyzed parameters whose values were highest in sperm samples stored in Bovidyl^®, regardless of storage time. In Bovidyl^®, BoviFree^®, and BioXcell^® extenders, TMOT values were estimated at 83%, 63%, and 59%, respectively, on D3. The extenders significantly influenced DNA integrity on D7. The percentage of dead sperm increased from D3. The quality of stored sperm cells was significantly influenced by the extenders’ biochemical composition. BoviFree^® and BioXcell^® contain glycerol which could contribute to deteriorating the quality of spermatozoa stored at 5 °C. Sperm cells stored in the egg yolk-based extender (Bovidyl^®) were characterized by the highest viability and functionality.

The aim of this study was to assess the quality and fertilizing potential of red deer epididymal spermatozoa stored in a liquid state for up to 11 days (D11). In Experiment 1, sperm quality was determined. In Experiment 2, the efficiency of in vitro fertilization (IVF) and artificial insemination (AI) of stored sperm were evaluated. An analysis of sperm quality on D5 of storage revealed a decrease (p < 0.05) in motility and morphology, and a higher proportion of apoptotic spermatozoa. On D1, D7 and D10, the total motility of sperm for IVF and AI was determined to be 82.6%, 71.0% and 64.8%, respectively. The results of IVF and AI demonstrated that the fertilizing potential of spermatozoa differs between days of storage. The percentage of blastocysts was higher when oocytes were fertilized on D1 (17.4 %) compared to D7 (8.5%) and D10 sperm (10.5%). Differences were noted in the pregnancy rates of inseminated hinds. The insemination with D1, D7 and D10 sperm led to live births (33% from D7 and D10). The results indicate that the quality of red deer epididymal spermatozoa remains satisfactory during ten days of storage in a liquid state, and that these spermatozoa maintain their fertility potential.

The aim of this study was to analyze the influence of season (rut and non-rut) on the antioxidant status of selected reproductive tissues in male European red deer (Cervus elaphus elaphus). Tissue samples were collected post mortem from the testes and epididymides (caput, corpus, and cauda) of 24 animals. The activity of antioxidant enzymes (superoxide dismutase—SOD, glutathione peroxidase—GPx, and catalase—CAT) and the mRNA expression of SOD1, SOD2, SOD3, GPx4, GPx5, and CAT were examined. In addition, these proteins were identified by western blot. ANOVA revealed that season, type of tissue, and the interaction between these factors significantly (p ≤ 0.05) influenced the activity and mRNA expression of the analyzed enzymes. The activity of SOD and GPx peaked in the corpus epididymis in the rut season and in the caput epididymis in the non-rut season. Regardless of season, the relative abundances of GPx4, SOD1, SOD2, and SOD3 mRNA were highest in the testis, and GPx5 mRNA—in the caput epididymis. The activity of SOD and CAT was significantly higher during the non-rut season compared with the rut season, but only in the caput epididymis. This study demonstrated that the activity of antioxidant enzymes and the relative mRNA expression varies across tissues and seasons to provide the reproductive system of European red deer with the required antioxidant protection. Further research is needed to expand our understanding of the antioxidant defense system in the reproductive tract of European red deer.

The aim of this study was to evaluate the motility, morphology, and antioxidant status of European red deer sperm stored in a liquid state (variant I) and in the epididymides (variant II). Spermatozoa were harvested post-mortem from the cauda epididymis. Sperm samples in both variants were stored for up to six days (D6) at 5 °C. Spermatozoa were assessed for motility, viability, morphology, activity of antioxidant enzymes (superoxide dismutase, SOD; glutathione peroxidase, GPx; catalase, CAT), and lipid peroxidation (malondialdehyde, MDA, content). Sperm samples were analyzed on storage days 0, 2, 4, and 6 (D0-D6). Storage time and storage method significantly (p ≤ 0.05) influenced the examined variables. On D2, a decrease in motility and acrosomal integrity was observed in both storage variants, whereas a decrease in viability and an increase in MDA content were noted in spermatozoa stored in the epididymides. On D4, higher values of SOD and GPx activity and MDA content were noted in variant I than in variant II. Catalase activity was very low. GPx is the key enzyme that participates in the reduction of hydrogen peroxide in sperm cells. Spermatozoa stored in a liquid state were characterized by higher motility and viability, improved morphology and antioxidant status than those stored in the epididymides; therefore, liquid storage is more recommended for short-term preservation of epididymal spermatozoa.