Octaarginine- and octalysine-modified nanoparticles have different modes of endosomal escape
Octaarginine- and octalysine-modified nanoparticles have different modes of endosomal escape
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DOI：
10.1074/jbc.m709387200
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发表时间：
2008-08-22
影响因子：
4.8
通讯作者：
Harashima, Hideyoshi
中科院分区：
文献类型：
--
作者：
El-Sayed, Ayman;Khalil, Ikramy A.;Harashima, Hideyoshi
本研究探讨了表面修饰与octagrinine肽（R8）的脂质体逃逸内吞囊泡的作用，使用octalysine（K8）作为控制阳离子肽;脂质体的内体逃逸的机制也进行了研究。包封在R8修饰的纳米颗粒中的浓缩质粒DNA的基因表达比K8修饰的纳米颗粒高1个数量级以上，并且比使用未修饰的纳米颗粒的基因表达高2个数量级。基因表达的差异不能归因于摄取的差异，因为R8和K8修饰的脂质体主要通过巨胞饮作用以相当的效率被摄取。R8-纳米颗粒逃逸到胞质溶胶的程度是K8-纳米颗粒的两倍。内体酸化的抑制抑制了R8-纳米颗粒内体逃逸，但增强了K8-纳米颗粒内体逃逸。使用活细胞中的光谱成像，我们表明R8和K8脂质体通过脂质体与内体膜之间的融合从内吞囊泡中逃脱。R8-脂质体在酸性和中性pH下有效融合，而K8-脂质体仅在中性pH下融合。在体外脂质混合和钙黄绿素释放实验中观察到类似的行为。细胞与不同标记的K8和R8修饰的纳米颗粒的共孵育证实了共同的摄取途径和不同的内体逃逸速率，特别是在较长的时间间隔。因此，可以得出结论，脂质体表面上的R8通过在中性和酸性pH下起作用的融合机制刺激从内吞囊泡有效逃逸;相反，K8主要在中性pH下介导逃逸。
The present study examines the role of surface modification with an octaarginine peptide (R8) in liposomal escape from endocytic vesicles, using octalysine (K8) as a control cationic peptide; the mechanism of endosomal escape of liposomes was also investigated. Gene expression of condensed plasmid DNA encapsulated in R8-modified nanoparticles was more than 1 order of magnitude higher than that of K8-modified nanoparticles, and 2 orders of magnitude higher than gene expression using unmodified nanoparticles. The difference in gene expression could not be attributed to differences in uptake, as R8- and K8-modified liposomes were taken up primarily via macropinocytosis with comparable efficiency. The extent of R8-nanoparticle escape to the cytosol was double that of K8-nanoparticles. Suppression of endosome acidification inhibited R8-nanoparticle endosomal escape, but enhanced that of K8-nanoparticles. Using spectral imaging in live cells, we showed that R8- and K8-liposomes escaped from endocytic vesicles via fusion between the liposomes and the endosomal membrane. R8- liposomes fused efficiently at both acidic and neutral pH, whereas K8- liposomes fused only at neutral pH. Similar behavior was observed during in vitro lipid mixing and calcein-release experiments. Co-incubation of cells with distinctly labeled K8- and R8-modified nanoparticles confirmed a common uptake pathway and different rates of endosomal escape particularly at longer time intervals. Therefore, it was concluded that R8 on the liposome surface stimulates efficient escape from endocytic vesicles via a fusion mechanism that works at both neutral and acidic pH; in contrast, K8 mediates escape mainly at neutral pH.