Exogenous spermidine effectively improves the quality of cryopreserved boar sperm.

Boar sperm are less resistant to the dramatic environmental changes that occur during in vitro preservation. Spermidine has various physiological functions including the anti-oxidative effect. The main objective of this study was to clarify whether spermidine could protect boar sperm from the attack of reactive oxygen species under cryopreservation treatment. We set the concentrations of spermidine at 0, 2, 4, 6, and 8 mmol/L and evaluated the effects of spermidine on sperm motility, viability, malformation rates, kinetic parameters, membrane integrity, mitochondrial activity, DNA integrity, H2 O2 content, malondialdehyde content, total antioxidant capacity, and antioxidant enzyme activity. Finally, the effects of spermidine on the sperm fertility were assessed by artificial insemination. The results showed that spermidine improved various physiological parameters of sperm in a dose-dependent manner. The quality and antioxidant capacity of sperm cryopreserved with 6 mmol/L spermidine were significantly less reduced (P < 0.05), and the contents of malformation rate, H2 O2 , and malondialdehyde content were significantly decreased (P < 0.05). The significant increase in the number of litters indicated the possibility that spermidine had important practical value in pig reproduction (P < 0.05). Therefore, the addition of appropriate concentrations of spermidine to cryopreservation extenders may effectively improve the quality of boar sperm for in vitro preservation.

Phosphocreatine addition to extender enhances the quality and antioxidant capacity of cryopreserved boar sperm.

Boar sperm are less resistant to drastic changes in the external environment during cryopreservation, mainly because their plasma membranes are rich in unsaturated fatty acids but lack cholesterol and are thus susceptible to lipid peroxidation caused by the attack of reactive oxygen species. This study evaluated the effect of adding phosphocreatine to cryopreservation extenders on boar sperm quality and antioxidant capacity. Different concentrations (0, 5.0, 7.5, 10.0 and 12.5 mmol/L) of phosphocreatine were added to the cryopreservation extender. After thawing, sperm were analysed for morphological parameters, kinetic parameters, acrosome integrity, membrane integrity, mitochondrial activity, DNA integrity and antioxidant enzyme activity. The results showed that 10.0 mmol/L phosphocreatine samples enhanced the boar sperm motility, viability, average path velocity, straight-line velocity, curvilinear velocity and beat cross frequency after cryopreservation and reduced the malformation rate compared to the control group (p < .05). The acrosome integrity, membrane integrity, mitochondrial activity and DNA integrity of boar sperm were higher than those of the control group after adding 10.0 mmol/L phosphocreatine to the cryopreservation extender (p < .05). Extenders containing 10.0 mmol/L phosphocreatine maintained high total antioxidant capacity; elevated the activities of catalase, glutathione peroxidase and superoxide dismutase; reduced malondialdehyde and H2 O2 content (p < .05). Therefore, adding phosphocreatine to the extender is potentially beneficial for boar sperm cryopreservation at an optimal 10.0 mmol/L concentration.

Methylprednisolone improves the quality of liquid preserved boar spermatozoa in vitro and reduces polymorphonuclear neutrophil chemotaxis and phagocytosis.

After artificial insemination, immune cells such as polymorphonuclear neutrophils will be recruited into the genital tract and induce endometrial inflammation, adversely affecting the spermatozoa. This study aimed to analyze the effect of methylprednisolone (MPS) on boar spermatozoa quality of in vitro liquid preservation and chemotaxis and phagocytosis of polymorphonuclear neutrophils toward boar spermatozoa. Various concentrations of MPS were added to the extender and analyzed for their effects on spermatozoa motility, kinetic parameters, abnormality rate, total antioxidant capacity (T-AOC) levels, H2O2 content, mitochondrial membrane potential and acrosome integrity. Testing of MPS on chemotaxis and phagocytosis of polymorphonuclear neutrophils toward spermatozoa induced by lipopolysaccharide (LPS). The results showed that an extender containing 2 × 10-7 mol/mL MPS was the most effective for preserving boar spermatozoa during in vitro liquid preservation at 17°C. It effectively improved spermatozoa motility, kinetic parameters, T-AOC levels, mitochondrial membrane potential and acrosome integrity, reducing the abnormality rate and H2O2 content. Meanwhile, the chemotaxis and phagocytosis of polymorphonuclear neutrophils toward spermatozoa under LPS induction were inhibited in a concentration-dependent manner. In conclusion, MPS has positive implications for improving in vitro liquid preserved boar spermatozoa quality, inhibiting chemotaxis and phagocytosis of polymorphonuclear neutrophils toward spermatozoa.

