低分子量聚乙亚胺介导的基因转染系统及动物皮肤组织基因转染试验

将带有绿色荧光蛋白(GFP)报告基因的真核表达质粒与阳离子聚合物聚乙亚胺(PEI)结合,用肝癌细胞株CM7221实验,研究其转染效率及可能引起的细胞毒性;进一步用此PEI/DNA复合物转染小鼠皮肤组织,通过报告基因检测,研究转染基因的表达位置及持续表达时间。结果发现,低分子量PEI介导的细胞转染效率最高可达55%,转染效率与PEI结构无关,但是随着分子量的增加,转染活性略有下降。同时,随着分子量的增加,PEI对细胞的毒性也相应的加大;动物皮肤转染实验显示,转染24h后,GFP基因在皮肤组织的毛囊、汗腺、皮脂腺等处高效表达,表达可持续7天。表明低分子量PEI是低毒性、高转染效率的有用非病毒转染载体,能够在动物皮肤组织中进行基因转移,这对皮肤疾病的基因治疗具有潜在的应用价值。

GENE DELIVERY SYSTEM BASED ON LOW MOLECULAR WEIGHT POLYETHYLENIMINE AND ITS TRANSFECTION ACTIVITY IN THE SKIN TISSUE

Low molecular weight polyethylenimine (LMW-PEI) was linked to an expressing plasmid contains a green fluorescence protein (GFP) reporter gene and effective gene transfer was observed in CM7221 cell line tested. We examined the relationship among the molecular weight, structure of PEI and their transfection activity and cytotoxicity on CM7221 cell line. We also examined the position and continuance time of the GFP reporter gene expressed in the skin tissue of mouse. Results showed that LMW-PEI/DNA complexes led to high levels of expression in the CM7221 cell line (about 55%). However, with the increasing of PEI molecular weight, the transfection activity of PEI was decreasing. There was an increasing cytotoxicity with the larger PEI molecules. Further research showed that LMW-PEI induced a significant and long-lasting (7 days) expression of the GFP reporter gene, in the hair vesicle, sweat, gland, sebaceous gland in the mouse skin tissues. The LMW-PEI described here is a new, highly efficient and non-cytotoxic vector. It would be a useful non-viral vector for gene delivery technology, particular useful as simple skin-specific vehicles of therapeutic genes.