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目的构建人信号转导和转录激活因子3(STAT3)真核表达载体,研究STAT3基因对人胃癌细胞株MGC803的影响。方法构建pcDNA3-STAT3真核表达质粒,经脂质体介导转染胃癌细胞株MGC803。RT-PCR和Western blotting检测转染后STAT3、P-STAT3(Y705)、P-STAT3(S727)、Survivin和PCNA在MGC803细胞中mRNA和蛋白水平的表达,CCK-8法检测pcDNA3-STAT3对细胞MGC803的影响。结果测序及酶切鉴定证实,真核表达质粒pcDNA3-STAT3构建成功。Western blotting和RT-PCR实验结果证实,与转染空载体组相比,转染重组质粒组STAT3、P-STAT3(Y705)、P-STAT3(S727)、Survivin和PCNA在蛋白水平和mRNA水平的表达均明显升高,差异有统计学意义(P<0.05)。CCK-8实验结果显示,与转染空载体组相比,转染重组质粒组人胃癌细胞株MGC803增殖能力明显升高,差异有统计学意义(P<0.05)。结论真核表达质粒pcDNA3-STAT3可以提高STAT3基因在人胃癌细胞株MGC803中的表达,并且增强胃癌细胞的增殖能力。
Objective To construct human eukaryotic expression vector of signal transducers and activators of transcription 3 (STAT3) and study the effect of STAT3 on human gastric cancer cell line MGC803. Methods The pcDNA3-STAT3 eukaryotic expression plasmid was constructed and transfected into gastric cancer cell line MGC803 by liposome. The expression of STAT3, P-STAT3 (Y705), P-STAT3 (S727), Survivin and PCNA in MGC803 cells were detected by RT-PCR and Western blotting. The expression of pcDNA3-STAT3 Effect of MGC803. Results sequencing and restriction enzyme digestion confirmed that the eukaryotic expression plasmid pcDNA3-STAT3 was successfully constructed. The results of Western blotting and RT-PCR confirmed that the expression levels of STAT3, P-STAT3 (Y705), P-STAT3 (S727), Survivin and PCNA at the protein and mRNA levels The expression was significantly increased, the difference was statistically significant (P <0.05). The results of CCK-8 assay showed that the proliferation of MGC803 cells transfected with the recombinant plasmid was significantly higher than that of the blank vector transfected cells (P <0.05). Conclusion The eukaryotic expression plasmid pcDNA3-STAT3 can increase the expression of STAT3 gene in human gastric cancer cell line MGC803 and enhance the proliferation of gastric cancer cells.