Pantazatos D P, Pantazatos S P, MacDonald R C
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, IL 60208-3500, USA.
J Membr Biol. 2003 Jul 15;194(2):129-39. doi: 10.1007/s00232-003-2031-y.
Cationic, O-alkylphosphatidylcholines, recently developed as DNA transfection agents, form bilayers indistinguishable from those of natural phospholipids and undergo fusion with anionic bilayers. Membrane merging (lipid mixing), contents release, and contents mixing between populations of positive vesicles containing O-ethylphosphatidylcholine (EDOPC) and negative vesicles containing dioleolylphosphatidylglycerol (DOPG) have been determined with standard fluorometric vesicle-population assays. Surface-charge densities were varied from zero to full charge. All interactions depended critically on surface-charge density, as expected from the adhesion-condensation mechanism. Membrane mixing ranged from zero to 100%, with significant mixing (>10 <70%) occurring between cationic vesicles that were fully charged and anionic vesicles that had fractional surface charges as low as 0.1. Such mixing with membranes as weakly charged as cell membranes should be relevant to transfection with cationic lipids. Unexpectedly, lipid mixing was higher at high than at low ionic strength when one lipid dispersion was prepared from EDOPC plus DOPG (in different proportions), especially when the other vesicles were of EDOPC; this may somehow be a consequence of the ability of the former mixture to assume non-lamellar phases. Leakage of aqueous contents was also a strong function of charge, with fully charged vesicles releasing essentially all of their contents less than 1 min after mixing. EDOPC was more active in this regard than was DOPG, which probably reflects stronger intermolecular interactions of DOPG. Fusion, as measured by contents mixing, exhibited maximal values of 10% at intermediate surface charge. Reduced fusion at higher charge is attributed to multiple vesicle interactions leading to rupture. The existence of previously published data on individual interactions of vesicles of the same composition made it possible for the first time to compare pairwise with population interactions, confirming the likelihood of population studies to overestimate rupture and hemifusion and underestimate true vesicle fusion.
阳离子型O - 烷基磷脂酰胆碱最近被开发用作DNA转染试剂，它形成的双层膜与天然磷脂的双层膜难以区分，并能与阴离子双层膜发生融合。含有O - 乙基磷脂酰胆碱（EDOPC）的正性囊泡群体与含有二油酰磷脂酰甘油（DOPG）的负性囊泡群体之间的膜融合（脂质混合）、内容物释放以及内容物混合，已通过标准荧光囊泡群体分析方法得以确定。表面电荷密度从零变化到完全带电。正如粘附凝聚机制所预期的那样，所有相互作用都严重依赖于表面电荷密度。膜混合程度从零到100%不等，在完全带电的阳离子囊泡与表面电荷分数低至0.1的阴离子囊泡之间发生显著混合（>10%<70%）。这种与像细胞膜一样弱带电的膜的混合应该与阳离子脂质转染相关。出乎意料的是，当一种脂质分散体由EDOPC加DOPG（不同比例）制备时，在高离子强度下的脂质混合比在低离子强度下更高，特别是当另一种囊泡是EDOPC时；这可能在某种程度上是前者混合物能够呈现非层状相的结果。水性内容物的泄漏也是电荷的强函数，完全带电的囊泡在混合后不到1分钟内基本上释放了所有内容物。在这方面，EDOPC比DOPG更活跃，这可能反映了DOPG更强的分子间相互作用。通过内容物混合测量的融合在中等表面电荷时表现出最大值10%。在较高电荷时融合减少归因于多个囊泡相互作用导致破裂。先前已发表的关于相同组成囊泡个体相互作用的数据首次使得能够将成对相互作用与群体相互作用进行比较，证实了群体研究可能高估破裂和半融合并低估真正囊泡融合的可能性。
