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目的探讨Tie2-siRNA表达载体对子宫内膜癌Ishikawa细胞株生物学行为的影响。方法将表达Tie2-siRNA的碱基片段插入含有巨细胞病毒启动子、新霉素抗性基因及绿色荧光蛋白报告基因的质粒中,应用脂质体包涵体技术将质粒转染入子宫内膜癌Ishikawa细胞株,以减少Tie2基因的表达,并采用逆转录PCR及Western blot方法鉴定。用MTT实验描绘各组细胞增殖曲线,流式细胞术检测各组细胞凋亡率。结果成功转染后Tie2基因的mRNA转录水平及蛋白表达水平抑制率分别为75.5%及56.4%,Ishikawa细胞增殖72-h抑制率为73.9%,凋亡率为35.9%。结论体外下调Tie2基因的转录及表达可抑制肿瘤生长,促进细胞凋亡。
Objective To investigate the effect of Tie2-siRNA expression vector on the biological behavior of Ishikawa cell line in endometrial carcinoma. Methods The base fragment expressing Tie2-siRNA was inserted into the plasmid containing cytomegalovirus promoter, neomycin resistance gene and green fluorescent protein reporter gene. Liposome inclusion body technology was used to transfect the plasmid into endometrial carcinoma Ishikawa cell lines to reduce the expression of Tie2 gene and identified by reverse transcription PCR and Western blot. The MTT experiment was used to characterize the proliferation curve of each group, and the apoptosis rate of each group was detected by flow cytometry. Results The mRNA and protein expression levels of Tie2 were 75.5% and 56.4% respectively after transfection. The inhibition rate of Ishikawa cells was 73.9% and the apoptosis rate was 35.9%. Conclusion Down-regulation of Tie2 gene transcription and expression in vitro can inhibit tumor growth and promote apoptosis.