Zeisig R, Shimada K, Hirota S, Arndt D
Max-Delbrück Center for Molecular Medicine, Berlin, Germany,
Biochim Biophys Acta. 1996 Dec 4;1285(2):237-45. doi: 10.1016/s0005-2736(96)00167-8.
A serious problem using liposomes for therapeutic purposes is the fast removal from blood circulation by components of the reticuloendothelial system (RES) most likely after opsonization of the vesicles. This study was performed to quantify the reduction in macrophage uptake in vitro of sterically stabilized liposomes (PEG-liposomes) prepared from hexadecylphosphocholine, cholesterol and poly(ethylene glycol2000) distearoylphosphoethanolamine (PEG2000DSPE) for the first time. The uptake was determined using HPC-liposomes of different defined size (125, 250 and 1000 nm) without and with sterical stabilization by incorporating 5 mol% of PEG2000DSPE. HPTS was used as fluorescence marker allowing the discrimination between general uptake and the part of liposomes internalized into the low pH-compartment (Daleke, L.D., Hong, K. and Papahadjopoulos. D. (1990) Biochim. Biophys. Acta 1024, 352-366). Liposomal uptake by J774 mouse macrophage-like cells was time-dependent. Both the uptake and internalization were clearly reduced for PEG-liposomes compared to plain liposomes. Sterical stabilization reduced the general uptake of liposomes in vitro by more than 50% and the internalization by about 50-60%. PEG-liposomes additionally showed a delay in internalization into the macrophages during the first 6 h. Size of used liposomes had only a minor influence on liposomal uptake but highest concentration of lipid was found for large multilammelar vesicles (MLV). The fixed aqueous layer thickness (FALT) was determined by zeta potential measurements of plain and sterically stabilised HPC-liposomes (100 nm) in solutions of different ion concentrations. The calculation of the thickness was based on the linear correlation between ln zeta (zeta-potential) and kappa (Debye Hückel-Parameter). FALT was calculated and found to be for plain HPC-liposomes 0.83 +/- 0.17 nm and for PEG-HPC-liposomes 3.57 +/- 0.17 nm. Exchange of the HPC by an alkylphospholipid with different head group has no or only minor effect (PEG-OPP-liposomes 3.44 +/- 0.31 nm). Thus the reduced uptake of HPC-LUVET correlates with an increased thickness of the fixed aqueous layer around these liposomes and could support the hypothesis that the thickness is an important property responsible for preventing opsonization and resulting finally in a reduced macrophage uptake.
将脂质体用于治疗目的的一个严重问题是，在囊泡最有可能被调理素化后，它们会被网状内皮系统（RES）的成分迅速从血液循环中清除。本研究首次对由十六烷基磷胆碱、胆固醇和聚（乙二醇2000）二硬脂酰磷脂酰乙醇胺（PEG2000DSPE）制备的空间稳定脂质体（PEG脂质体）在体外巨噬细胞摄取的减少进行了量化。通过使用不同确定尺寸（125、250和1000nm）的HPC脂质体，在添加和不添加5mol%PEG2000DSPE进行空间稳定的情况下，测定摄取情况。HPTS用作荧光标记物，以区分一般摄取和内化到低pH区室的脂质体部分（Daleke，L.D.，Hong，K.和Papahadjopoulos，D.（1990）Biochim.Biophys.Acta 1024，352 - 366）。J774小鼠巨噬细胞样细胞对脂质体的摄取是时间依赖性的。与普通脂质体相比，PEG脂质体的摄取和内化明显减少。空间稳定使脂质体在体外的一般摄取减少了50%以上，内化减少了约50 - 60%。PEG脂质体在最初6小时内还表现出内化到巨噬细胞中的延迟。所用脂质体的大小对脂质体摄取的影响较小，但大单层囊泡（MLV）的脂质浓度最高。通过在不同离子浓度的溶液中对普通和空间稳定的HPC脂质体（100nm）进行zeta电位测量来确定固定水层厚度（FALT）。厚度的计算基于ln zeta（zeta电位）和kappa（德拜休克尔参数）之间的线性相关性。计算得出普通HPC脂质体的FALT为0.83±0.17nm，PEG - HPC脂质体的FALT为3.57±0.17nm。用具有不同头部基团的烷基磷脂替换HPC没有或只有很小的影响（PEG - OPP脂质体为3.44±0.31nm）。因此，HPC - LUVET摄取的减少与这些脂质体周围固定水层厚度的增加相关，这可能支持这样的假设，即厚度是防止调理素化并最终导致巨噬细胞摄取减少的一个重要特性。
