CAJ | 학술논문

目的考察烷基化低分子量聚乙烯亚胺作为基因递送载体的安全性和有效性。 　　方法以1-溴十二烷和分子量为1800的分枝状 PEI （bPEI1.8K）为原料，合成 bPEI1.8K-C12，并用1H-NMR 对其结构进行确认；采用 MTT 法考察 bPEI1.8K-C12的细胞毒性；红细胞溶血实验考察 bPEI1.8K-C12的生物相容性；测定bPEI1.8K-C12/DNA 复合颗粒的粒径分布和 zeta 电位；采用激光共聚焦显微镜观察 bPEI1.8K-C12/DNA 复合颗粒的细胞摄取行为；采用琼脂糖凝胶阻滞电泳考察 bPEI1.8K-C12对DNA 的固缩能力；并用荧光素酶报告基因和绿色荧光蛋白报告基因考察 bPEI1.8K-C12的体外转染效率。 　　结果经1H-NMR 确认，成功合成了 bPEI1.8K-C12；MTT结果表明 bPEI1.8K-C12对人乳腺癌 MCF-7细胞的毒性与bPEI1.8K 相当；红细胞溶血实验结果表明高浓度 bPEI1.8K-C12静脉注射时具有潜在溶血性；质量比相同时，bPEI1.8K-C12/DNA 复合颗粒的平均粒径和 zeta 电位均比相应的bPEI1.8K/DNA 大；烷基化修饰后，bPEI1.8K-C12对 DNA 的固缩能力降低，MCF-7细胞对 bPEI1.8K-C12/DNA 复合颗粒的摄取效率大大增加，bPEI1.8K-C12递送报告基因质粒的体外转染效率显著高于 bPEI1.8K，甚至与 lipofectamine2000相当。 　　结论 bPEI1.8K-C12是一种安全高效的基因递送载体，具有较高的进一步开发前景。
목적고찰완기화저분자량취을희아알작위기인체송재체적안전성화유효성。 　　방법이1-추십이완화분자량위1800적분지상 PEI （bPEI1.8K）위원료，합성 bPEI1.8K-C12，병용1H-NMR 대기결구진행학인；채용 MTT 법고찰 bPEI1.8K-C12적세포독성；홍세포용혈실험고찰 bPEI1.8K-C12적생물상용성；측정bPEI1.8K-C12/DNA 복합과립적립경분포화 zeta 전위；채용격광공취초현미경관찰 bPEI1.8K-C12/DNA 복합과립적세포섭취행위；채용경지당응효조체전영고찰 bPEI1.8K-C12대DNA 적고축능력；병용형광소매보고기인화록색형광단백보고기인고찰 bPEI1.8K-C12적체외전염효솔。 　　결과경1H-NMR 학인，성공합성료 bPEI1.8K-C12；MTT결과표명 bPEI1.8K-C12대인유선암 MCF-7세포적독성여bPEI1.8K 상당；홍세포용혈실험결과표명고농도 bPEI1.8K-C12정맥주사시구유잠재용혈성；질량비상동시，bPEI1.8K-C12/DNA 복합과립적평균립경화 zeta 전위균비상응적bPEI1.8K/DNA 대；완기화수식후，bPEI1.8K-C12대 DNA 적고축능력강저，MCF-7세포대 bPEI1.8K-C12/DNA 복합과립적섭취효솔대대증가，bPEI1.8K-C12체송보고기인질립적체외전염효솔현저고우 bPEI1.8K，심지여 lipofectamine2000상당。 　　결론 bPEI1.8K-C12시일충안전고효적기인체송재체，구유교고적진일보개발전경。
Objective Preliminary evaluation of the safety and efficiency of alkylated low molecular polyethylenimine as a gene delivery vector. Methods bPEI1.8K-C12 was synthesized with branch PEI (Mw 1800; bPEI1.8K) and 1-bromododecane, and structurally confirmed with 1H-NMR. Cytotoxicity and biocompatibility of bPEI1.8K-C12 were evaluated by MTT method and hemolysis assay, respectively. Size distribution and zeta potential of bPEI1.8K-C12/DNA polyplexes were determined, and cellular uptake of the DNA polyplexes was visualized with confocal laser scanning microscope (CLSM). DNA condensation ability of bPEI1.8K-C12 was studied by gel retardation assay, and in vitro gene delivery efficiency was investigated with luciferase and green fluorescent protein reporter gene, respectively. Results bPEI1.8K-C12 was successfully synthesized after confirmation by 1H-NMR. Cytotoxicity of bPEI1.8K-C12 on MCF-7 cells was as low as bPEI1.8K, and systematic delivery of high dose bPEI1.8K-C12 may cause hemolysis in vivo. The average size of bPEI1.8K-C12/DNA polyplexes was bigger than that of bPEI1.8K/DNA polyplexes at the same weight ratio, and the zeta potential of bPEI1.8K-C12/DNA polyplexes was higher than that of bPEI1.8K/DNA polyplexes at the same weight ratio. After alkylation, the DNA condensation ability of bPEI1.8K-C12 was reduced, but the cellular uptake of bPEI1.8K-C12/DNA polyplexes was greatly enhanced. In vitro gene transfection efficiency of bPEI1.8K-C12/DNA was significantly higher than that of bPEI1.8K, and was comparable to that of lipofectamine2000. Conclusion bPEI1.8K-C12 is a safe and efficient gene delivery vector, and holds great promise for further development.