Department of Pharmaceutics, Institute of Pharmacy, University of Bonn, 53121 Bonn, Germany.
Department of Neurosurgery, University of Bonn, Bonn, Germany.
Colloids Surf B Biointerfaces. 2020 Oct;194:111193. doi: 10.1016/j.colsurfb.2020.111193. Epub 2020 Jun 24.
Direct cytoplasmic delivery is essential for susceptible molecules as proteins and some nucleic acids to improve their therapeutic efficacy in cells. Using liposomes for their delivery proved challenging due to known uptake by endocytosis followed by partial or complete lysosomal breakdown. Thus, "fusogenic" liposomes (FL) composed of the neutral lipid dioleoylphosphatidylethanolamine (DOPE) combined with the cationic lipid 1, 2-dioleoyl-3-trimethylammoniumpropane (DOTAP) were tested in different ratios for their cell membrane fusion ability and their cytoplasmic delivery was compared to "pH-sensitive" liposomes in murine brain endothelial cells (bEnd.3). They were loaded with cargos of different molecular sizes (calcein/ enhanced green fluorescent-protein (EGFP)/ EGFP coding plasmid) and their intracellular delivery was quantitatively and qualitatively analyzed. FL composed of equimolar ratios of DOPE and DOTAP showed the most efficient cytoplasmic delivery of all cargos by fusing with the cell membranes within the first 15 min of addition. Their EGFP plasmid delivery to cells was quantified to be 58.2 ± 9.5 % of the total EGFP load and calcein delivery was measured in buffer to be 64.1 ± 4.0 % of the total calcein load, and reduced in blood to 26.1 ± 0.6 %. Thus our tested FL allowed a fast and abundant cytoplasmic delivery of cargos independent of their molecular sizes while avoiding endocytosis, although they also underwent fast fusion with erythrocytes. Seemingly, these carriers could be used as a powerful delivery tool for in-vitro purposes.
直接细胞质递送对于蛋白质和一些核酸等易感性分子至关重要，可提高其在细胞中的治疗效果。由于内吞作用后的部分或完全溶酶体分解，使用脂质体进行递送被证明具有挑战性。因此，测试了由中性脂质二油酰基磷脂酰乙醇胺（DOPE）与阳离子脂质 1,2-二油酰基-3-三甲基铵丙烷（DOTAP）组成的“融合脂质体”（FL）在不同比例下的细胞膜融合能力，并将其与“pH 敏感”脂质体在小鼠脑内皮细胞（bEnd.3）中的细胞质递送进行了比较。它们被装载了不同分子大小的货物（钙黄绿素/增强型绿色荧光蛋白（EGFP）/EGFP 编码质粒），并对其细胞内递送进行了定量和定性分析。由 DOPE 和 DOTAP 等摩尔比组成的 FL 在添加后的最初 15 分钟内与细胞膜融合，显示出所有货物中最有效的细胞质递送。它们向细胞中递送 EGFP 质粒的定量为总 EGFP 负载的 58.2±9.5%，缓冲液中递送钙黄绿素的定量为总钙黄绿素负载的 64.1±4.0%，在血液中减少到 26.1±0.6%。因此，我们测试的 FL 允许快速而丰富的细胞质递送货物，而与货物的分子大小无关，同时避免了内吞作用，尽管它们也与红细胞快速融合。显然，这些载体可作为一种强大的体外递送工具。
