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背景与目的:在大多数肿瘤组织中都发现了survivin的异常高表达现象,这说明survivin在肿瘤发生中具有重要作用。本文的目的在于研究突变型survivin对肿瘤细胞恶性表型的影响,并初步探讨其可能机制。方法:构建突变型survivin的表达载体,转染HeLa细胞,通过G418筛选,得到稳定表达的细胞克隆;利用流式细胞仪分析HeLa细胞的凋亡;通过Westernblot检测突变型survivin对细胞周期蛋白的影响。结果:突变型survivin表达质粒在HeLa细胞中表达良好,利用相应的抗体可以检测到其表达的蛋白;表达突变型survivin的HeLa细胞与正常对照相比,克隆形成能力明显减弱;瞬时转染突变型survivin的HeLa细胞可以自主发生凋亡;而且,突变型survivin还可以使多核HeLa细胞增多,survivin-N端的作用比survivinT34A更明显;突变型survivin可以影响周期蛋白的cyclinD1,使其蛋白水平分别下降了68%和12%。结论:突变型survivin可以部分逆转HeLa细胞的恶性表型,cyclinD1蛋白水平下降可能是其发挥作用的重要机制之一。
BACKGROUND & AIM: Abnormal high expression of survivin was found in most tumor tissues, indicating that survivin plays an important role in tumorigenesis. The purpose of this paper is to study the effect of mutant survivin on the malignant phenotype of tumor cells and to explore its possible mechanism. Methods: The mutant survivin was constructed and transfected into HeLa cells. The stable clones were obtained by G418 screening. The apoptosis of HeLa cells was analyzed by flow cytometry. The effect of survivin on the expression of cyclin was analyzed by Western blot . Results: The mutant survivin expression plasmid was well expressed in HeLa cells. The expression of the survivin mutant plasmid was detected by using the corresponding antibodies. Compared with the normal control, the HeLa cells expressing the mutant survivin showed a marked decrease in clonogenic capacity. The transient transfection mutant survivin HeLa cells can autonomously apoptosis; Moreover, mutant survivin can also increase the number of multi-nuclear HeLa cells, the role of survivin-N end than survivinT34A more obvious; mutant survivin can affect the cyclinD1 cyclinD1 protein levels decreased 68% and 12%. Conclusion: Survivin can partially reverse the malignant phenotype of HeLa cells. The decrease of cyclinD1 protein level may be one of the important mechanisms.