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目的探讨转录因子E2F1表达对人乳腺癌细胞系MCF-7增殖及侵袭能力的影响。方法将构建并已鉴定的E2F1-siRNA真核表达载体(pSIREN-E2F1)和阴性对照载体(pSIRENC)利用脂质体介导转染技术转染人乳腺癌细胞系MCF-7,经嘌呤霉素筛选,得到稳定抑制E2F1表达的乳腺癌细胞(SIR组),并设阴性载体转染细胞(SIRC组)和未转染细胞(MCF-7组)为对照。3组细胞采用RT-PCR、Western blot法检测E2F1基因表达情况;采用Transwell侵袭实验、MTT实验检测细胞侵袭能力和增殖能力。结果建立了稳定抑制E2F1基因表达的乳腺癌细胞(SIR细胞);SIR组细胞E2F1mRNA、E2F1蛋白表达水平及细胞穿过膜细胞数(0.857±0.015、0.776±0.012、87.000±5.000)均明显低于SIRC组(1.563±0.031、1.512±0.011、143.000±11.000)和MCF-7组(1.544±0.018、1.494±0.013、152.000±9.000),差异均有统计学意义(P<0.05);培养第2天开始,SIR组细胞增殖能力明显低于SIRC组和MCF-7组(P<0.05)。结论采用siRNA技术抑制人乳腺癌细胞E2F1表达可明显抑制乳腺癌细胞的增殖能力,降低其侵袭能力,E2F1可作为乳腺癌发生过程中有价值的生物学指标。
Objective To investigate the effect of transcription factor E2F1 on proliferation and invasion of human breast cancer cell line MCF-7. Methods The constructed and identified E2F1-siRNA eukaryotic expression vector (pSIREN-E2F1) and negative control vector (pSIRENC) were transfected into human breast cancer cell line MCF-7 by liposome-mediated transfection. (SIR group). The transfected cells (SIRC group) and untransfected cells (MCF-7 group) were used as control. 3 groups of cells using RT-PCR, Western blot detection of E2F1 gene expression; using Transwell invasion assay, MTT assay cell invasion and proliferation ability. Results The expression of E2F1 gene in breast cancer cells (SIR cells) was inhibited stably. The expression of E2F1 mRNA and E2F1 protein in SIR group was significantly lower than that in SIR group (0.857 ± 0.015,0.776 ± 0.012 and 87.000 ± 5.000) SIRC group (1.563 ± 0.031,1.512 ± 0.011,143.000 ± 11.000) and MCF-7 group (1.544 ± 0.018,1.494 ± 0.013,152.000 ± 9.000), the difference was statistically significant (P <0.05); on the second day Initially, the cell proliferation in SIR group was significantly lower than that in SIRC group and MCF-7 group (P <0.05). Conclusion The inhibition of E2F1 expression in human breast cancer cells by siRNA can significantly inhibit the proliferation and reduce the invasion ability of breast cancer cells. E2F1 can be used as a valuable biological marker in the development of breast cancer.