Notch1信号途径在食管鳞癌细胞株EC9706的作用研究

目的：研究Notch1基因在食管癌细胞株中的表达及其对食管鳞癌细胞EC9706凋亡的影响.方法：通过免疫细胞化学检测Notch1基因在EC9706细胞中的表达.通过RT-PCR扩增Notch1基因,构建真核表达载体pcDNA3.1-Notch1,转染EC9706细胞,利用Western blotting检测转染pcDNA3.1-Notch1载体12h,24h,48h,72h的食管癌细胞株与转染空载体食管癌细胞株中Notch1的表达,并观察每一时期食管癌细胞株的凋亡情况.结果：在食管癌细胞株中Notch1基因低表达,此外,转染Notch1后的细胞株Notch1表达量与转染空载体相比有明显差异（F=80.442,P＜0.05）.经组间两两比较,实验组食管癌细胞株的Notch1的表达量在12h时比对照组食管癌细胞株的表达量显著增高（P＜0.01）,前者约为后者的4.6倍.但48h至72h时Notch1表达量实验纽与对照组食管癌细胞株无显著性差异（P＞0.05）.进一步通过倒置显微镜发现48h至72h细胞出现大量凋亡的现象.上述结果说明12h Notch1表达量的增加,使食管癌细胞EC9706中的Notch1信号途径得以激活,该途径激活后导致48h到72h时的细胞大量凋亡.结论：Notch1在食管癌细胞中的激活引起细胞大量凋亡,提示Notch1基因有可能成为治疗食管癌的新靶点.

The Role of the NOTCH1 Signaling Pathway in the Esophageal Squamous Carcinoma Cell Line EC9706

Objective: To study the expression of Notch1 in the esophageal squamous carcinoma cell line EC9706 and the effect of activated Notch1 on apoptosis of EC9706 cells. Methods: The expression of Notch1 in EC9706 was detected using immunocytochemistry. RT-PCR was used to amplify the Notch1 gene and then the eukaryotic expression vector pcDNA3.1-Notch1 was constructed and was used to transfect the EC9706 cells. Western blotting was used to detect Notch1 in the transfected cells,parental cells,and empty vector-transfected cells at 12h,24h,48h,and 72h. Apoptosis was also observed. Results: The expression of Notch1 protein was very low in the EC9706 cells. There was a significant difference in the expression level of Notch1 between EC9706 transfected with Notch1-pcDNA3.1 and EC9706 transfected with pcDNA3.1 alone (F=80.442,P<0.05). The expression level of Notch1 in EC9706 transfected with Notch1- pcDNA3.1 at 12h was 4.6 times higher than in EC9706 transfected with empty vector (P<0.01). However,at 48h and 72h there was no significant difference in the expression of Notch1 between the EC9706 transfected with Notch1-pcDNA3.1 and cells transfected with empty vector(P>0.05). Inverted microscopy showed considerable apoptosis of EC9706 occurred at 48h and 72h. These results indicate that the enhancement of Notch1 expression activates Notch1 signal pathways in EC9706 and promotes apoptosis at 48h and 72h. Conclusion: The activation of Notch1 in esophageal cancer cell gives rise to a great deal of apoptosis,suggesting that the Notch1 gene could be a new target for esophageal cancer treatment.