Investigation of the protective effects of different forms of selenium in freezing dog semen: Comparison of nanoparticle selenium and sodium selenite.

Esin B ，Kaya C ，Akar M ，Çevik M ... -  《-》

Antioxidant effects of supplementation of 3,4-dihydroxyphenyl glycol on sperm parameters and oxidative markers following cryopreservation in canine semen.

Supplementing the extender with antioxidants with low molecular weight can enhance the quality of the post-thaw sperm during the freezing process. This study was aimed at determining the impacts of 3,4-dihydroxyphenyl glycol (DHPG) on the spermatozoa of the canine undergoing freeze-thawing process. In this study, 24 ejaculates were obtained from three mixed-breed dogs and were diluted in a Tris-based extender. The diluted semen was divided into aliquots for supplementation of 10, 30, 50 and 70 µg/ml of DHPG, control (without antioxidant) and control sham (DMSO). After being extended, the semen was equilibrated at a temperature of 4°C and then transferred to the straws and kept 4 cm above the liquid nitrogen for 20 min and was finally immersed in the liquid nitrogen. They were cryopreserved for seven days; then, sperm parameters including sperm motility evaluation, motility characteristics, viability, DNA and plasma membrane integrity, total antioxidant capacity (TAC), reduced glutathione content (GSH), antioxidant enzymes (superoxide dismutase [SOD], catalase [CAT] and glutathione peroxidase [GPx]) activity malondialdehyde (MDA) levels were evaluated. This study showed that spermatozoa cryopreservation with 50, 30 and 70 µg/ml of DHPG concentrations had better progressive motility, Curvilinear Velocity, Linearity, viability, intact plasma membrane and the levels of TAC, GPx and GSH were higher than the control group. The 50, 30 and 70 µg/ml of DHPG concentrations led to the significant decrease of DNA damage compared to the control group. Total motility, average path velocity, straight-line velocity and CAT activity were significantly improved in 30 and 50 µg/ml of DHPG concentrations, compared to the control group. Also, the 50 and 30 µg/ml of DHPG concentrations, decreased MDA levels compared to the other groups, significantly. In conclusion, our study showed that the addition 50 µg/ml of DHPG to the canine semen extender improved the semen characteristics and oxidative markers in the cryopreservation process.

Shakouri N ，Soleimanzadeh A ，Rakhshanpour A ，Bucak MN ... -  《-》

Cryopreservation of epididymal cat spermatozoa: effects of in vitro antioxidative enzymes supplementation and lipid peroxidation induction.

Reactive oxygen species and lipid peroxidation reaction, causes of sperm damage, can be diminished by action of antioxidative enzymes. This study aimed to investigate effects of (1) the antioxidative enzymes; catalase, glutathione peroxidase and superoxide dismutase, on epipididymal cat sperm quality and (2) the lipid peroxidation reaction induced by a transition metal (ferrous ion (II); Fe(2+)) on sperm quality during the cryopreservation process. Epididymal spermatozoa harvested from 39 male cats were pooled and divided into 13 aliquots (n=13). Each aliquot was resuspended with either a Tris egg yolk extender I (control; EE-I), or the Tris egg yolk extender I supplemented with 200 U/mL catalase (EE-CAT), or 10 U/mL glutathione peroxidase (EE-GPx), or 600 U/mL superoxide dismutase (EE-SOD), and then cryopreserved. After thawing, each sperm sample was subdivided into two groups; with and without lipid peroxidation induction (EE-I plus Fe(2+), EE-CAT plus Fe(2+), EE-GPx plus Fe(2+) and EE-SOD plus Fe(2+)). Subjective sperm motility, membrane, and acrosome integrity were evaluated at the time of collection, after cooling, and at 0, 2, 4, and 6h after thawing. Motility patterns assessed by computer-assisted sperm analysis (CASA), mitochondrial activity, and DNA integrity were evaluated during post-thaw incubation, whereas percentage of lipid peroxidation was detected at 0 and 6h after thawing. The results demonstrate that catalase supplementation reduced linear motility and subjective motility immediately and 2h after thawing (P<0.05). Catalase supplementation, however, improved DNA integrity at 4h (P<0.05). Supplementation with glutathione peroxidase, compared to the control group, had a statistically significant positive effect on subjective motility at 0 and 6h, linear motility at 6h, mitochondrial activity at 6h, membrane integrity at 2 and 6h, and DNA integrity at 4h after thawing. Although superoxide dismutase had a positive effect on sperm membrane integrity at 2h after thawing (P<0.05), it significantly reduced membrane integrity after cooling, linear motility at thawing, and acrosome integrity at 2h after thawing. None of the three selected antioxidative enzymes significantly influenced acrosome integrity and none reduced the level of lipid peroxidation. Furthermore, induction of the lipid peroxidation reaction by Fe(2+) negatively affected most of the sperm quality parameters, i.e., motility and DNA integrity, during post-thaw sperm incubation (P<0.05). After thawing, there were, however, no significant differences between the control plus Fe(2+) and the antioxidative enzymes supplementation plus Fe(2+) groups. We can conclude that (1) glutathione peroxidase exhibits positive effects on post-thaw epididymal cat spermatozoa; but (2) none among the selected antioxidative enzymes could improve all sperm quality parameters; and (3) the lipid peroxidation reaction may be one cause of post-thaw epididymal sperm damage in cats, but the concentrations of antioxidative enzymes used in this study could not protect cat spermatozoa from lipid peroxidation induction.

