Mechanism Of Early Embryonic Death Induced By Intrauterine Bubbles In Mice

Embryo transfer technique is the last step in embryo engineering and plays a decisive role in embryo pregnancy rate.In mice,cattle,sheep and human embryos,there is usually a certain amount of air being transplanted into the uterus with embryos.Whether these bubbles affect the outcome of embryo transfer is still unclear.Studies have shown that in guinea pig,rat,hamster,and other rodents,when estradiol and progesterone level in certain conditions,intrauterine air can induce decidualization of endometrium.At the same time,the results of mouse embryo transfer showed that the intrauterine bubbles could lead to early embryo death.Decidualization is the decisive factor for the embryos successful implantation and development.So in order to explore the effect of the intrauterine bubble on the embryo development,we transfered air into the uterine of pseudopregnancy 3.5d mice and normal pregnancy 3.5d mice,and collected 7.5d decidual tissue,The mechanism was studied by morphological observation,endometrial permeability test,tissue section,2-DE,RNA-Seq and RT-PCR verification.The results showed that the intrauterine bubbles could induce the uterus horns obviously thickening,weight increase,increased diameter,and obvious vascular remodeling,large number of polykaryocytes appearance,irregular distribution of all kinds of cells,which could not distinguish the primary and secondary decidua regions,massive death of embryos in the horn of the uterus.The results of image analysis of 2-DE showed that there was no protein difference between decidua induced by intrauterine bubble and normal embryo induced decidua within the detected range.The results of RNA-Seq showed that there were 64 differentially expressed genes between the two groups,of which 59 differentially expressed genes were highly expressed in the normal decidua tissue.The highest 10 expressed genes among differentially expressed genes ware H19,Prl7a1,Prl3d1,Prl4d1,Prl2c3,Krt8,Mmp9,Prl2c2,Plet1,Cts7.GO analysis of differentially expressed genes showed that the differentially expressed genes were mainly enriched in placental development,trophoblastic giant cell differentiation,cell response to interleukin 1,cell response to tumor necrosis factor,and regulation of growth hormone activity,In the process of cell protein decomposition,protein hydrolysis and other biological processes,the enriched genes are highly expressed genes in normal decidual tissue.The results of RNA-Seq were verified by RT-PCR.The results showed that the expression patterns of these 8 genes was consistent with the results of RNA-Seq.The results showed that the bubbles in the uterus led to abnormal decidualization in the endometrium,and the gene expression pattern changed,which affected the normal development of the embryo and eventually led to the death of the embryo.These results are not only helpful to improve the success rate of mouse embryo transfer,but more importantly,it provides new ideas for the research of human and other animal embryo transfer.