| Background Breast cancer is the most common malignant tumors of women in the world, accounting for about 18% of the women who suffer from the cancer. The latest statistics pointed out that about 120 million women were diagnosed with breast cancer of the whole world. Compared with the developed countries, China has a low incidence of the breast cancer, but in the recent years the incidence rate has increased significantly. At present, there are about 47 million patients suffered from breast cancer. Because of the improvement of the diagnosis and treatment, more and more women are able to survive. The absolute number of people dying from breast cancer each year is declining. The 5 year survival rate is about 50%-60%. However the domestic and foreign related research reports that invasion and metastasis are still the main challenge in the treatment of breast cancer of clinical work, which is the main reason for the cause of treatment failure and death in patients. At the present stage, the metastasis mechanism of breast cancer is still not very clarified and the emergence of the drug resistance to the molecular target agents has arisen. Therefore, it is very urgent to explore the metastasis mechanism of breast cancer and find newtarget genes for the treatment of breast cancer. TM4SF1(Transmembrane-4-L-six-family-member-1) is a member of the four transmembrane protein L6 superfamily, and it showed abnormal high expression in the in lung cancer, pancreatic cancer, prostate cancer, colorectal cancer and other malignant tumors, while it has a relatively low expression in normal tissues and normal vascular endothelial cells, and TM4SF1 is closely related to the proliferation, adhesion, migration, invasion of the tumor cells and tumor angiogenesis. Recently, TM4SF1 is widely studied as a new target for the treatment for a variety of malignant tumors.Objective 1. To compare the expression of TM4SF1 in human breast cancer cell lines MCF-7 and MDA-MB-231, and to explore the correlation between the gene and the degree of malignancy; 2. When TM4SF1 was silenced, to analyse the changes of the proliferation and migration ability about the two kinds of cells.Methods The TM4SF1 m RNA relative expression of human breast cancer cell line MCF-7 and MDA-MB-231 was investigated by using Real-time PCR and the results were compared with that of MCF-10 A cells; The application of si RNA was used to transient transfect two kinds of cells lines MCF-7 and MBA-MB-231; Western blot assay was used to analyze the effect of transfection and its protein expression; CCK-8 assay and wound healing assay were used to measure the difference of cell proliferation and migration ability respectively when TM4SF1 was silenced.Results 1. The efficiency of transient transfection: when TM4SF1 was silenced, the relative expression of TM4SF1 m RNA was significantly lower than that in the NC SiGroup and the control group, which indicated that the gene was successfully inhibited(P<0.01). 2. The relative expression m RNA of three kinds of cell lines: the relative expression of TM4SF1 m RNA was(0.016 ± 0.004),(1.159 ± 0.218) and(1.396 ± 0.199) in MCF-10 A, MCF-7 and MDA-MB-231 cells respectively; and the relative expression of TM4SF1 m RNA in MCF-7 and MDA-MB-231 were higher than those in MCF-10 A cells(F=150.447, P < 0.01); the relative expression of TM4SF1 m RNA in highly malignant triple negative breast cancer cell line MDA-MB-231 TM4SF1 m RNA was higher than that in low-grade estrogen receptor positive cell line MCF-7(P < 0.05). 3. Blot Western results: the bands of TM4SF1 protein bands in the silenced group were significantly darker than the negative control group and blank control group, which showed that the two breast cancer cell lines had been successfully transfected. 4. CCK-8 results: when TM4SF1 was silenced on 0h of MCF-7 cells, the cell proliferation ability of group si NC and group si TM4SF1 was:(0.113 ± 0.015) and(0.112 ± 0.010), respectively; on 24 h, the ability of corresponding group was:(1.074 ± 0.081) and(1.028 ± 0.042); on 48 h, the ability of corresponding group was:(1.114 ± 0.061) and(1.021 ± 0.055); on 72 h, the ability of corresponding group was:(1.070 ± 0.101) and(0.909 ± 0.075). when TM4SF1 was silenced on 0h of MDA-MB-231 cells, the cell proliferation ability of group si NC and group si TM4SF1 was:(0.110 ± 0.010) and(0.112 ± 0.009), respectively; on 24 h, the ability of corresponding group was:(1.296 ± 0.072) and(1.199 ± 0.055); on 48 h, the ability of corresponding group was:(1.335 ± 0.054) and(1.247 ± 0.071); on 72 h, the ability of corresponding group was:(1.261 ± 0.047) and(1.123 ± 0.097). Above each time, the data in group si NC and group si TM4SF1 was organized with two independent samples t test, the results were no statistically significant(P>0.05). 5. The scratch test results: when TM4SF1 was silenced on 24 h of MCF-7 cells, the migration rate in si NC group was(42.359 ± 3.639)%, and the rate was(23.412 ± 6.397)% in si TM4SF1 group, and t test results: t=4.459, P = 0.011; when TM4SF1 was silenced on 24 h of MDA-MB-231 cells, the migration rate in si NC group was(51.799 ± 5.429)%, and the rate was(30.549 ± 5.978)% in si TM4SF1 group, and t test results: t=4.558, P = 0.010.Conclusion 1. The expression of TM4SF1 in breast cancer cells shows relatively higher than that in normal breast cells 2. TM4SF1 enhances the migration ability of breast cancer cells and has no significant correlation with the cell proliferation ability; 3. TM4SF1 may be positively correlated with the degree of malignancy. |
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