Expression And Clinical Significance Of HMAM MRNA In Peripheral Blood And VEGF Protein In The Tissues From Patients With Breast Cancer

Objective: The spread and metastasis of tumor cell were critical cases involved in the decease of breast cancer, which was a malignant disease with complicated and systemic features.Normal imageology methods could only find that the size of tumor tissue was over 1.0cm,but then there were already metastasis of other systems or organs. The change of giant molecule markers level in the blood had always taken place in the advanced stage of disease.Thus to find a specific marker was very important to detect micrometastasis in the early stage of disease. The human mammaglobin (h-MAM) gene was recently cloned by Fleming and Watson by differential screening and rapid amplification of cDNA ends. It maped to chromosome 11q12-13, and the amino acid sequence it encoded was similar to that of epithelial secretory proteins belonging to the uteroglobin family, all of which were encoded by genes clustered on the same chromosomal region. Mammaglobin expression had been observed in breast epithelial cells and up-regulated in breast cancer, but not in normal blood or bone marrow. Recently the expression of hMAM mRNA was detected in peripheral blood from patients with breast cancer by nested-RT-PCR method,which suggested hMAM mRNA was specific expression in breast cancer,and the positive expression of hMAM mRNA in peripheral blood might predict hemato- genous metastatic spreading of tumor cells,and hMAM mRNA might be potential biological marker for detecting breast cancer micrometastasisVascular endothelial growth factor (VEGF) appeared to be one of the most critical factor for blood vessel development in breast cancer. VEGF was stimulated to express mainly by hypoxia and was a specific mitogen of endothelial cell. VEGF could stimulate hyperplasia endothelialis and immigration, emerge fresh blood vessel, elevate vascular permeability, strengthen endotheliocyte migration in blood vessel epimatrix and promote metastasis of the tumor. Did VEGF have a positive correlation with the expression of micrometastasis in the blood of breast cancer? Now these researches were rare.The study aimed to observing the dynamic expression of hMAM mRNA in the blood of breast cancer by nested RT-PCR and the expression of VEGF protein in the breast cancer tissue by SP immunohistochemistry; analyze the expression of hMAM mRNA and VEGF protein and the correlation with clinical pathology factors ,such as lymphoid node metastasis,TNM stage and C-erbB-2 . And this may provide theory basement and experiment evidence assisting the study on prognosis of breast cancer and the therapy method to prevent the micrometastasis and spread of the tumor.Methods: 50 cases of breast cancer were collected, which were from Jul 2006 to sep 2007; 20cases of breast fibroadenoma, 10 cases of healthy volunteers, 5 cases of gastric carcinomas, 5 cases of colorectal carcinomas, 5 cases of esophageal carcinomas, 5 cases of lung carcinomas, 5 cases of ovarian carcinomas and 5 cases of endometrial carcinomas were collected as negative control . Breast carcinoma cell line MCF-7 was used as positive control .The expression of hMAM mRNA was assessed by nested RT-PCR; After 2～3 cycle chemotherapy ,the expression of hMAM mRNA was assessed again in the blood of 50 cases of breast cancer . The expression of VEGF protein was assessed by SP immunohistochemistry.All statistical calculations were carried out using the statistical software named Statistical Product and Services Solutions(SPSS) 13.0. The Chi-square test was used to analyze data.Results: The positive rates of hMAM Mrna was 34.0% in peripheral blood of breast cancer,hMAM mRNA was not expressed in peripheral blood from other cancers,breast benign lesions or healthy volunteers.hMAM mRNA expression was correlated with lymph nodes status and TNM stage,the positive expression rate of hMAM mRNA in peripheral blood of breast cancer patients with stageⅢandⅣwas significantly higher than those patients with stageⅠandⅡ(P=0.015). hMAM mRNA expression was not correlated with age,tumor size,estrogen receptor(ER)status,progesterone receptor(PR)status,p53,PCNA or HER-2. After 2～3 cycle chemotherapy,13 cases in 17cases of positive hMAMmRNA turned to be negative,there was statistical difference between pre-chemotherapy and post-chemotherapy.VEGF antigen was mostly observed in the cytoplasm or membrane of cancer cells, and seldom in the cytoplasm of endothelium cell in the cancer stroma by the semiquantitative method of immunohistochemistry. The positive rate was 64.0%, which was obviously higher than that of control group(P<0.05). The expressions of VEGF protein in breast cancer tissue had a positive correlation with the expressions of hMAMmRNA in the blood of breast cancer.The expressions of VEGF protein was correlated with tumor size, status of lymph node,clinical TNM stage (P < 0.05),but not with the age (P> 0.05). And the expressions of VEGF by ICH had a positive correlation with the expressions of C-erbB-2 (P<0.05).Conclusions: hMAM mRNA was specific expression in breast cancer, and the positive expression of hMAM mRNA in peripheral blood might predict hematogenous metastatic spreading of tumor cells,and hMAM mRNA may be potential biological marker for detecting breast cancer micrometastasis. The expressions of VEGF protein in breast cancer tissue was correlated with the expressions of hMAMmRNA in the blood of breast cancer,which suggested that breast cancer patients with the positive expression of VEGF protein may develop micrometastasis in the blood more easily.the change of hMAMmRNA expression level may predict the efficicency of chemotherapy after operation and hMAMmRNA may become an important independent prognostic marker.