超声介导自制白蛋白微泡和声诺维对体外报告基因转染效率的比较研究

目的:探讨超声介导微泡破裂法促进外源基因的安全转移的方法,为临床上肿瘤等疾病的基因治疗研究提供新的思路。方法:以绿色荧光蛋白基因为标记基因,以人大肠癌细胞株SW480为靶细胞,分别以自制白蛋白微泡和声诺维(SonoVue)微泡作为载体,通过超声介导微泡破裂法促进绿色荧光蛋白GFP基因在人大肠癌细胞株的定向转染,以激光共聚焦显微镜来定性和半定量观察GFP在靶细胞的表达情况,以椎虫蓝染色法检测超声作用于细胞的安全性。结果:当超声的强度为0.75W/cm2,作用时间为40s时,对细胞比较安全;自制白蛋白微泡和声诺维微泡的浓度为10%时,分别达到最佳的基因转染效率,两者无显著性差异,但后者的相对表达强度较高。结论:超声介导微泡破裂法促进外源基因的转移是一种比较安全而有效的基因转染方法。

Comparative study on efficiency of gene transfection mediated by ultrasound enhanced self-made albumin microbubble or SonoVue microbubble destruction in vitro

Objective:To develop a novel method to effectively deliver green fluorescence protein(GFP) gene into human colon cancer cell line SW480 in vitro by ultrasound mediated microbubble destruction. Methods:Plasmid(pcDNA3.1/GFP) was transfected into the human colon cancer cell line SW480 with ultrasound combined with albumin microbubble or SonoVue microbubbles. Expression of the GFP gene was quantified by laser confocal microscopy analysis. Cell viability was assayed by Trypan Blue staining. Results:Ultrasound combined with microbubbles could enhance gene transfer in cultured cells, but the effect varied according to the ultrasound condition and microbubble concentration. Optimal gene expression occurred with microbubble at the concentration of 10% and ultrasound parameter at 0.75 W/cm2 for 40 s. Under this condition, the transfection rate of GFP gene with ultrasound mediated albumin microbubble destruction method was similar to that with SonoVue(P > 0. 05) and the relative fluorescence intensity of GFP gene was lower than that of transfection with SonoVue(P < 0. 01). Conclusion:Ultrasound-mediated albumin microbubble or SonoVue microbubble destruction method is a promising strategy for gene delivery.