Methyl-β-cyclodextrins and liposomes as water-soluble carriers for cholesterol incorporation into membranes and its evaluation by a microenzymatic fluorescence assay and membrane fluidity-sensitive dyes
Methyl-β-cyclodextrins and liposomes as water-soluble carriers for cholesterol incorporation into membranes and its evaluation by a microenzymatic fluorescence assay and membrane fluidity-sensitive dyes
复制标题
DOI：
10.1006/abio.1998.2594
复制
发表时间：
1998-05-01
影响因子：
2.9
通讯作者：
Ojeda, F
中科院分区：
文献类型：
--
作者：
Härtel, S;Diehl, HA;Ojeda, F
我们测试了一种基于装载胆固醇的甲基- β -环糊精的装载方法，并将其与使用装载胆固醇的脂质体的方法进行了比较，使用甲基- β -环糊精，我们将微粒体膜中的胆固醇含量增加到原始含量的近四倍。用含胆固醇的脂质体代替，我们实现了140%的升高。为了直接检测膜胆固醇，我们对最初用于精确检测血清胆固醇的微酶荧光法进行了轻微修改。由于不需要进行脂质提取，该方法对脂质体和微粒体中的胆固醇检测是可靠的。此外，我们比较了芘、芘-甲醇、二芘、1-6-苯基-1,3,5，-六atrien和1-（4-三甲基氨基苯基）-6-苯基-1,3,5，-六atrien这三种荧光染料的灵敏度，以便通过施加在脂质基质上的动态相关变化间接检测胆固醇。这些染料不仅在膜上的位置不同，而且它们的动力学行为也不同。我们校准了脂质体中确定胆固醇含量的染料，并使用最合适的染料来跟踪和量化脂质体和微粒体膜中的胆固醇掺入。(C) 1998学术出版社。
A variety of methods to incorporate cholesterol into Lipid membrane systems have been applied with varying success, We tested an incorporation method based on cholesterol-loaded methyl-beta-cyclodextrins and compared it to a method that uses cholesterol-loaded liposomes, With methyl-beta-cyclodextrin, we increased the cholesterol content in microsomal membranes to almost the fourfold of the original content. With cholesterol-loaded liposomes instead, we achieved an elevation of 140%. Short incubation times and well-defined carrier properties favor the beta-cyclodextrin method, For direct detection of membrane cholesterol, we slightly modified a microenzymatic fluorescence assay originally developed for precise cholesterol detection in serum. Without the need to perform lipid extraction, this assay was reliable for cholesterol detection in liposomes and in microsomes. Additionally, we compared the sensitivity of the fluidity-sensitive fluorescent dyes pyrene, pyrene-methanol, bis-pyrene, 1-6-phenyl-1,3,5,-hexatrien, and 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5,-hexatrien in order to detect cholesterol indirectly by the dynamically relevant changes exerted on lipid matrices. These dyes differ not only in their membrane location but also in their dynamical behavior. We calibrated the dyes in Liposomes of defined cholesterol content and used the most suited ones to follow and quantify the cholesterol incorporation into liposomal and microsomal membranes. (C) 1998 Academic Press.