The Effects Of Silencing P21 Gene By RNA Interference On IKK/IKB/NF-KB Signal In Breast Cancer Cell MCF-7

ObjectiveTo evaluate the influence of silencing P21 gene by RNA interference on the proliferation, invasion, the drug susceptibility and the IKK/IKB/NF-KB path expression in breast cancer MCF-7 cell line.Methods1. Cultivate human breast cancer MCF-7 cell, select the cells in logarithmic growth phase. The optimal segment targeting P21 gene was designed and transfected into breast cancer MCF-7 cells by Lipofectamine TM2000.2. Extracted protein after transfection and P21 protein in the transfected cells were detected by Western blotting.3. Cultured cells after transfection and detected cell proliferation at different time points by MTT; Assessment of Adriamycin susceptibility of breast cancer MCF-7cells was detected by MTT.4. Cell invasion force of breast cancer MCF-7cells was evaluated by Transwell.5. Extracted protein after transfection and NF-KB signaling pathways were detected by Western blotting.Results1. After RNA interference Breast cancer cell MCF-7, Western blotting shows that compared with NC group, the amount of P21 protein expression significantly decreased(P < 0.05) in si RNA Transfection group compared with NC group.2. MTT was used to detect cells after transfection. The si RNA transfection group of cells in 24, 36, 48, 72 h absorbance values were(0.63+/-0.004),(0.70+/-0.003),(0.87+/-0.005),(1.32+/-0.004),respectively. Cells in negative control group in 24, 36, 48, 72 h absorbance values were(0.60+/-0.002),(0.65+/-0.003),(0.83+/-0.003),(1.39+/-0.002), respectively.The difference is not obvious.3. Use MTT to detect transfected cells, si RNA transfection group of cells in different concentration of adriamycin IC50 of 72 h, IC50 is 1.513?g /ml. Negative IC50 in control cells is 1.530?g /ml. The difference is not obvious.4. Use Transwell to detect transfected cell's invasion force. The si RNA transfection group of cells in the membrane cell number is(26.7+/-8.50), negative control group in cell membrane cell number is(325.7+/-38.50), There is statistical difference with the si RNA transfection group and the control group(P < 0.05).5. Western shows that, between the si RNA transfection group and the control group, the expression of IKBa, IKKb, NF-KB protein has no statistical difference(P > 0.05), the expression of P- NFKB protein has statistical difference(P < 0.05).Conclusions1. P21 si RNA can efficiently inhibit the expression of P21 of breast Cancer MCF- 7 cells.2.When P21 gene expression is downregualted, proliferation ability and Adriamycin susceptibility of breast cancer cell MCF-7 has no change. Targeted P21 in the process of breast cancer chemotherapy may have no means.3.When P21 gene expression is downregualted, invasion ability of breast cancer cell MCF-7 to reduce. Targeted P21 may reduce breast cancer metastasis and improve the prognosis of patients with breast cancer.4.When P21 gene expression is downregualted, NF-KB phosphorylation level in breast cancer cell MCF-7 to reduce. Combinative manipulation of P21 and NF-KB pathway may be the novel and promising therapeutic targets for the patients with metastatic breast cancer.