Expression Rules Of Dicer In Endometrium Of Early Pregnant Mouse

ObjectiveEmbryo implantation is a complex and important life processes, it isidentified as the key to the success of pregnancy.In recent years, to explorethe mechanism of embryo implantation become hot topics in the study ofreproductive medicine, but it is still not completely clear. With the progressof epigenetics, people found that miRNA play a regulatory role to uterusreceptive and blastocyst invasive behavior in the process of implantation.Mature of miRNAs need the participation of key enzymes (nucleic acidenzyme Dicer) in the body, Dicer gene have played an important role in avariety of physiological and pathological events. This experiment regardDicer gene as the research object, its expression in endometrium in micebefore and after pregnancy implantation as the research scope, in order tofound the expression rule of Dicer, and provide some basis for the furtherexplore to the function of Dicer gene in the process of embryo implantationand its molecular mechanism.Methods1. Animal model establishment and material collection: Establish unpregnancy, pregnancy, pseudopregnancy mouse model, collecting theendometrial tissue of unpregnancy, pregnancy1d~7d, pseudopregnancy1d~7d of mice,-80℃refrigerator frozen for later use.2. The RNA extraction and real-time fluorescence quantitative PCR(RT FQ-PCR): Extraction the total RNA of endometrium by using Trizolpyrolysis liquid, reverse transcription synthesis its cDNA. According to thesequence of Dicer mRNA, designed corresponding PCR upstream anddownstream primers, using β-actin as a internal, testing the expression ofDicer mRNA in mouse endometrial tissue.3. Immunohistochemical staining: Take the endometrial tissue toparaffin embedding and sectioning, testing the Dicer protein expression inthe endometrium of mice quantitative and orientation.4. Protein extraction and Western Blot: RIPA cracking liquid was usedto extract proteins of endometrium, measuring the concentration of proteinby the BCA method. To detect the expression of Dicer protein inendometrium.Result1. Real-time fluorescent quantitative PCR results showed that, inpregnancy mice, during the implantation window and afer it(d4～d6),expressions of Dicer mRNA was significantly higher than that beforeimplantation(d1), and expressions of d5is the highest; inpseudopregnancy mouse, compared to d1, the expressions of Dicer mRNA was also increased in d4～d7.2. Immunohistochemical results showed that, in endometrium ofunpregnancy mouse, Dicer have a small amount of expression in all cavityepithelium and glandular epithelium and stromal cells; the strongexpression of Dicer began to appear in endometrial stromal cells ofpregnancy mouse of d1. To d4and d5, the expression of Dicer wassignificantly enhanced in epithelium and stroma cells in endometrial,stromal cells are stronger than the epithelial, and continued to theendometrial was decidualization; in pseudopregnancy mouse, expression ofDicer from d4to D7is significantly enhanced, d4, d5is mainlyexpressed in the matrix, d6, d7is mainly seen in the epithelium.3. Western blot results showed that, in pregnancy mice, during theimplantation window and afer it(d4～d7), expressions of Dicer proteinwas significantly higher than that before implantation(d1), and expressionsof unpregnancy and d2and d3do not change much; in pseudopregnancymouse, compared to d1, the expressions of Dicer protein was alsoincreased in d4～d7.Conclusion1. During the implantation of d4～d7, expressions of Dicer mRNAand protein was significantly higher than that before implantation(d1).2. During the implantation window and afer it, expressions of Dicerwas significantly higher than that before implantation, and it has been demonstrated in the stromal.3. There are differences in time and location in the expression of Dicerin endometrium of early pregnant mouse, the expression rule of Dicer inendometrial stromal cell may be one of the implantation mechanisms.