| Background:Breast cancer is one of the frequent malignant tumors in female. Despite important advances in diagnosis and therapy, breast cancer patients'overall survival has not been improved markedly. Oncogenes and anti-genes are involved in the carcinogenesis and development. So screening genes related to the carcinogenesis and development of breast cancer is important for elucidating the mechanism of breast cancer and seeking new tumor markers.Our previous microarray analysis verified:COL11A1 and MMP3 was significantly up-regulated in breast cancer tissues compared with adjacent normal breast, and down-regulated in lymph node metastasis tissues compared with paired primary breast cancer tissues.Objective:Large sample cases were used to confirm changes of COL11A1 and MMP3 gene expression along with the carcinogenesis and development of breast cancer and further assessed its clinical significances.Methods:Using real-time RT-PCR, COL11A1 and MMP3 mRNA expression were assessed in 107 primary breast cancer specimens,30 cases of benign breast tissues,30 paired adjacent normal breast and primary breast cancer tissues,30 paired primary breast cancer and matched lymph node metastasis tissues. To determine the mRNA location of COL11A1, we used in situ hybridization to detect the mRNA expression of COL11A1. To confirm the consistency between protein and mRNA expression of COL11A1 and MMP3, the protein expression of COL11A1 and MMP3 was further determined by western blot in sets of specimens with matched adjacent normal breast, primary breast cancer and lymph node metastasis tissues from the same patients and seven breast cancer cell lines:MDA-MB-231, MDA-MB-435, ZR-75-1, SK-BR-3, T47-D, MCF-7, MDA-MB-453.Results:1. The dynamic changes of COL11A1 during the carcinogenesis and development of breast cancer and its clinical significances.(1) COL11A1 mRNA expression level was significantly higher in primary breast cancer (n=30) compared with matched adjacent normal breast tissues (t=4.442, P=0.000), and COL11A1 mRNA expression level (n=30) was significantly lower in lymph node metastasis tissues compared with matched primary breast cancer specimens (t=3.050, P=0.005).(2) Expression of COL11A1 mRNA was 2.7-fold higher in benign breast tissues than in adjacent normal samples (t=2.622,P=0.012), Expression of COL11A1 mRNA was 67.47-fold higher in primary breast cancer samples than in normal samples (t=7.056, P=0.000),25-fold higher than in benign breast samples (t=6.866, P=0.000),9-fold higher than in lymph node metastasis.(3) The COL11A1 mRNA expression showed the significant correlation with involved lymph nodes (P=0.046), clinical stage (P=0.004), progesterone receptor status (PR). No correlation was found with COL11A1 mRNA expression levels and other clinical characteristics such as patients' age,tumor size, histological grade, Her-2 and estrogen receptor (ER) status.(4) 29 of 30 (96.9%) adjacent normal breast tissues and 25 of 30 (83.3%) benign breast tissues showed lower levels of COL11A1 mRNA expression, but 87 of 107 (81.3%) primary breast cancer specimens showed higher levels of COL11A1 mRNA expression. COL11A1 mRNA expression has high sensitivity (81.3%) and specificity (83%) for discriminating between benign breast disease and primary breast cancer.(5) Breast cancer patients with high levels of COL11A1 mRNA have more incidents of metastasis, recurrence, or death when compared with patients with low levels of COL11A1 mRNA in the primary tumor.(6) Most of the hybridization signal was derived from breast cancer cells, and some staining was also found on the stroma in and around breast cancer cells. Primary breast cancer tissues cells show intense cytoplasmic staining, while only minimal staining in lymph node metastasis tissues cells.(7) The COL11A1 protein expression patterns follow the same trend as observed with the gene expression patterns. The ability of invasion of breast cancer cell line has no relationship with the expression of COL11A1 mRNA. 2. The dynamic changes of MMP3 during the carcinogenesis and development of breast cancer and its clinical significance.(1) The MMP3 mRNA of primary breast cancer was up-regulated compared with its matched adjacent normal breast tissue in 53.33% cases (16/30), down-regulated in 23.33% cases (7/30), and equal with the normal breast tissue in 23.33%(7/30). In lymph node metastasis tissues MMP3 mRNA expression level (n=30) was significantly lower compared with matched primary breast cancer specimens (t=5.855, P=0.000).(2) Expression of MMP3 mRNA was 11.66-fold higher in benign breast tissues than in adjacent normal samples(t=1.321, P=0.197);Expression of MMP3 mRNA was 2.93-fold higher in primary breast cancer samples than in normal samples (t=3.684, P=0.000), has no correlation with benign breast samples (t=1.081, P= 0.288),8.57-fold higher than in lymph node metastasis(t=7.027, P=0.00).(3) No correlation was found with MMP3 mRNA expression levels and other clinical characteristics such as patients'age, tumor size, clinical stage, involved lymph nodes, histological grade, Her-2 status, progesterone receptor status (PR), and estrogen receptor (ER) status. However, along with the carcinogenesis and development of breast cancer, an evidently increased trend in MMP3 expression was observed.(4) Breast cancer patients with high levels of MMP3 have more incidences of metastasis, recurrence, or death when compared with patients with low levels of MMP3 RNA in the primary tumor.(5) The MMP3 protein expression patterns follow the same trend as observed with the gene expression patterns.(6) The invasion ability of breast cancer cell lines positively correlated with MMP3 mRNA expression levels.Conclusions:These findings indicate that the dynamic changes of COL11A1 and MMP3 expression are involved in the carcinogenesis and development of breast cancer, which suggests that COL11A1 and MMP3 may be potential prognostic makers for the patients with breast cancer. Additionally, COL11A1 mRNA level may be a useful diagnosistic marker for patients with breast cancer. |
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