Epithelial Ovarian Cancer-Related Genes Involved In Limmune Escape Igs, FOXP3 Expression Studies

Ovarian tumors is a common female genital malignancy, one of the three. 5-year survival rate around 30%, has become a threat to women's lives and health of the main tumor. Among them, the source of the tumor epithelial primary ovarian tumors account for 50% -70%. Therefore, the epithelial ovarian cancer research of great significance. The current general view is that cell-mediated immunity in the anti-tumor immunity play a leading role, including CD8+, CD4+ T cells, NK cells, macrophages and DC of the anti-tumor effects. And humoral immunity can also be caused by apoptosis or growth inhibition of the signal generated, complemen-t-mediated target cell lysis (CDC) and ADCC to support the role of T cells in anti-tumor effects. Epithelial ovarian cancer patients more than the existence of abnormal immune function, mainly for cancer patients by inhibiting cell-mediated immunity in general, the effector cells, such as CD4+ T cells, CD8+ T cells, NK cell function to reduce, and in the humoral immune show aspects of the peripheral blood of patients with difficult to understand Ig levels. For epithelial ovarian cancer reveals the immune response of the induction and maintenance of the lower of the relevant mechanisms, the paper conducted the following experimental study of two parts:1 IgG, IgA, IgM, IgD, IgE expressions in the tissues of epithelial ovarian carcinomaLong time ago clinical tumors have found that some patients with the IgG, IgA, IgM and plasma of circulating immune complexes (CIC) levels were significantly higher, even of monoclonal immunoglobulin of hyperlipidemia, these antibodies can be normal cells or tumor cell-specific binding, it is known as tumor-reactive antibodies, these antibodies often precedes the increase in poor prognosis. 20th century 80's, it was found that the existence of a tumor in patients with immune suppression factor can inhibit the function of host cells, this inhibitory effect was found in the Ig and blood, and it has a prevalence of blood through the removal of Ig molecules to treat cancer method, experimental results showed that after removal of Ig tumor necrosis can be significant.In 2001 Professor Zhang Youhui study group had ascites from ovarian cancer patients with isolated an immune inhibitory factor, it can effectively inhibit the proliferation of peripheral blood T lymphocytes, and unexpectedly found that the factor of SDS-PAGE electrophoresis patterns with IgG completely consistent. Further through the flow cytometry found that ascites IgG positive cancer cells was stronger than that of the lymphocytes. The traditional theory of immunology, Ig gene only in the course of B cell development before rearrangement occurs, but in recent years, research has suggested that malignant epithelial cells can occur in Ig gene rearrangement and generation of Ig molecules.The experimental application of immunohistochemistry in epithelial ovarian cancer tissue microarray (epithelial ovarian cancer with 177 cases) and 50 cases of benign ovarian epithelial tissue in IgG, IgA, IgM, IgD, IgE specific staining, and the following research: (1) observation of IgG, IgA, IgM, IgD, IgE in patients with epithelial ovarian carcinoma characteristics. Igs was found in epithelial ovarian cancer in high-expression will be positive staining mainly in cytoplasm of cancer cells, and the existence of a large number of Igs secreted extracellular form; (2) of the Ig staining intensity score, compared IgG, IgA, IgM, IgD, IgE in epithelial ovarian cancer and benign ovarian epithelial tissue differences, results showed that IgA, IgM, IgD, IgE in epithelial ovarian cancer tissues was significantly higher than benign ovarian epithelium (P< 0.001); (3) Analysis of IgG, IgA, IgM, IgD, IgE staining intensity and tumor histological grade of relevance, results showed that IgG, IgA, IgM, IgD, IgE expression intensity and histological grade of tumor was positively correlated (P < 0.001), tumor of the lower degree of differentiation, Ig stronger expression.This experiment the following conclusions: (1) Epithelial ovarian cancer cells express high IgG, IgA, IgM, IgD, IgE, and IgG, IgA, IgM, IgD, IgE expression in the existence of a large number of exocrine cells of the phenomenon. Epithelial ovarian cancer cells to secrete Igs to the cancer tissue is epithelial ovarian cancer to participate in an important mechanism for immune escape. (2) Epithelial ovarian cancer, the lower the degree of malignancy, IgG, IgA, IgM, IgD, IgE were in weaker expressions; the higher the degree of malignancy, IgG, IgA, IgM, IgD, IgE were in stronger expressions. Abnormal expressions of IgG, IgA ,IgM, IgD, IgE in ovarian epithelial carcinoma indicate what were relation of occurring and development of ovarian epithelial carcinomaIn this experiment IgG, IgA, IgM, IgD, IgE were studied in the field of histological observation of large sample studies for the