Effects Of Early Pregnancy On NOD-like Receptor And Insulin/AKT/S6K Pathways In The Endometrium Of Sheep

Early pregnancy is the key stage of pregnancy identification and embryo implantation.Most of the embryo loss occurs during this period.A successful pregnancy requires a great deal of information exchange between the endometrium and the fetus,including immune communication,metabolic coordination and hormone collaboration.NOD-like receptors are a class of pattern recognition receptors,and recent studies have shown that they belong to a new class of maternal effect genes.Insulin/AKT/S6 K signal pathway is closely related to the regulation of cell cycle,autophagy,glucose metabolism and protein synthesis.Endometrium is the key place for pregnancy identification and embryo implantation in sheep and a variety of reproductive hormone receptors are involved in the regulation of uterine physiological state.The purpose of this study was to explore the molecular events in early pregnancy by studying the expression of the NODlike receptor,the key nodes of Insulin/AKT/S6 K signal pathway and reproductive hormone receptor in sheep endometrium in early pregnancy.Twenty-four healthy small-tailed Han sheep about 18-month-old were randomly divided into 4 groups,with 6 sheep in each group.After synchronous estrus,three groups’ sheep were artificially inseminated,while the fourth group was not.The endometrium of sheep was collected on the 13 th,16th and 25 th day of pregnancy,and the endometrium of sheep on the 16 th day of non-pregnancy(estrous cycle).The corresponding expression levels of the NOD and Insulin/AKT/S6 K signal pathways and reproductive hormone receptors were detected by RT-q PCR,Western Blot and immunohistochemistry.The results show that:1.The expression of some members of NOD family changed in the endometrium of sheep during early pregnancy.The relative expression of CIITA,NAIP,NOD1,NOD2,NLRP1 and NLRP3 was down-regulated compared to the non-pregnant sheep,while the relative expression of NLRP7 was up-regulated;Immunohistochemistry(IHC)showed that CIITA was located in the luminal epithelium,superficial glandular epithelium and stroma,NOD2 was located in the spongy layer and basal layer of endometrium.2.In the endometrium of sheep during early pregnancy,the expression of node genes in Insulin/AKT/S6 K signaling pathway showed inconsistent expression.In the early stage of pregnancy,INSR and PI3 K showed multi-stage changes that increased at first and then decreased,the expression of PDK1 was down-regulated,the activity of AKT1 was enhanced,and the activities of m TOR,4EBP1 and S6 K were inhibited.IHC results showed that p-AKT1 and m TOR were located in luminal epithelium and superficial glandular epithelium.3.In the endometrium of sheep during early pregnancy,the expression of FSHR was down-regulated,LHR was up-regulated,ESR,GHR and THR were increased and then down-regulated,while the change of ADRB3 was not obvious.IHC results showed that FSHR was located in superficial glandular epithelium and stromal cells,and GHR was located in luminal epithelium and superficial glandular epithel.To sum up,the changes in the expression of NOD-like receptor family members,Insulin/AKT/S6 K pathway and reproductive hormone receptors in the endometrium of sheep during early pregnancy,indicate that NOD and Insulin/AKT/S6 K pathways and reproductive hormone receptors are involved in the regulation of maternal immune tolerance,and embryo implantation in sheep.