超声联合阳离子微泡造影剂体外增强基因转染

Ultrasound combined with cationic microbubbles enhanced in vitro gene transfer

目的探讨超声介导自制阳离子微泡破坏、增强基因转染的效果,并与普通脂质微泡对比,寻找一种更可靠、完善的基因载体工具。方法采用水浴机械震荡法,加入DC-胆固醇制备阳离子微泡,观察其物理特性并检测其体外载基因能力。将微泡和质粒DNA加入人脐静脉血管内皮细胞后分为6组,观测不同微泡浓度对细胞存活率的影响,应用荧光显微镜观察增强型绿色荧光蛋白的表达,流式细胞仪检测基因转染率,MTT法分析细胞存活率。结果自制阳离子微泡形态规整、分散度好,最大载基因率为39.7%。超声联合阳离子微泡能够增强基因转染,与普通脂质微泡组相比差异有统计学意义;随着微泡浓度增加,细胞损伤增大。结论采用自制阳离子微泡可在体外实现较高效率的基因转染。

Objective To observe the effects of ultrasound mediated destruction enhanced gene transfer with self-made cationic microbubbles comparing with common lipid microbubbles,so as to explore a more reliable and perfect gene vector.Methods Cationic lipid ultrasound microbubbles was prepared by mechanical oscillation with DC-cholesterol.Its physical properties and gene loading ability were detected.Microbubbles and plasmid DNA with HUVEC were divided into 6 groups,while radiation parameters were optimized.Cell viability of different concentration microbubbles was assessed.Expression of the enhanced green fluorescent protein was observed with fluorescent microscopy,and gene loading ability was detected with flow cytometry.The cell viability was measured using MTT assay.Results Regular shape and good dispersion were observed in self-made cationic microbubbles.The maximum ratio of loading gene of microbubbles was 39.7%.Between common lipid microbubbles and ultrasound mediated cationic microbubbles destruction,statistical differences were found.Decrease in viability of HUVEC was observed with increasing MB concentration.Conclusion The self-made cationic microbubbles have high gene loading ability,being able to enhance the efficiency of in vitro gene transfer.