Development of fusogenic liposomes which can deliver any substances into the cells through membrane fusion

许多研究者已经研究了脂质体作为将物质转运到细胞中的载体。然而，常规脂质体不直接与细胞膜融合。即使当它们通过在表面上添加抗体或配体而主动结合到细胞表面时，它们也通过内吞作用被摄入细胞。因此，大多数包封在脂质体中的物质被溶酶体酶降解，导致其效率损失。另一方面，仙台病毒的包膜含有负责与细胞膜结合和融合的蛋白质，仙台病毒基因通过膜融合被导入细胞。因此，引入的基因不被溶酶体酶分解。基于这些发现，开发了在脂质体表面上具有仙台病毒包膜蛋白的融合脂质体。融合脂质体可以通过直接膜融合将其包封的材料引入细胞质中。在这项研究中，我们检查了 关于我们 将仙台病毒颗粒与单脂质体（PA：PC：CHOL=1：4：5（mol））融合，制备了单层融合脂质体，并对融合脂质体的性质进行了研究。具有磷脂酸（PA）的负表面电荷的脂质体给出了最好的结果。胆固醇的量（30-50%）和唾液酸的存在不影响递送效率。此外，我们还根据融合脂质体颗粒密度的差异，对融合脂质体进行了纯化。纯化的融合脂质体在大小和结构上与仙台病毒颗粒相似，并且与仙台病毒一样有效地与细胞融合。这些脂质体可以与来自各种物种（人、猴、兔、犬、牛、小鼠和仓鼠）和来自各种组织（成纤维细胞、上皮细胞、内皮细胞、肝细胞、神经元细胞和淋巴瘤细胞）的细胞以几乎相同的效率融合。融合脂质体可以将任何可以包裹在脂质体中的物质（包括基因或蛋白质）导入细胞，是将遗传信息直接导入动物组织的新工具。少

Many investigators have examined liposomes as carriers for transporting substances into cells. However, conventional liposomes do not directly fuse with the cell membrane. Even when they are actively bound to the cell surface by adding an antibody or ligand on the surface, they are taken into the cell by endocytosis. Therefore, most substances encapsulated in liposomes are degraded by lysosome enzymes, resulting in loss of their efficiency. On the other hand, the envelope of the Sendai virus contains a protein responsible for binding and fusion to the cell membrane, and Sendai virus genes are introduced into cells via membrane fusion. Therefore, the introduced genes are not decomposed by lysomome enzymes. Based on these findings, fusogenic liposomes which possess the envelope proteins of the Sendai virus on the liposomal surface were developed. Fusogenic liposomes can introduce their encapsulated materials into the cytoplasm through direct membrane fusion. In this study, we examined th … More e preparation and characterization of unilamellar fusogenic liposome, and the efficiency of delivery of proteins and DNA by unilamellar fusogenic liposomes.A fusogenic liposome was generated by fusion between a Sendai virus particle and a simple liposome (PA : PC : CHOL=1 : 4 : 5 (mol)). The liposomes with a negative surface charge of phosphatidic acid (PA) gave the best results. The amount of cholesterol (30-50%) and the presence of sialic acid did not affect the efficiency of delivery. In addition, we purified fusogenic liposomes based on the difference of the density of the particles. The purified fusogenic liposomes are similar in size and structure to the Sendai virus particle, and fuse with cells as efficiently as the Sendai virus. These liposomes can fuse with cells derived from various species (human, monkey, rabbit, canine, cattle, mouse and hamster) and from various tissues (fibroblasts, epithelial cells, endothelial cells, hepatocytes, neuronal cells and lymphoma cells) with almost the same efficiency. They can deliver any substances that can be encapsulated in the liposomes, including genes or proteins, into the cells.In conclusion, the fusogenic lipsomes are new tools for direct introduction of genetic information into animal tissues. Less