The Study Of Molecular Mechanisms Underlying The Efects Of Certain Protective Agents On Boar Sperm Preserved At 4 ℃

In modern pig husbandry,artificial insemination(AI)is extensively used throughout the world.Numerous studies have shown that the semen used for AI must be diluted with suitable extenders and should be preserved at 17 ℃,4 ℃ or in liquid nitrogen.Thus,with the goals of avoiding the freezing injury of boar sperm preserved in liquid nitrogen and prolonging the survival time in vitro,many researchers are committed to the study of liquid storage at 4 ℃.Due to a low cholesterol/phospholipid ratio,boar spermatozoa are particularly vulnerable to low temperatures,which can lead to oxidative stress caused by the inappropriate formation of ROS.At present,little is known about the protective mechanisms of antioxidants towards ROS toxicity in sperm preserved at low temperatures.The main work of this research is as follows:Firstly,we investigated the underlying molecular mechanisms of the protective effects of BSA and sikm milk(SM)on boar sperm and found that BSA and SM probably first induced sperm protein modification changes and then enhanced mitochondrial membrane potential,subsequently increasing intracellular ATP content and improving sperm motility.Moreover,there was no significant difference between the protective effects of BSA and SM in boar semen.Thus,the present study was expected to provide a theoretical basis for the comprehensive understanding of the protective mechanisms of BSA and SM from the perspective of energy metabolism and protein modification.Hopefully,our research suggests a practical feasibility for using SM instead of BSA as a boar extender supplement and will reduce production costs.Secondly,to gain further insight into the protective effects of APS,MLT and CNMs on boar sperm,the present study was performed to assess the changes in sperm quality parameters,antioxidant capacity,ATP content and protein phosphorylation levels.Here,we demonstrated that supplementation with APS,MLT and CNMs could effectively preserve boar sperm quality parameters such as sperm motility and mitochondrial membrane potential.Moreover,we found that the positive effects of APS,MLT and CNMs on boar sperm quality were mainly due to the elimination of excessive mitochondrial ROS,the improvement of antioxidant capacities and the enhancement of ATP levels.Finally,by conducting a series of studies on protein phosphorylation,we also discovered that APS,MLT and CNMs could protect boar sperm from oxidative stress and energy deficiency through inhibiting the protein dephosphorylation caused by ROS via the c AMP-PKA signaling pathway.In a word,this is the first exploration of the molecular mechanism underlying the protective roles of APS,MLT and CNMs towards ROS toxicity from the perspective of oxidative stress and protein modification.This study comprehensively provides novel insights into the action mechanism of the protective effects of antioxidants on sperm stored at 4 ℃ and reveals the practical feasibility of using APS,MLT and CNMs as a boar semen extender supplement for assisted reproductive technology.