叶酸修饰的聚乙烯亚胺-聚乙二醇共聚物载体在体内外转染效率的研究

目的探讨叶酸修饰的不同相对分子质量聚乙酰亚胺构建的聚乙烯亚胺-聚乙二醇5000(PEI-PEG)作为基因载体在体内外的转染效率。方法在体外实验中,分别测试3个相对分子质量FPPs(branched PEI:0.6kDa,10kDa和Linear PEI 25kDa)在最佳N/P、有无血清、血清的浓度和叶酸受体靶向性等条件下对口腔癌上皮细胞(KB)转染率的影响;在体内实验中,FPP(0.6kDa)分别压缩pLuc-p53基因形成的复合物通过瘤内注射和尾静脉注射2种方式考察聚合物的转染效率。结果在体外实验中,转染率最佳条件分别是N/P=20、有血清且转染率随着白蛋白(ALB)或胎牛血清(FBS)浓度的增加有上升趋势;其中3个相对分子质量PEI组成的FPP转染率分别为FPP(10kDa>25kDa>0.6kDa);FPP有较强的叶酸靶向性;在体内实验中,与BPP/pLuc和PEI(25kDa)/pLuc相比,FPP(10kDa)/pLuc呈现出较高的转染率;在48h内,随着时间的推移,转染效率增加;FPP(10kDa)/p53能改善荷瘤小鼠;与裸pLuc相比,3种相对分子质量的FPPs没有毒性。结论具有靶向性的FPP非病毒载体能成为临床传递基因治疗载体的候选。

Study of transgene efficiency of folate modified polyethyleneimine-polyethylene gly-col in vit ro and in vivo

Objective To investigate the transgene efficiency mediated by folate modified different molecular weight polyethylenei‐mine‐polyethylene glycol5000 (PEI‐PEG) in vitro and in vivo .Methods In vitro ,the ratio of N/P in complexes ,medium with serum or without serum ,serum concentration ,and folate receptors targeting were studied for gene transfection efficiency of three molecu‐lar weight folate modified PEI (branched 0 .6 kDa ,10 kDa and Linear 25 kDa)‐PEG (FPP) in KB cells .In vivo ,the transfection efficiency of FPP(10 kDa) compressed pLuc/p53 was investigated through intratumoral injection and tail vein injection in bearing tumor nude mice .Expression of Caspase‐9 and Bax in treated nude mice was researched by western‐bolting .Results The higher transfection efficiency exhibited in FPP(10 kDa) when N/P=20 and in serum ,and it was increasing with serum concentration .Fur‐thermore ,the transfection efficiency of FPP was FPP (10 kDa>25 kDa>0 .6 kDa) .Meanwhile ,FPP(10 kDa) had folate receptors tar‐geting in KB cells .In vivo ,higher transfection efficiency of FPP(10 kDa) compressed pLuc/p53 were gained though intratumour and tail vein injection ,and FPP(10 kDa)/p53 can improve bearing tumor nude mice .Transfection efficiency of all non‐virus vectors was increased with time ongoing .FPPs had almost no toxicity .Conclusion Targeted FPPs could be used as a clinic gene delivery sys‐tem in future .