Zhu Hong-jian, Zhang Zhi-qing, Zeng Xiang-fu, Wei Shou-shun, Xu Chun-xiao, Huang Guo-jin, Guo Ying-lu
Institute of Urology, Peking University, Beijing 100034, China.
Zhonghua Zhong Liu Za Zhi. 2004 Jan;26(1):22-5.
To study the effect of gene expression of mouse uroplakin II (UPII) promoter on human bladder cell cancer cell line.
The mRNA expression of different cell lines was quantified by RT-PCR. Green fluorescent protein (GFP) and luciferase (Luc) were used as reporter genes. The plasmids carrying UPII or GFP were constructed and transfected into human cell lines of bladder transitional cell cancer (BIU-87), kindey cancer (GRC-1), vascular endothelium (EC), lung cancer cell line (A549) and skin fibroblast cell line (Hs27). GFP activity of cells was detected by confocual microscopy and flow cytometry (FCM). Luciferase value was measured by luminometer (microplate) and luciferase to beta-galactosidase ratios (L/G values) were used for evaluating transfection efficiency.
RT-PCR showed high expression level of UPII mRNA in bladder cancer cell line BIU-87, whereas low level or no expression in nonbladder cancer cell lines. The activity of GFP in bladder cancer (BIU-87) cell was higher than that in the other cell lines (5 - 10/HP versus 0 - 2/HP), with 4.34% positive cells in BIU-87 detected by FCM, but no positive cell was found in the other cell lines. L/G values indicated that the luciferase expression in human bladder cancer cells transfected with mouse UPII promoter was 1.8 - 8.2-fold as high as that in the nonbladder cell lines.
Mouse UPII promoter gene can be expressed in a tissue-specific fashion in human urinary bladder cancer. It is capable of initiating transcription of reporter genes in human bladder cancer cell line.
研究小鼠尿血小板溶素II（UPII）启动子基因表达对人膀胱癌细胞系的影响。
采用逆转录聚合酶链反应（RT-PCR）定量检测不同细胞系的mRNA表达。以绿色荧光蛋白（GFP）和荧光素酶（Luc）作为报告基因。构建携带UPII或GFP的质粒，并转染至人膀胱移行细胞癌（BIU-87）、肾癌（GRC-1）、血管内皮细胞（EC）、肺癌细胞系（A549）和皮肤成纤维细胞系（Hs27）。通过共聚焦显微镜和流式细胞术（FCM）检测细胞的GFP活性。用光度计测量荧光素酶值，并采用荧光素酶与β-半乳糖苷酶比值（L/G值）评估转染效率。
RT-PCR显示，UPII mRNA在膀胱癌细胞系BIU-87中高表达，而在非膀胱癌细胞系中低表达或无表达。膀胱癌（BIU-87）细胞中的GFP活性高于其他细胞系（5 - 10/HP对0 - 2/HP），FCM检测发现BIU-87中有4.34%的阳性细胞，而其他细胞系未发现阳性细胞。L/G值表明，用小鼠UPII启动子转染的人膀胱癌细胞中荧光素酶表达比非膀胱细胞系高1.
