Optimization of the Electroformation of Giant Unilamellar Vesicles (GUVs) with Unsaturated Phospholipids
Optimization of the Electroformation of Giant Unilamellar Vesicles (GUVs) with Unsaturated Phospholipids
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DOI：
10.1007/s00232-015-9828-3
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发表时间：
2015-10-01
影响因子：
2.4
通讯作者：
Mir, Lluis M.
中科院分区：
文献类型：
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作者：
Breton, Marie;Amirkavei, Mooud;Mir, Lluis M.
巨单层囊泡（GUV）是广泛使用的细胞膜模型。GUV具有细胞样直径，并且含有构成细胞膜的相同磷脂。获得这些囊泡最常用的方案被称为电形成，因为该方案的关键步骤在于将电场施加到磷脂存款上。尚未考虑由于施加电场而导致的不饱和磷脂的潜在氧化，尽管GUV膜中氧化脂质的存在可能影响其渗透性和机械性能。通过质谱分析，我们证明了电形成技术可以导致构成囊泡的多不饱和磷脂氧化。然后，使用流式细胞术，我们表明电场的幅度和持续时间影响囊泡的数量和大小。根据我们的研究结果，磷脂的氧化水平随着它们的不饱和度以及电场的振幅和持续时间而增加。然而，当脂质氧化水平超过25%时，囊泡的直径减小，并且当脂质氧化水平达到40%时，囊泡破裂或重组，并且它们的生产速率降低。总之，经典的电铸方法应始终优化，作为所用磷脂的函数，特别是用于生产用作细胞膜模型的多不饱和磷脂的巨型脂质体。
Giant unilamellar vesicles (GUV) are widely used cell membrane models. GUVs have a cell-like diameter and contain the same phospholipids that constitute cell membranes. The most frequently used protocol to obtain these vesicles is termed electroformation, since key steps of this protocol consist in the application of an electric field to a phospholipid deposit. The potential oxidation of unsaturated phospholipids due to the application of an electric field has not yet been considered even though the presence of oxidized lipids in the membrane of GUVs could impact their permeability and their mechanical properties. Thanks to mass spectrometry analyses, we demonstrated that the electroformation technique can cause the oxidation of polyunsaturated phospholipids constituting the vesicles. Then, using flow cytometry, we showed that the amplitude and the duration of the electric field impact the number and the size of the vesicles. According to our results, the oxidation level of the phospholipids increases with their level of unsaturation as well as with the amplitude and the duration of the electric field. However, when the level of lipid oxidation exceeds 25 %, the diameter of the vesicles is decreased and when the level of lipid oxidation reaches 40 %, the vesicles burst or reorganize and their rate of production is reduced. In conclusion, the classical electroformation method should always be optimized, as a function of the phospholipid used, especially for producing giant liposomes of polyunsaturated phospholipids to be used as a cell membrane model.