Expression Of Oct4 And Ssea-1 In Breast Cancer

Backgrounds and objects:Oct4(octamer binding factor4) with molecular weight 18-KD contains 324 amino acids,who also be known as POU5F1/OTF3/Oct3,are one part of POU family.It is located at human chromosome 6p21.3 and possesses POU5F1 gene encoding open reading frame.POU transcription factor also known as DNA-binding protein is capable of activating the gene which has cis-(cis) gene transcription response elements at the promoter or enhancer region transcribing,which activates or inhibits gene expression,and maintains cell totipotency.This type of gene cis-response element binding sequence contained Oct,which also be called eight ATGCAAAT polymer.Oct4 expresses in oocytes,mouse preimplantation embryos,ectoderm of gastrula primordial germ cells and embryonic stem cells,but not in the progeny cells,which suggests that it has a close relation with the differentiation of cell pluripotent.In adult tissues,Oct4 mostly expresses in a number of cells who have the properties of stem cell,such as the individual cells at basal cell layer of the skin/mammary stem cells and stomach stem cells,but have declining expression or no expression in those differentiated cells,for example,which has a low expression when theⅠ-type mammary epithelial stem cells differentiate into typeⅡcells.Oct4 may be a totipotent differentiation-specific gene marker,or a indispensable gene for totipotent differentiation.Tai found the Oct4 in the weak and high tumorigenicity Human breast epithelial stem cells,pancreatic cancer cell lines Capan-2 and Pan-1,liver cancer cell lines Mahlava,Hella cancer cell line(HeLa) and Human breast cancer cell line -7 (MCF-7).Some experiments suggest that cells express Oct4 gene are potentially pluripotent stem cells,which may be the target cells that cause the process of tumors. Recent studies show that Oct4 play a role in the origin of a variety of malignant tumors.Some Swiss 3 T3 cells which have been conducted the Oct4 gene would transformate at malignant direction,and cause the formation of tumor in the subsequent experiment.All of these demonstrate that the abnormal expression on Oct4 gene may lead to malignant transformation of normal cells,what is more,it may cause the occurrence of a wide range of tumors through the activation and up-regulation its downstream target genes.SSEA-1(stage specific embryonic antigen 1),also be known as the Lewis x(Le~x, CD15),it is a trisaccharide with the structure Galβ(1-4)(Gal) Fucα(1-3)(fucose) GlcNAcβ(1-3)(GalNAC).In mammals,the SSEA-1 emerges at the late stage of 8-cell,and have a significant expression in the morula,followed by the expression in inner cell mass and endoderm,which disappear in the trophectoderm and emerge on the surface of primordial germ cells laterally.SSEA-1 can be detected in the endometrium,fallopian tubes,tubules,sperm,cerebellum,neural stem cells of adult mice,on the surface of nerve cells,glial cells,gastric epithelial cells,peripheral white blood cells and red blood cell of adult and a variety of tumors such as breast cancer, colon cancer,Hodgkin's lymphoma,and acute promyelocytic leukemia.The characteristics of SSEA-1 antigen and sialyl SSEA-1 express on the surface of tumor cells have been used to diagnose metastasis and prognosis monitor during treatment in tumor patients.It has been suggested that Lex is involved in cellular recognition during fertilization,embryogenesis and neural development;it also mediate the process of cell recognization and adhesion/the recognization between blastocyst and the endometrial cells/recognization and adhesion among the animal epithelial cell, thus cause the completion of blastocyst nidation and the formation of pykno-structure. Furthermore LeX on the surface of human sperm-egg has a confirmed effect on the sperm-egg fusion. Maria Teresa Elola,et al confirmed that MCF-7 and human polymorphonuclear neutrophils(PMN) could combine with human umbilical vein endothelial cells (HUVEC)through SSEA-1,the convidence is that this adhesion can be blockaged by anti-SSEA-1 monoclonal antibody MCS-1.Thus this theory gives us a possible way in preventing the metastasis of tumor cells,which is inhibiting target through the corresponding monoclonal antibody.Breast cancer is one of the most common female malignant tumors,and its incidence is increasing year by year.In Europe and the United States,it accounts for 25%～30%of female malignant tumor.The statistics showed that about 1.3 million people who were diagnosed with breast cancer each year all over the world in the