Therapeutic Effect Of Zuogui Pill And Umbilical Cord Mesenchymal Stem Cells Transplantation On Injuried Endometrium

Part1 Study on the expression of PI3 K,AKTm RNA in thin endometriumObjective: To investigate the expression of PI3K?AKTm RNA in thin endometrium tissues,and to explore the effect of PI3K/AKT signal pathway on thin endometrium.Methods:Sixty endometrium specimens were collected from Department of Gynecology of our hospital from June 2015 to June 2016,including 30 cases of normal endometrium tissues and 30 cases of thin endometrium tissues.The western medicine diagnostic criteria of thin endometrium and normal endometrium refers to reference.After hysteroscopy surgery,collected the endometrial tissues and stored at-70?.Results:The expression levels of PI3 K,AKT mRNA,in thin endometrium group were significantly decreased,as compared with those in normal endometrium group(P<0.05).Conclusions:PI3K/AKT signaling pathways exist both in thin and normal endometrium tissue.The expression levels of PI3K?AKT in thin endometrium at proliferation period are significantly decreased,Speculated that PI3K/AKT signaling pathway may be involved in the growth and differentiation of endometrial tissue,may be play an important role in the pathogenesis and development of thin endometrium.Part2 Therapeutic effect of Zuogui Pill and umbilical cord mesenchymal stem cells transplantation on injuried endometriumObjective:Zuogui Pill is the representative prescription of nourishing yin and tonifying kidney yin,wrote by the famous expert of tonifying and warming in Ming dynasty.The prescription has the function of nourishing yin and invigorating kidney,replenishing vital essence and nourishing the Bone marrow,zuogui pill also can regulate the hypothalamus pituitary target gland axis,so is widely used in gynecological clinical disease,bone disease,nervous system diseases,etc.Umbilical cord mesenchymal stem cells,With a strong ego proliferation and multi-directional differentiation potential and rich of sources,does not involve the ethical issues,is becoming a new representative of adult stem cells.So,it may become the seed cells for the treatment of refractory injuried endometrial diseases.Research proves that PI3K/AKT signaling pathway as a signal molecule,plays a key role in cell proliferation,differentiation,apoptosis,migration and so on.PI3K/AKT signaling pathway ofen over expressed in endometrial carcinoma and endometriosis,so it may be involved in endometrial tissue growth and repair process.By building injuried endometrial model of SD rats,to exploring the traditional Chinese medicine(TCM)Zuogui pill combination between umbilical cord mesenchymal stem cells to repair injuried endometrial.Study the mechanism of Zuogui pill combination between umbilical cord mesenchymal stem cells repair injuried endometrial,to provide the oretical basis for the future widely applications of thin endometrium and Severe intrauterine adhesions.Methods:There is no unified standards of injuried endometrial model,the method of using 100 ? hot water scald endometrial is adopted to establish the SD rat injuried endometrium model.All building rats scald SD rats the left side of the Y type uterus.The experimental rats were divided into 5 groups randomly,six rats in each group.The group A(sham operation group):With the scalp needle catheter through the left side of the uterus after the exposuring of Y type uterus,then pulled the catheter,then Sewn up step by step;The group B(model group): after the completion of the building does not do any processing;The group C(Chinese traditional medicine group): 3 days after the completion of the building,then each rats started to gavage Zuogui pill according to 1.62 g/Kg dose daily,continuous lavage 28 days;The group D(uterine injection group): 3 days after the completion of the building,each rat had a laparotomy again,and injected 5×107 human umbilical cord mesenchymal stem cells in situ uterine.The group E(combined group): 3 days after the completion of the building,each rat had a laparotomy again,and injected 5×107 human umbilical cord mesenchymal stem cells in situ uterine,and after injection,each rats started to gavage Zuogui pill according to 1.62 g/Kg dose daily,continuous lavage 28 days.28 days after the treatment,all the rats of 5 groups were put to death,then taken out the left side of the Y type uterus of rats.Abandoned the part near ovaries and vagina,Cut into two segments,one period of endometrial tissue for HE staining and sirius red staining and immunofluorescence test,another part of endometrial tissue for Western Blot detection,detection the expression of PI3 K,and AKT protein.Results:1.Compared with the model group,HE staining showed that the endometrium thickness and the number of glands were increased in the Zuogui pills group,Uterine injection group and combined group to different extents,P<0.05.Of these groups,the endometrium thickness and the number of glands and the endometrial morphology in the uterine injection group and combined group were the most close to normal levels,but were still not fully recovered.Compared with the model group,sirius red staining showed that endometrial fibrosis mitigation in all of the three treatment groups,especially in uterine injection group and combined group.2.Red fluorescent of CD29 mostly appeared in the endometrial stroma of SD rats,a few appeared in the endometrial glandular epithelium and cavity epithelium.Compared with sham operation group,the model group and the chinese traditional medicine group,fluorescence intensity of CD29 were obviously enhancement in uterine injection group and combined group.Green fluorescent of AKT mostly appeared in cavity epithelium and the endometrial glandular epithelium of SD rats,a few appeared in the endometrial stroma.Compared with sham operation group and model group,fluorescence intensity of AKT were enhancement significant in Chinese traditional medicine group,uterine injection group and combined group,p<0.05.3.Compared with the sham operation group and the model group,the PI3 K ? AKT ? p-AKT protein in Chinese traditional medicine group,uterine injection group and combined group were obviously enhancement,p<0.05.Compared with Chinese traditional medicine group,the PI3K?AKT?p-AKT protein in uterine injection group and combined group were obviously enhancement,p<0.05;but there were no significant differences between uterine injection group and combined group.Conclusions:Our data demonstrate that Zuogui pills,human umbilical cord mesenchymal stem cells transplantation can induce regeneration of the endometrial cells and repair endometrial tissue.There still a few human umbilical cord mesenchymal stem cells stay in endometrial stroma and cavity epithelium after 28 days of human umbilical cord mesenchymal stem cells transplantation.Both Zuogui pill and human umbilical cord mesenchymal stem cells transplantation repair endometrial tissue may be related to activation of PI3K/AKT Signaling pathways.Zuogui pill combined with umbilical cord mesenchymal stem cells maybe a mew hope for patients with thin endometrium or Severe intrauterine adhesions.