Proteomic Analysis Of Exosomes In Pig Pregnancy Of Uterine Cavity Fluid

During the implantation of pig embryos,there is a close signal communication between the embryo and the mother,and many regulatory factors have complex and coordinated changes in the process of pregnancy recognition,thus promoting the synchronous response of the embryo,endometrium and uterine microenvironment.The signal communication between the embryo and the mother requires the signal transmission of related factors in the uterine cavity fluid.In recent years,exosomes,as communication media between cells,play an important role in the microenvironment of the reproductive system.However,it has not been reported whether exosomes and the types of proteins are present in the uterine cavity fluid during the early gestation period in pigs.Therefore,the analysis and identification of the components and functions of exosomes in the uterine cavity of pig in early pregnancy.laying a foundation for the discovery of the important factors regulating the implantation of pig embryos and provided a new idea for the improvement of the success rate of embryo implantation.In this study,uterine cavity exosomes were isolated and identified,and i TRAQ quantitative proteomics method was used to quantify and analyze the uterine cavity exosome proteins in pure white sows during estrus 9 days,gestation 9 days,gestation 12 days and gestation 15 days,and the following results were obtained:(1)TEM results showed that the exosomes of uterine cavity were membraniform vesicles,and WB identification results showed that the exosome marker CD63 was positively expressed.Particle size analysis showed that the diameter of exosomes from uterine cavity fluid was between 100-150 nm.(2)The analysis of iTRAQ proteomic and quantitative biological information showed that the expression patterns of exosome proteins in uterine cavity at 9 days of estrus and 9 days of gestation were similar,and the expression patterns of exosome proteins in uterine cavity at 12 days of gestation and 15 days of gestation were similar.(3)Western Blot analysis showed that ACP5 and MEP1 B proteins were present in four stages of uterine cavity exosomes.The immunohistochemical results of four stages of endometrial sections showed that ACP5 and MEP1 B proteins were expressed on the surface of the endometrium and the trophoblast of pig embryos.(4)MEP1B protein was expressed in the pig endometrium at 9 days of estrus,9 days of gestation,12 days of gestation and 15 days of gestation,and it increased with the increase of gestation time.Besides,the si RNA of MEP1 B protein had an inhibitory effect on the activity of pig endometrium cells and embryonic trophoblast cells.In conclusion,There are exosomes in the uterine cavity fluid of pigs,and the expression of exosome proteins is different at different periods of estrus and early pregnancy,so MEP1 B protein is probably related to embryo implantation.