end of 20 century,and 400,000 would die of it.In our country,the breast cancer has a second/fifth more high incidence rate in town/village,even which is higher in some large city.Breast cancer has become the greatest threat to women's health.Since the 1990',with early diagnosis and comprehensive treatment of the progress,especially progress in postoperative adjuvant therapy,the breast cancer mortality in North America/the United Kingdom and other countries had a little decrease.However, because of tumor etiology,biological behavior and development,as well as the complexity and heterogeneity,clinical heterogeneity of the tumor and other factors, the treatment to malignant tumors had not confirmed effect,breast cancer is no exception,therefore,it is very necessary for finding breast cancer prognostic factors and treatment to new target on the basis of combined treatment and the individual principles.Nowadays,even it had individual report about the study on cell lines,but the research on Oct4 and SSEA-1 in breast cancer is at the preliminary stage,whether it expresses in breast cancer? What is its significance? We know little about all of this. In view of this,the purpose of this research,include:1.Explore whether the Oct4 and SSEA-1 express in the breast cancer cell lines and breast cancer;2.Explore the expression of Oct4 and SSEA-1 and its significance in cell lines and tissues; Materials and Methods:1.Materials:Tissue specimens and cell linesTissue specimens:exairesis tissues from 2005 to 2007 were got from the First Affiliated Hospital of Zhengzhou University,which included 93 cases of breast infiltrating ductal carcinoma(all these female patents were diagnosed for the first time, and did not receive preoperative chemotherapy and radiotherapy),32 cases of breast benign lesions,2 cases of seminoma organizations.All specimens were confirmed by pathology experts.Cell lines:Human breast cancer cell line 7,MDA-MB-468 was given as a present by the Henan Key Laboratory of Cancer Pathology.After anabiosis,these cells were cultured in 10%fetal bovine serum,100U/ml penicillin,and DMEM high glucose culture medium with 100U/ml streptomycin with the saturated humidity,37℃, 5%CO2 incubator,the transfer of culture was enforced with the help of 0.25%trypsin after the culture cell began growing adherently,the logarithmic phase cells were chosen for the next step.2.Detected the expression of Oct4 and SSEA-1 with the Immunohistochemical PV two-step method and SP method.3.Chose the logarithmic phase cells to dissociate and centrifuge,coated them into a cell mass with plasma and thrombin,conventional fixed,embedded,sliced,the detection method was same with tissue one.4.Score method:Generally considered two indexes(percentage of cells staining positive accounted for the observed same kind cells and intensity of staining) to determine semi-quantitative results.Group on the basis of the proportion of positive cells:the number of positive cells≤10%for 1/;11%～50%for 2;51%～75%for 3;＞75%for 4.Judge the degree of positive intensity on the basis of coloration:no staining for 0;amber for 1;buffy for 2;brown for 3.The cross product of both were divided into 0～2(-);3～4(+);6～8(++);above8(+++).5.Statistical treatment:All data were deal with statistical software SPSS13.0, statistical methods was Chi-square test.The difference was significant when P＜0.05. Results:1.Histological section1.1 expression rate of Oct4 is 45.2%(42/93)in breast cancer;3.1%(1/32)in breast benign lesions.Compare with breast benign lesions,the expression rate of Oct4 in breast cancer is significantly higher(P＜0.05).The expression rates of Oct4 in different molecular subtypes of breast cancer are 34.5%in Luminal A;33.3%in Luminal B;66.7%in HER2 over-expression;88.2%in Basal-like,all is different statistically.1.2 The positive rates of SSEA-1 in breast cancer and breast benign lesions are both less than5%.2.Cell slices2.1 The positive rates of Oct4 in both cell lines are less than 1%.2.2 The positive rates of SSEA-1 in both cell lines are more than90%.Conclusions:1.Oct4 expresses in Breast cancer;the expression rate in Breast cancer is significantly higher than it in benign breast disease;Basal-like type has the highest expression rate.2.The expression rate of SSEA-1 is low in breast cancer and benign lesions, which is not a suitable test target for clinic.3.The expression rate of Oct4 and SSEA-1 in breast cancer and breast cancer cell lines is different,the result got from cell lines may not be suitable for entities organizations,the further research is necessary.