流体力学注射介导的重组pcDNA-IFN-λ3-EGFP质粒在小鼠体内的表达

Expression of recombinant plasmid pcDNA-IFN-λ3-EGFP in target organs transfered by intravenous hydrodynamics-based injection in mice

目的研究流体力学尾静脉注射对目的基因器官靶向性的影响。方法将BALB/c小鼠按随机数字表分组法分为流体力学注射组和空质粒对照组。流体力学注射组将100μg/只带有增强型绿色荧光蛋白(EGFP)基因的表达质粒pcDNA-IFN-λ3-EGFP溶液2ml在5s内快速注入尾静脉,对照组在5s内注入空质粒pcDNA3.1的生理盐水2ml。注射后在不同时间内用过量麻醉剂处死小鼠,取肝、肺、肾、心、脑等组织作冰冻切片,于荧光显微镜下观察。结果流体力学注射pcDNA-IFN-λ3-EGFP质粒在肝组织中表达IFN-λ3-EGFP融合蛋白,注射8h后即有较强的表达,24h后表达最强,2d后明显减弱,1周后消失。其他组织及对照组未检测到绿色荧光蛋白表达。结论流体力学注射方法是肝靶向性的活体基因转移方法,绿色荧光蛋白可作为该方法进行目的基因研究的一个可靠和方便的示踪剂。

Objective To investigate the target organ for gene transfer by an intravenous injection of green fluorescent protein （GFP） - expressed plasmid DNA using hydrodynamics - based procedure. Methods BALB/c mice were selected and randomly divided into hydrodynamics - based injection group and plasmid control group. 2 mL GFP - expressed plasmid （ 100μg for each animal） was intravenously injected into the mice of hydrodynamics - based transfer group rapidly in 2 seconds; the control group was injected with 2 mL control plasmid saline solution in 5 seconds. The tissues were collected, the expression levels of GFP in liver, lung, kidney, heart, and brain were determined under fluorescent microscope by frozen sections. Results The hydrodynamics - based injection of naked GFP plasmid DNA into mice induced high expression level of GFP gene in liver. The fluorescent was strongest on 24 h after injection. It weakened significantly in 2 days and disappeared in 1 week. There are no fluorescent detected in other tissues and in the animals of control groups. Conclusion Hepatic delivery of foreign gene can be accomplished by hydrodynamics - based injection, and GFP is a reliable and convenient tracer agent in this procedure.