氟化聚乙烯亚胺衍生物构成的胶束载体的构建、表征及其在跨血-脑屏障递送中的作用研究

Construction and characterization of nanomicelle carriers composed of fluorinated polyethyleneimine derivatives and their role in delivery across the blood-brain barrier

目的:探讨由氟化聚乙烯亚胺(polyethyleneimine,PEI)衍生物构成的纳米胶束的构建、表征及其在跨血-脑屏障(blood-brain barrier,BBB)进行基因递送中的作用。方法:使用PEI和七氟丁酸酐(heptafluorobutyric anhydride,HFAA)通过化学反应合成PEI-HFAA,随后通过酰胺反应连接芥子酸(sinapic acid,SA)得到产物PEI-HFAA-SA(简称SPF),最终在SPF外包裹聚山梨酯80(polysorbate 80,PS80)得到最终产物PEI-HFAA-SA@PS80(简称SPFT)。通过傅里叶变换红外吸收光谱、核磁共振氟谱和核磁共振氢谱等对SPFT的分子键和元素组成进行分析,并通过动态光散射实验、琼脂糖凝胶阻滞实验和扫描电镜观察对其水合直径、质粒吸附及保护能力、载体质粒复合体的稳定性和形貌进行进一步表征。探究SPFT在小鼠神经胶质瘤细胞Neuro 2a中的基因转染效率和细胞毒性,通过尾静脉注射携带绿色荧光蛋白表达质粒的SPFT至C57BL/6J小鼠中,观察其在脑组织中的分布情况以及BBB内细胞的基因转染效果。结果:以SA和HFAA修饰以及PS80包裹的方法合成了SFPT。检测结果显示,SPFT水动力粒径100~200 nm,并且对质粒有一定的携带和保护作用。SPFT携带质粒在体外表现出良好的转染能力和生物相容性。体内实验显示,尾静脉注射后的SPFT在小鼠大脑中有聚集现象,能够携带质粒穿越BBB并进行基因递送。结论:SPFT具有一定生物相容性和良好的穿越BBB及基因递送能力,为脑部疾病的治疗药物递送提供了新的思路。

Objective:To investigate the construction,characterization,and role of nanomicelles composed of fluorinated polyethyleneimine(PEI)derivatives in gene delivery across the blood-brain barrier(BBB).Methods:PEI-heptafluorobutyric anhydride(HFAA)was synthesized through a chemical reaction between PEI and HFAA,followed by amide reaction with sinapic acid(SA)to obtain PEI-HFAA-SA(referred as SPF).Finally,PEI-HFAA-SA@PS80(referred as SPFT)was obtained by encapsulating polysorbitol 80(PS80)within SPF.The molecular bonds and elemental composition of SPFT were analyzed using Fourier transformation infrared absorption spectroscopy,fluorine nuclear magnetic resonance(NMR),and hydrogen NMR spectroscopy.The hydrodynamic particle size,plasmid adsorption and protection capacity,stability and morphology of the carrier-plasmid complex were further characterized by dynamic light scattering experiments,agarose coagulation experiments and scanning electron microscopy observations,respectively.The gene transfection efficiency and cytotoxicity of SPFT were investigated in mouse glioma cell line Neuro 2a.SPFT carrying green fluoresant protein expression plasmid was injected into C57BL/6J mice by tail vein to observe its distribution in brain tissues and the effect of gene transfection within the BBB.Results:SA and HFAA were modified to synthesize SFPT,which was then wrapped in PS80.SPFT had a hydrodynamic particle size of 100 to 200 nm while exhibiting significant loading capacity for plasmids along with effective protection against degradation.In vitro experiments revealed that SPFT possessed excellent transfection ability and biocompatibility.In vivo experiments showed that SPFT accumulated in the brain successfully and crossed the BBB to deliver the gene effectively after tail vein injection into mice.Conclusion:SPFT exhibits a good biocompatibility and demonstrates an efficient gene delivery across the BBB,presenting a novel approach for drug administration in neurological disorders.