Dexamethasone affects the chemotaxis and phagocytic activity of neutrophils for boar spermatozoa and the quality of liquid preserved boar semen in vitro.

This study aimed to investigate the effect of dexamethasone (DEX) on chemotaxis and phagocytic activity of neutrophils for spermatozoa and semen quality of preserved boar semen. The different concentrations of dexamethasone were added to boar semen dilutions to detect its effects on the chemotaxis of neutrophils and phagocytosis of neutrophils and sperm motility sperm malformation rate, plasma membrane integrity, mitochondrial activity, and spermatozoa motility parameters. The study results showed that the experimental groups of DEX significantly inhibited the phagocytosis and chemotaxis of PMNs for spermatozoa. With the increased concentration of DEX, there was an inhibition effect on PMNs activity. In addition, under 17 °C storage conditions, the addition of DEX 1 × 10-6 mol/mL concentration has the best preservation effect on boar semen, which can effectively improve the sperm motility, movement parameters, plasma membrane integrity, mitochondrial activity, T-AOC activity, and significantly reduce the sperm malformation rate and H2O2 content. Therefore, the most suitable concentration of DEX to preserve boar semen at 17 °C is 1 × 10-6 mol/mL. The significant increase of conception rate of sows and litter size of piglets proved that DEX has practical application value. Thus, it is shown that the use of DEX to inhibit uterine inflammation is effective and feasible for sperm damage providing new methods for developing low-dose artificial insemination technology.

Separation of motile sperm for in vitro fertilization from frozen-thawed bull semen using progesterone induction on a microchip.

This study presents a novel method for the separation of motile sperm from non-progressive motile and immotile sperm and in vitro Fertilization (IVF). This separation of bull sperm was accomplished by inducing chemotaxis along a progesterone release agent in a 7.5-mm microchannel microchip composed of a biocompatible polydimethysiloxane layer and a glass gradient. The selected sperm was applied directly for IVF. In the first experiment, we tested the effect of different lengths of microchannnel (5mm, 7.5mm and 10mm) on quality parameter of separated sperm. The results showed that separated sperm using 7.5-mm microchannel chip were improved in sperm motility, swimming velocity, and beat frequency compared with other groups. In the second experiment, a medium containing sperm from swim-up method and outlet reservoir of our 7.5-mm microchannel chip was collected and mitochondrial activity of the sperm was determined by fluorescence microscopy. The sperm from the microchip had higher mitochondria activity (47.6%±6.0%) than the sperm from the swim-up method (23.6%±4.7%) (P<0.05). There were significant differences in rate of acrosome intactness between the swim-up method and the microchip (36.0%±4.1% vs. 66.8±2.1%, respectively, P<0.05). In the third experiment, we compared sperm penetration in the microchip-IVF system with a standard IVF method (droplet-IVF). The microchip-IVF group had the highest percentages of oocytes penetrated (82.2%±1.6% vs. 63.5%±2.4%) and monospermic oocytes (67.8%±3.4% vs. 42.4%±1.5%). In addition, early developmental competence of oocytes to the blastocyst stage was higher when the oocytes were inseminated in the microchip-IVF system compared with those inseminated in a standard droplet-IVF system. These results demonstrate that our microchip based on a sperm chemotaxis system is useful for motile sperm separation from frozen-thawed bull semen for IVF. Therefore, the optimized microchip system provides a good opportunity to sort motile bull sperm for IVF.