DNA TRANSFECTION MEDIATED BY CATIONIC LIPOSOMES CONTAINING LIPOPOLYLYSINE - CHARACTERIZATION AND MECHANISM OF ACTION
DNA TRANSFECTION MEDIATED BY CATIONIC LIPOSOMES CONTAINING LIPOPOLYLYSINE - CHARACTERIZATION AND MECHANISM OF ACTION
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DOI：
10.1016/0005-2736(94)90066-3
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发表时间：
1994-01-19
影响因子：
3.4
通讯作者：
HUANG, L
中科院分区：
文献类型：
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作者：
ZHOU, XH;HUANG, L
聚阳离子脂质，脂聚（L-赖氨酸）（LPLL）介导有效DNA转染的能力取决于处理细胞的刮擦（Zhou等人（1991）Biochim. Biophys. Acta 1065，8-14）。发现通过在脂质体组合物中包括辅助脂质可以避免机械处理。在测试的辅助脂质中，六方相形成脂质，二油酰磷脂酰乙醇胺（DOPE），产生了最高的活性。通过改变LPLL的亲脂性和阳离子脂质体与DNA的比例进一步优化转染效率。最佳的DNA-脂质体复合物的转染活性提高了6倍，如果细胞用干扰内吞过程的试剂进行预处理。同时，用抑制膜融合的肽预处理细胞使活性降低约60%。这些结果表明，DNA-脂质体复合物被内吞机制吸收，并且DNA的细胞质递送涉及可能在内体隔室中的融合相关事件。转染过程通过薄切片电子显微镜观察。结果发现，复合物进入细胞质主要是通过破坏内体稳定，偶尔通过穿透质膜。因此，我们的研究结果不同于先前的假设，该假设表明转染是通过脂质体与被处理细胞的质膜的融合介导的。
The ability of a polycationic lipid, lipopoly(L-lysine) (LPLL), to mediate efficient DNA transfection depended on scraping of the treated cells (Zhou et al. (1991) Biochim. Biophys. Acta 1065, 8-14). It was found that the mechanical treatment could be avoided by including a helper lipid to the liposome composition. Among the helper lipids tested, a hexagonal phase forming lipid, dioleoylphosphatidylethanolamine (DOPE), gave rise to the highest activity. The transfection efficiency was further optimized by varying the lipophilicity of the LPLL and the ratio of the cationic liposome to DNA. Transfection activity of the optimal DNA-liposome complexes was enhanced by up to 6-fold if cells were pretreated with agents interfering with the process of endocytosis. Meanwhile, pretreatment of cells with a peptide which inhibits membrane fusion decreased the activity by about 60%. These results indicated that DNA-liposome complexes are taken up by an endocytosis mechanism and that cytoplasmic delivery of DNA involves a fusion-related event probably in the endosome compartment. The transfection process was visualized by thin-section electron microscopy. It was found that the complexes entered the cytoplasm mainly by destabilizing endosomes and occasionally by penetrating through the plasma membrane. Therefore, our findings differ from a previous hypothesis which suggests that transfection is mediated by fusion of the liposomes with the plasma membrane of the treated cells.