Thuwanut P ，Chatdarong K ，Johannisson A ，Bergqvist AS ，Söderquist L ，Axnér E ... -  《-》

Quercetin supplemented casein-based extender improves the post-thaw quality of rooster semen.

The advantageous influence of quercetin (Q) supplementation in an extender has not yet been evaluated for rooster semen cryopreservation. This research was purposely conducted in order to assess the effect of different quercetin concentrations added into an extender on the sperm quality of the rooster subsequent to a freezing-thawing process. After the freezing-thawing process, spermatozoa quality parameters (membrane functionality, acrosome integrity, motility, viability, and abnormal morphology), endogenous enzymes (SOD, CAT, and GPx), mitochondrial activity, DNA fragmentation index, lipid peroxidation (MDA), and ROS were all evaluated. A total of 75 neat pooled ejaculates (3 ejaculates/rooster) were collected from 25 arbor acres roosters (24 wks) twice a week using abdominal massage technique, then divided into five equal aliquots and diluted with an extender containing different doses of Q (CS-Q) as follows: casein extender without Q (control only), casein extender containing 0.040 mg/mL quercetin (CS-Q 0.040), 0.020 mg/mL quercetin (CS-Q 0.020), 0.010 mg/mL quercetin (CS-Q 0.010), and 0.005 mg/mL quercetin (CS-Q 0.005). Our results depicted that adding to the extender with a 0.010 mg/mL Q enhanced (P < 0.01) sperm motility, membrane function, viability, mitochondrial activity, intact acrosome (P < 0.05), SOD (P < 0.001), CAT, and GPx (P < 0.01) compared to the control group at post-thaw. Compared to the control group and other treatment groups after the freeze-thawing process, the addition of 0.005 mg/mL Q into the extender also showed higher (P < 0.05) improvement in the quality of sperm parameters and a higher (P < 0.01) SOD and CAT but did not improve mitochondrial activity and sperm viability. In addition, there was a lower degree of DNA fragmentation index, lower (P < 0.05) lipid peroxidation and ROS in frozen-thawed sperm treated with 0.010 mg/mL and 0.005 mg/mL Q than in control and the other treatment groups. In addition, 0.020 mg/mL Q supplementation into the extender also reduced DNA fragmentation and improved GPx activity compared to the control group at post-thaw. Different concentrations of Q 0.010 and 0.005 mg/mL added to the extender reduced the percentage of abnormal spermatozoa compared to the other groups. The results of this study showed for the first time that the inclusion of an extender with a suitable quercetin concentration of 0.010 mg/mL improved the post-thawed quality of rooster semen.

Appiah MO ，Li W ，Zhao J ，Liu H ，Dong Y ，Xiang J ，Wang J ，Lu W ... -  《-》

The effects of different levels of catalase and superoxide dismutase in modified Beltsville extender on rooster post-thawed sperm quality.

Oxidative damage of sperm by means of reactive oxygen species generated by the cellular components of semen is one of the main reasons for decreased sperm motility and fertility during the freeze-thawing process. This study was conducted to determine the influence of catalase (CAT) and superoxide dismutase (SOD) on rooster sperm motility, viability and MDA level after freezing and thawing. Semen samples from 10 sexually-mature Ross 308 breeder roosters were collected and pooled, divided into nine equal parts and diluted with modified Beltsville extender containing no antioxidants (control), or supplemented with 50, 100, 200 and 300 μg/mL CAT, or 50, 100, 200 and 300 U/mL SOD. After thawing, sperm motility and motion parameters were assessed using a CASA system. Sperm viability and MDA level were assessed by eosin-nigrosin and MDA test, respectively. The results of this experiment showed that the extender supplemented with 100 and 200 μg CAT, and 50 U SOD had the highest sperm motility (P<0.05) in sperm motility. Also, addition 100, 200 and 300 μg CAT, and 50 U SOD can improve significantly viability after freeze-thaw. Extender supplemented with 100 μg CAT had significantly lower MDA level compared to control and 300 μg CAT. In conclusion, the results of the present study demonstrate that addition of CAT (100 μg/mL) and SOD (50 U/mL) independently have beneficial effect on quality of post-thawed rooster semen.

Amini MR ，Kohram H ，Zare-Shahaneh A ，Zhandi M ，Sharideh H ，Nabi MM ... -  《-》