脂质体介导人β防御素2基因转染膀胱上皮的实验研究  

Experimental Study of Liposome-mediated Human β-defensin-2 Gene Transfer to Bladder Epithelial Cells

作　　者：赵俊丽[1] 王志平[2] 王俭勤[1] 

机构地区：[1]兰州大学第二医院肾病内科,兰州市730030 [2]兰州大学第二医院泌尿研究所,兰州市730030

出　　处：《医学分子生物学杂志》2011年第1期56-60,共5页Journal of Medical Molecular Biology

基　　金：甘肃省自然科学基金（No.0803RJZA007）;兰州大学中央高校基本科研业务费专项资金（No.1zujbky.2010149）

摘　　要：目的 探讨脂质体介导人β防御-2（humaβ-defensin2，hBD2）基因体内、外转染膀胱上皮细胞的可行性。方法采用脂质体法体外转染人膀胱上皮细胞株T24及膀胱灌注大鼠体内转染，ELISA检测细胞上清及大鼠尿液中hBD：的表达；RT—PCR及Western印迹检测细胞及大鼠膀胱中hBD：的表达；组织免疫荧光检测hBD，在膀胱黏膜的表达。结果RT—PCR及Western印迹检测显示，脂质体介导pCAGG—hBD2体、内外转染膀胱上皮细胞后均可表达重组hBD2；转染细胞上清及大鼠尿液中hBD2浓度分别为（36．5±3．2）ng／10。细胞和（4．77±1．4）ng／ml；转基因hBD2主要表达于膀胱黏膜上皮层。结论脂质体介导hBD2基因体内、外转染膀胱上皮细胞后均能获得高效表达，为泌尿系感染的防御素基因治疗提供了实验依据。Objective To assess the feasibility of in vitro and in vivo liposome mediated gene transfeetion of human β-defensin-2 （ hBD2） gene into human bladder epithelial cells and rat blad der transitional epithelium. Methods Transfection was performed using a cationic Liposome-Trans- FastTMTransfeetion Reagent. The level of secreted hBD2 in cell supernatants and rat urine was checked by ELISA. The expression of the transgene hBD2 in situ was detected by immunohistochemistry. RT-PCR and Western blotting were used to confirm the hBD2 expression in transfeeted cells and rat bladders. Results ELISA demonstrated a high level of hBD2 production in transfeeted cells （36.5 ±3.20） ng/10^6eells and a detectable level of hBD2 in rat urine （4.77 1.4 ng/ml） Expression of transgene hBD2 in transfeeted cells and rat bladders were confirmed by both RT-PCR and Western blotting. Immunohistoehemistry revealed that the transgene hBD2 was expressed in the entire epithelial layer of the transduced bladders. Conclusion Liposome-mediated hBD2 plasmid DNA transfection system is an efficient method for antimierobial gene therapy of UTI.

关 键 词：脂质体介导人β防御-2 脂质体 膀胱上皮细胞 

分 类 号：R349.6[医药卫生—基础医学]