The Study Of Biological Characteristic In Decidual And Peripheral Blood NK Cells Of ICP Patients

Backgroud:Intrahepatic cholestasis of pregnancy (ICP) is thought to a complication of late pregnancy, with character of all over the body Itch of skin, liver disfunction, distinct degree morbus arcuatus. It is associated with an increased risk of fetal distress, causing premature deliveries, fetal distress, fetal deaths and stillbirths. Presently, the cause of ICP and the mechanisms of oxygen deficiency of infants are not fully elucidated, no drug treatment has achieved the ideal goals. Only after etiological factor and mechanisms of placental hypofunction are identified completely, effective measure of precaution and therapy may be designed.Though studies suggest that the disturbance of maternal immune system function may relate to the pathogenesis of the disease, its etiology and pathogenesis are unknown. Recent research on maternal-fetal interface has been proposed, the main content is composed of maternal uterus deciduas and fetal trophoblast, all kinds of cells and secreted cytokine are composed of special immunological microenvironment on at the maternal-fetal interface, which is crucial for embryo implantation and gestational maintenance. In early pregnancy, NK cells are enriched at sites where fetal trophoblast infiltrates the decidua. The appearance of decidual NK cells suggest that one of the functions of these cells is control of placentation. Histologic studies indicate NK cells are in direct contact with the invading trophoblast by adhesion molecules CD56. Decidual NK cells mainly distribute in decidua which links fetal and maternal interface and which can directly recognize fetal antigen. The specific phenotype and function of dNK cells may contribute to keep immunotolerance in fetal-maternal interface, also can they secrete an array of cytokines facilitate decidual blood rebuilding and maintain the balance of Th1/ Th2 cytokines. To study the functions and status of dNK cells undoubtedly not only helps us to comprehend the physiological phenomena of pregnancy and but also to provide new ways to research the mechanism of pathological pregnancy, as well as the development in reproduction immunology.None of the aforementioned studies refered to the change of percentage, phenotype, function and Th1/Th2 cytokine balance in peripheral blood natural killer (pNK) cells and decidua natural killer (dNK) cells in the patients with ICP. To further shed light on the immunological mechanisms of ICP, we focus our attention on local immune reactions at the maternal-fetal interface and system immunity reactions between normal late pregnant women (NLP) and ICP. We study the percentage and the phenotype in the decidua natural killer (dNK) cells in the patients with ICP, investigate the change of Th1/Th2 cytokine levels of decidual mononuclear cells and peripheral blood mononuclear cells in pregnant women with ICP, to futher investigate the change of Th1/Th2 cytokine levels in peripheral blood natural killer (pNK) cells and decidua natural killer (dNK) cells, so that we can investigate whether disturbed local immune reactions and system immunity reactions are also present in pregnant women with ICP. Undoubtedly, Elucidation of immunologic microenvironment at maternal-fetal interface with ICP must be an original pathway to investigate the mechanism of ICP.Objective:To detect the change of dNK cells and pNK cells, to compare the percentage, phenotype, Th1/ Th2 cytokine secretion and cytotocxic action between dNK cells and pNK cells between normal late pregnant women (NLP) and ICP, which can help us to further discover the important interaction between dNK cells and trophoblast cells immunotolerance local immune reactions at the maternal-fetal interface and system immunity. Methods:1) The same pregnant women third-trimester parietalis decidua and peripheral blood were obtained during elective caesarean sections (CS) in ICP patients (n= 15) and NLP (n=15). Peripheral blood mononuclear cells (PBMC) were centrifugated using a standard gradient sedimentation technique cell suspensions were prepared by an electromechanical dispersal method and centrifugated using a standard gradient sedimentation technique. And percentage and phenotype of dNK cells were detected by flow cytometric analysis.2) To detect the mRNA expression of HLA-G, HLA-E and MICA at trophoblasts from elective caesarean sections (CS) in ICP patients (n=6) and NLP (n=6) by the method of RT-PCR. They are ligands of NKG2A and NKG2D respectively.3) The concentration of Th1-type cytokine (INF-γ) was increased in patients suffering from ICP by enzyme linked immunosorbent assay, Whereas no differences were found in the level of IL-4.4) To detect the expression of intracellular Th1/Th2 cytokines IFN-γand IL-4 in dNK cells and pNK cells by the flow cytometry.5) The cytotoxic activity of dNK cells and pNK cells against targets was measured in by the LDH assay.Results:1) Compared to controls, the parietalis deciduas from ICP patients were characterized with the increased the percentage of CD3-D56+NKcells, CD56+CD16+ NK cells, CD56+CD16- NK cells, CD56+NKG2D+ NK cells. Whereas no differences were found in the percentage of CD56+NKG2A+cells.2) Decidual natural killer cells had high expression of NKG2A, which ligand HLA-G and HLA-E mRNA were also expressed in trophoblasts tissue. Although dNK cells highly expressed NKG2D, there was no MICA mRNA expression.3) The concentration of Th1-type cytokine (INF-γ) was significantly increased in patients suffering from ICP. Whereas no differences were found in the levels of IL-4.4) The expression of intracellular Th1/Th2 cytokines IFN-γand IL-4 were significantly high in dNK cells in ICP pregnant women. IFN-γand IL-4 are the most important cytokines in regulating Th1/Th2 immunological balance, suitable quantity of which play an important role in regulating immunological balance at fetal-maternal interface5) Their cytotoxic activity of dNK cells and pNK cells in ICP team against targets was significantly increased than those of dNK and pNK cells cells in NLP team.Conclusions:Decidual NK cells and peripheral NK cells in ICP team are the important natural lymphocytes, whose percentage, phenotype, Th1/Th2 cytokine secretion and cytotocxic activity are obviously different from decidual NK cells and peripheral NK cells NLP team, and also they tightly touch with the invading trophoblasts. dNK cells and their secreting cytokines compose the center of decidual immunological regulation through development, regulation, receptor expression, biological effect. Abnormal immune responses not only appear insystem immunity but also mainly manifest fetal-maternal interface immunity. From the above results, we conclude that dNK cells play an important role in regulating immunological tolerance at fetal-maternal interface and system immunity...