first time. The results of this study will provide a theoretical basis Igs are involved in mechanism of epithelial ovarian cancer cell immune escape mechanisms and clues to the biological treatment of Igs for the targets in epithelial ovarian cancer cells2 FOXP3 expression in epithelial ovarian cancer cellsCD4+CD25+ regulatory T cells (Treg) is the body to"active"approach to maintaining stability in autoimmune channels can inhibit inappropriate immune response, immune response can limit the scope, extent and time , while maintaining its own stability and the prevention of autoimmune diseases and transplant rejection control play very important role in the protection. CD4+CD25+ Treg cells suppress the immune system in different types of cell differentiation and effector functions: conventional CD4+ and CD8+ T lymphocytes , NKT cells, B cells, dendritic cells and monocytes / macrophages. However, recent studies have found in the tumor micro-environment in the CD4+CD25+ Treg cells increased significantly, and inhibit tumor immune effect. Fork-like transcription factor FOXP3 is the first immunosuppressive cells, CD4+CD25+ Treg development in the thymus, peripheral maintenance of the expression and function of an important regulator of genes that will enable primary CD4+CD25- T cells into CD4+CD25+ Treg. In 2001, Bnmkow foxp3 genes was reported for the first time studying the of sf mice. Sf mice is due to foxp3 gene DNA-binding domain containing the two base frameshift insertion, caused by client protein C deficiency and the forkhead domain of transcription can not be regulation. sf mice CD4+ T cell-mediated immunity imbalance, fatal disease of lymphocyte proliferation and with multiple organ infiltration, lymphocyte infiltration and organ non-specific IL-2, IL-4, IL-10, IFN-γ, etc. a large number of secreted cytokines. Homologous human FOXP3 gene mutations can cause human diseases of immune disorders, endocrine diseases, enteropathy, X chromosome-linked syndrome (IPEX), performance for the full immune disorders, associated with autoimmune endocrine diseases such as early onsetⅠtype diabetes and thyroiditis, in some cases accompanied by severe atopic atopy, including eczema, food allergy and eosinophilic inflammation.Usually found that the immune effects of tumor cells such as CD4+ T cells, CD8+ T cells, NK cell proliferation and activity were of the lower inhibitory ability. Lower of immune response exists in the tumor micro-environment.In order to explore the local epithelial ovarian cancer-related immune mechanisms Participation, this study conducted the following experiment:(1)Study of FOXP3 in ovarian epithelial carcinoma tissues Application of immunohistochemical staining method we detected FOXP3 expreession in 46 cases of epithelial ovarian cancer. 36 cases of epithelial ovarian cancer was detected in FOXP3 expression, FOXP3 staining was brown showed the positioning of the nucleus in cancer cells.(2)Study of FOXP3 expression in epithelial ovarian carcinoma cell lines Detecting FOXP3 expression in CAOV3 in SKOV3 cells. Application of reverse transcription polymerase chain reaction (RT-PCR) we detected the expression of two cells'FOXP3 mRNA expressions, sequencing, sequence and open reading frame (ORF) of the ratio; Application of immunocytochemical staining FOXP3 protein in SKOV3 cells and CAOV3 cells.The results of the two cells were detected in the expression of FOXP3 mRNA. In SKOV3 cells FOXP3 mRNA fragments were in the same sequence of the GeneBank FOXP3 mRNA; CAOV3 cells FOXP3 mRNA fragments have a mutation, resulting in a leucine into a phenylalanine; FOXP3 protein in SKOV3 cells located in the nucleus, and in CAOV3 cells located in cytoplasm.(3)FOXP3-specific siRNA interference experiment Observation of the expression of FOXP3 mRNA expression and FOXP3 protein expression by flow cytometry in SKOV3 cells after siRNA interference 48,72,96,120 hours after siRNA interference,comparing with Non-silencing siRNA group. The results showed that 48h after siRNA interference can effectively inhibit the expression of FOXP3 mRNA and inhibit the FOXP3 protein expression after interfere 48,72,96,120 hours in siRNA group than Non-silencing siRNA group. In this study, the following conclusions:(1) Epithelial ovarian cancer cells to express FOXP3, may be of an important mechanism epithelial ovarian cancer cells escape immune effector cells'attack constantly proliferation, invasion and metastasis.(2) Epithelial ovarian cancer cells can express FOXP3, may be one of important mechanism the increasing CD4+ CD25+ Treg cells in the location of epithelial ovarian cancer tissue.In this study, we first the time demonstrate epithelial ovarian cancer and ovarian cancer cell lines can express transcription factor FOXP3, will provide important theoretical basis for reveal the tumor low immune inhibition mechanisms in the locations of epithelial ovarian cancer tissue and important clues for epithelial ovarian cancer immune therapy provide.