Deciphering the dialogue between the bovine blastocyst and the uterus:embryo-induced alterations in extracellular vesicle protein content from an ex vivo model and the in vivo environmentOA

Backgroud Efficient communication between the embryo and the endometrium is essential for the successful establishment and maintenance of pregnancy.Uterine-derived extracellular vesicles(EVs)contribute to embryomaternal communication,supporting early embryonic development.This study aimed to:(i)compare the protein cargo of uterine fluid EVs(UF-EVs)from CYCLIC and PREGNANT heifers;(ii)characterize the protein profile of conditioned medium(CM)-EVs from endometrial explants cultured alone(EXPL)or co-cultured with five d 7 blastocysts(EXPL+EMB)in vitro;and(iii)compare the EV protein cargo between the in vivo and in vitro models(i.e.,EXPL vs.CYCLIC and EXPL+EMB vs.PREGNANT).Results We identified 1,459 and 1,752 proteins in the UF-EVs of CYCLIC and PREGNANT heifers,respectively.Among these,12 were exclusive to CYCLIC,and 18 were exclusive to PREGNANT.Among the 1,329 proteins identified in both groups,16 were differently abundant;ten were more abundant,and six were less abundant in UF-EVs from PREGNANT heifers.In vivo,the changes in UF-EV protein cargo induced by the presence of a blastocyst were related to inflammatory and immune responses,endometrial receptivity,and support of early embryonic development by promoting cell polarity,cell–cell adhesion,and stem cell differentiation.In vitro,we identified 1,501 proteins in the CM-EVs from EXPL,1,975 in the CM-EVs from EXPL+EMB,and 82 in the CM-EVs from EMB.Additionally,50 proteins were unique to EXPL+EMB,and another 33 were differentially abundant due to the synergistic interaction between the embryo and the endometrium.These proteins are involved in embryonic development,regulation of stem cell differentiation,establishment and maintenance of cell polarity,interferon tau(IFNT)-mediated cell signaling,endometrial receptivity,and immune modulation.Although there are qualitative and quantitative differences between in vivo and in vitro-derived EVs,UF-EVs from CYCLIC heifers compared to CM-EVs from EXPL,as well as UFEVs from PREGNANT heifers compared to CM-EVs from EXPL+EMB shared common proteins.Conclusions These findings highlight the pivotal role of EVs in embryo-maternal communication,suggesting that their protein cargo may actively contribute to the modulation of the uterine environment to support early embryonic development.Understanding these molecular interactions could provide valuable insights into the mechanisms of implantation and pregnancy establishment.