稳定表达磷酸化Stat3蛋白的EC-1细胞株的获取  

Establishment of EC-1 cell line stably expressing p-Stat3 protein

作　　者：贺红柳[1,2] 轩小燕[1,2,3] 李刚[4] 王志强[1,2] 宋一民[1,2] 李蕴蕴[1,2] 李珊珊[1,2] 

机构地区：[1]郑州大学第一附属医院病理科 [2]河南省肿瘤病理重点实验室,郑州450052 [3]郑州大学基础医学院微生物学与免疫学教研室,郑州450001 [4]中国人民解放军153中心医院胸外科,郑州450007

出　　处：《郑州大学学报（医学版）》2013年第3期297-300,共4页Journal of Zhengzhou University(Medical Sciences)

基　　金：国家自然科学基金资助项目81071970

摘　　要：目的:建立稳定表达磷酸化Stat3(p-Stat3)蛋白的EC-1细胞株。方法:应用脂质体Lipofectamine2000将pXJ40-Stat3质粒转染入EC-1细胞中,G418筛选得稳定转染pXJ40-Stat3的EC-1细胞株,IL-6处理20min以获得稳定表达p-Stat3蛋白的EC-1细胞株(转染pXJ40-Stat3+IL-6处理组)。同时以稳定转染pXJ40-Stat3(转染pXJ40-Stat3组)、IL-6处理组(不转染)、空载体组和空白组细胞作为对照。分别用RT-PCR和Western blot法检测各组细胞中Stat3 mRNA和p-Stat3蛋白的表达。结果:5组细胞中Stat3 mRNA和p-Stat3表达差异有统计学意义(FStat3 mRNA=5.971,P=0.010;Fp-Stat3=7.334,P=0.005)。转染pXJ40-Stat3+IL-6处理组EC-1细胞中Stat3 mRNA表达最高(0.72±0.01),其次是转染pXJ40-Stat3组(0.65±0.01),均高于其他3组。转染pXJ40-Stat3+IL-6处理组EC-1细胞中p-Stat3蛋白表达最高(0.72±0.03),其次为IL-6处理组(0.65±0.01),均高于其他3组。结论:成功建立稳定表达p-Stat3蛋白的EC-1细胞株。Aim ： To establish a EC-1 cell line stably expressing p-Stat3 protein. Methods： EC-1 cells were transfeet- ed with eukaryotic expression vector pXJ40-Stat3 by Lipofectamine 2000, screened by G418, and then stimulated by IL-6 for 20 rain to obtain the cell line stably expressing p-Star3 （pXJ40-Stat3 ＋ IL-6 group）. At the same time EC-1 ceils trans- fected with pXJ40-Stat3 but not stimulated by IL-6（pXJ40-Stat3 group） , EC-1 cells stimulated by IL-6 alone, EC-1 ceils transfected with blank plasmid and blank EC-1 cells were used as control. The expressions of Star3 mRNA and p-Star3 pro- tein in each group were detected by RT-PCR and Western blot. ReSults： The expression of Star3 mRNA and p-Stat3 had significant difference among the five groups（ Fstat3 mRNA = 5. 971 ,P = 0. 010 ;Fp-stat3 = 7. 334 ,P = 0. 005 ）. The expression of Stat3 mRNA in pXJ40-Stat3 ＋ IL-6 group was the highest in the five groups and it was （0.72 ±0.01 ） , followed by that in pXJ40-Stat3 group（0.65 ±0.01 ） , and no significant differences were found in the other three groups. The expression of p- Stat3 in pXJ40-Stat3 ＋ IL-6 group was also the highest in the five groups and it was （0.72 ± 0.03 ）, followed by that in IL- 6 alone group（0.65 ±0.01 ） , and no significant differences were found in the other three groups. Conclusion： The EC cell line stably expressing p-Stat3 has been successfully constructed and high expression of p-Stat3 could be obtained after IL-6 treatment.

关 键 词：食管鳞状细胞癌 EC-1细胞 pXJ40-Stat3 磷酸化Stat3蛋白 转染 

分 类 号：R735.1[医药卫生—肿瘤]