Effect Of Punicalagin On Boar Semen Quality During Room Temperature Preservation

Since 1980 s,the application of artificial insemination(AI)in pig breeding and reproduction has gained worldwide recognition and has laid a solid foundation for the rapid development of the pig industry.The preservation of semen is a key technique of artificial insemination,and the quality of semen preservation is a key factor to ensure the success rate of AI.Due to the high proportion of polyunsaturated fatty acids(PUFAs)in the plasma membrane,the pig sperm is relatively sensitive to low temperature and reactive oxygen species(ROS),which determines that the sperm should be preserved in liquid environment in room temperature.Excess ROS produced during sperm storage at normal temperature causes oxidative damage to sperm acrosome and plasma membrane,reduce motility and exercise capacity of sperm,resulting in sperm death,which finally lead to decreased semen preservation quality and artificial insemination;Furthermore,the bacterial contamination during room temperature preservation also causes sperm death.In summary,sperm oxidative damage and bacterial contamination are the main problems are needed to be solved in the current preservation of pig semen at room temperature.In response to the above problems,with the hope to reduce oxidative stress and bacterial contamination and prolong preservation time,the pomegranate extract punicalagin was added to Beltsville Thawing Solution(BTS)to treat pig sperm at the room temperature.What's more,this study also serves as a theoretical underpinning for research and development of preservation solution at room temperature in China.The main results for this study are as follows:1.Punicalagin supplement relieved oxidative damage by removing ROS,resulting in increased sperm motility.The pig sperm was stored in BTS with different concentrations(0,10,15,and 20 ?mol/L)of punicalagin respectively at room temperature for 7 days.The results showed that sperm motility,acrosome integrity,and plasma membrane integrity were significantly increased with 10,15 and 20 ?mol/L punicalagin supplement(P < 0.05),peaking at 15 ?mol/L(P < 0.01).Moreover,the contents of ROS and malondialdehyde(MDA)in sperm were significantly reduced at all punicalagin supplement groups(P < 0.05),accompanied with increased total antioxidant capacity(T-AOC)(P < 0.05),and 15 ?mol/L punicalagin treatment benefited most(P <0.01).2.Punicalagin inhibited the growth of bacteria in pig sperm.The data demonstrated that the number of bacteria in 15 ?mol/L punicalagin supplement group was significantly lower than that in the control group(P < 0.05),especially for the two dominant species in sperm preservation solution.3.The punicalagin enhanced pig sperm movement performance by activating the sperm AMPK pathway,increasing energy supply.The sperm cell variability was measured after 7-day preservation at room temperature.The data showed that punicalagin supplement,especially with the concentration of 15 ?mol/L,inproved sperm movement,including Progressive(PRO),Straight Linear Velocity(VSL),Curvilinear Velocity(VCL),Average Path Velocity(VAP)(P < 0.05).In addition,the ATP content in all punicalagin supplemented groups were significantly higher than that in the control group(P < 0.01).Furthermore,the mRNA and protein levels of the mitochondrial respiratory chain-related genes NRF-1 and PGC-1 increased significantly(P < 0.05),with significant increase of the phosphorylation level of AMPK(P < 0.05).In summary,this study demonstrated that the supplement of 15 ?mol/L punicalagin improved oxidative stress via increasing oxidative damage,activated the AMPK signaling to enhance exercise performance and inhibit bacterial growth,improving pig sperm preservation at time temperature.