人UroplakinII与小鼠UroplakinII启动子活性和组织特异性对比

朱宏建; 曾祥福; 魏守顺; 郭应禄

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人UroplakinII与小鼠UroplakinII启动子活性和组织特异性对比

首发时间：2006-11-30

朱宏建 ¹ 曾祥福 ¹ 魏守顺 ¹ 郭应禄 ²

1、武警总医院泌尿外科
2、北京大学第一医院，北京大学泌尿外科研究所

摘要：膀胱癌是泌尿外科最常见的恶性肿瘤，基因治疗已成为治疗恶性肿瘤的重要方法之一。本研究旨在探讨人UroplakinII (UPII) 启动子与小鼠UPII启动子在人细胞株中的启动活性和组织特异性强弱。方法: 构建以人或小鼠UPII启动子为调控基因，以绿色荧光蛋白（GFP）和荧光素激酶（Luc）为报告基因的重组质粒，应用脂质体转染技术，将重组质粒转染入膀胱移行细胞癌细胞（BIU-87）和肾癌细胞（GRC-1）、脐静脉血管内皮细胞（EC）、肺癌细胞（A549）、皮肤成纤维细胞（Hs27），利用共聚焦显微镜、流式细胞仪和微板检测仪观察细胞表达GFP和Luc的情况。结果: 含人UPII启动子重组质粒转染后，BIU-87表达GFP较多，10-15/HP，亮度较强，流式细胞仪测定表达量为10.1%。而GRC-1、EC细胞表达极少，0-5/HP，亮度暗，流式细胞仪测定表达量为0和1.8％。Luc在BIU-87中的表达量是其它组织细胞的2.2倍以上。含小鼠UPII启动子重组质粒转染后，BIU-87表达GFP较多，5~10/HP，亮度较强，流式细胞仪测定表达量为4.34%。而EC细胞表达极少，0~2/HP，亮度暗，流式细胞仪测定表达量分别为0%。Luc在BIU-87中的表达量是其他组织细胞的1.8~8.2倍，人UPII启动子与小鼠UPII启动子相比，无论是GFP还是Luc差异有显著性（P<0.01）。结论: 人UPII 启动子比小鼠UPII启动子在人细胞株中具有更高的启动活性和更低的组织特异性。为靶向性基因治疗膀胱癌提供了实验依据。

关键词： UroplakinII 启动子膀胱癌

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Compare activity and tissue-specific between human UroplakinII promoter and mouse UroplakinII promoter in human bladder cell

Zhu Hong jian ¹ Zeng Xiangfu ¹ Wei Shoushun ¹ Guo Yinglu

1、The General Hospital of Armed Police Forces

Abstract：Objective Differential expression of the desired gene product in the target tissue is central to the concept of gene therapy. One approach is to use a tissue-specific promoter to drive therapeutic genes. The feasibility of tissue-specific gene therapy for bladder cancer and its transcriptional control were investigated using human UroplakinII (UPII) promoter. Methods Green fluorescent protein (GFP) and luciferase (Luc) controlled by hUPII promoter or mUPII promoter were used as reporter genes. The plasmid carrying hUPII or mUPII and GFP was constructed and transfected into BIU-87, GRC-1 and EC. GFP activity of cells was detected by confocal microscope and flow cytometry (FCM). The plasmids carrying Luc and hUPII or mUPII were constructed and transfected into BIU-87, EC，A549 and Hs27. Luciferase values were measured by luminometer (microplate). L/G values were acquired for transfection efficiency in all cells by dividing the luciferase values by the β-galactosidase levels. Results The activity of GFP controlled by hUPII promoter in BIU-87 cells is higher than in other cells (10-15/HP versus 0-5/HP). 10.1% positive cells in BIU-87 could be detected by FCM. Little positive cells were found in other cell lines. L/G values showed the luciferase expression from hUPII promoter in human bladder cancer cells is significantly higher than in nonbladder cell lines. Furthermore, The activity of GFP controlled by hUPII promoter in bladder cancer (BIU-87) cells is higher than in other cells (5~10/HP versus 0~2/HP). 4.34% positive cell in BIU-87 could be detected by FCM. No positive cell is found in other cell lines. L/G values show the luciferase expression from mUPII promoter in human bladder cancer cells is 1.8~8.2 folds higher than in nonbladder cell lines. Conclusion hUPII promoter has better activity and worse tissue-specific than mUPII promoter. The use of a highly specific promoter-driven gene vector will provide a unique strategy for bladder cancer gene therapy.

Keywords： UroplakinII Promoter Bladder cancer

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1. 教育部博士点基金（20030001025）

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朱宏建，曾祥福，魏守顺，等. 人UroplakinII与小鼠UroplakinII启动子活性和组织特异性对比[EB/OL]. 北京：中国科技论文在线 [2006-11-30]. https://www.paper.edu.cn/releasepaper/content/200611-912.

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论文编号	200611-912
论文题目	人UroplakinII与小鼠UroplakinII启动子活性和组织特异性对比
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