论文部分内容阅读
目的研究抑癌基因WWOX对Lewis肺癌细胞c-jun蛋白表达及其转录活性的影响,探讨WWOX基因的抑癌机制。方法采用脂质体转染法将WWOX基因重组真核表达质粒转染Lewis肺癌细胞,RT-PCR和Western blot法检测WWOX基因mRNA的转录水平和蛋白的表达水平;免疫组化法检测WWOX基因转染后Lewis细胞中c-jun蛋白的表达水平;半定量RT-PCR法检测c-jun调控的4种肿瘤相关基因p21、cyclinD1、FasL及VEGF mRNA的转录水平。结果重组真核表达质粒pcDNA4.0/Myc-His-WWOX转染Lewis细胞后,WWOX基因在mRNA和蛋白水平上均得到表达;与未转染细胞和空载体转染细胞相比,WWOX基因转染细胞胞浆中c-jun蛋白的表达量升高,而细胞核中c-jun蛋白的表达量未见明显差异;p21基因mRNA的转录水平升高,cyclinD1、FasL和VEGF基因mRNA的转录水平降低。结论WWOX基因可在Lewis细胞中表达,其转染Lewis肺癌细胞后,不直接调控c-jun蛋白的表达量,但可影响其转录活性。
Objective To study the effect of tumor suppressor gene WWOX on the expression of c-jun protein and its transcriptional activity in Lewis lung carcinoma cells and to explore the mechanism of WWOX gene suppression. Methods WWOX recombinant plasmid was transfected into Lewis lung carcinoma cells by lipofection method. The mRNA and protein levels of WWOX mRNA and protein were determined by RT-PCR and Western blot respectively. Immunohistochemistry was used to detect WWOX gene transcripts The expression of c-jun protein in Lewis cells was detected by RT-PCR. The transcriptional levels of p21, cyclinD1, FasL and VEGF mRNA were detected by c-jun semi-quantitative RT-PCR. Results WWOX gene was expressed at both mRNA and protein levels after transfection of Lewis cells with recombinant eukaryotic expression vector pcDNA4.0 / Myc-His-WWOX. Compared with untransfected cells and empty vector transfected cells, WWOX gene transfection The expression of c-jun protein in the cytoplasm of the infected cells increased, but the expression of c-jun protein in the nucleus had no significant difference. The transcription level of p21 mRNA increased and the mRNA transcription levels of cyclinD1, FasL and VEGF decreased . Conclusion WWOX gene can be expressed in Lewis cells. The transfection activity of WWOX gene is not directly regulated by Lewis lung carcinoma cells.