超声微泡介导pEGFP-N1质粒转染人牙周膜成纤维细胞的实验研究

Ultrasound-mediated lipid microbubble destruction enhances pEGFP-N1 expression in human periodontal ligament fibroblasts cell

目的探讨超声微泡造影剂在一定能量的超声波辐照下,介导质粒pEGFP-N1转染人牙周膜成纤维细胞(HP-DLFs)的效率及安全性。方法体外原代培养HPDLFs,以EGFP基因为报告基因,脂质微泡造影剂为载体,用超声辐照介导质粒pEGFP-N1转染HPDLFs。实验组超声+微泡+质粒组根据转染条件不同分成不同亚组,对照组为质粒组、微泡+质粒组、超声+质粒组和脂质体+质粒组。转染48h后在倒置荧光显微镜下观察绿色荧光蛋白GFP表达。同时用MTT法检测HPDLFs的活力。结果超声微泡介导的质粒对HPDLFs的转染效率与脂质体介导的质粒转染效率相似,明显高于其他对照组。超声+微泡+质粒组中HPDLFs的活力明显高于脂质体+质粒组。结论在一定条件下,超声微泡能安全、有效地介导外源基因的转染与表达。

Objective To investigate whether ultrasound-mediated microbubble destruction can safely and effectively deliver pEGFP-N1 plasmid to human periodontal ligament fibroblasts. Methods The primary cultured human periodontal ligament fibroblasts were divided into five groups. Experiment groups were ultrasound＋microbubble＋plasmid. Based on different conditions of transfection, experiment groups were further divided into sub groups. Control groups included naked plasmid, ultrasound＋ plasmid, microhubble＋ plasmid and liposome＋ plasmid. After 48 h, GFP expression in the human periodontal ligament fibroblasts was detected using phase-contrast fluorescence microscopy. MTT was adopted to measure the cells vitality of every group. Results The transfeetion efficiency of ultrasound＋mierobubble＋plasmid group was similar to that of the liposome＋plasmid group, which were both higher than that of the other control groups, but the cells vitality of the ultrasound＋ microbubble＋ plasmid group was higher than that of the liposome＋ plasmid group. Conclusion Under some specific conditions, ultrasound mediated mierobubble destruction method can safely and effectively enhance the reporter gene transfeetion and expression.