Ricchelli F, Jori G, Gobbo S, Tronchin M
C.N.R. Centre of Hemocyanins and other Metalloproteins, Department of Biology, University of Padova, Italy.
Biochim Biophys Acta. 1991 May 31;1065(1):42-8. doi: 10.1016/0005-2736(91)90008-v.
Unilamellar liposomes of dipalmitoylphosphatidylcholine (DPPC) have been chosen as suitable models of cell membranes in studies aimed at defining the influence of specific parameters on the distribution properties of selected hydrophobic photosensitizers, namely hematoporphyrin (HP) and protoporphyrin (PP), in normal and tumour tissues. To better mimic in vivo situations, DPPC liposomes were sometimes mixed with cardiolipin (Card) or cholesterol (Chol). Two techniques were mainly used: the quenching of porphyrin fluorescence by methyl viologen, which can discriminate different dye populations inside the vesicles as well as their degree of accessibility to the external medium, and the polarization of porphyrin fluorescence, which gives information on the dye microenvironment through its degree of rotational freedom. The nature of the porphyrin binding sites in each phospholipid monolayer is found to be a function of the degree of hydrophobicity and the concentration of the dye as well as the chemical composition of the liposomes. In DPPC and DPPC-Chol liposomes, all PP molecules are deeply embedded into very rigid, hydrophobic domains of the inner lipid monolayer. Only in the presence of cardiolipin, for [PP] greater than 2.5 microM, a partial shift of the dye molecules towards the outer lipid monolayer is observed. HP mostly localizes at the inner lipid/water interface in all liposomes: at very low concentrations ([HP] approximately equal to 0.5 microM) the dye is bound to the polar heads of the lipids through its carboxylate groups, leaving the rest of the molecule dissolved in the inner aqueous pool. At higher concentrations, HP molecules change their orientation: the ionized propionic chains still interact with the polar heads while the hydrophobic core lies in the lipid phase in DPPC and DPPC-Card vesicles. HP incorporated into DPPC-Chol mixed liposomes projects from the inner lipid phase into the aqueous compartment in all the concentration range studied by us. A very small fraction of HP population (corresponding to 5-10% of the overall fluorescence) is localized at the water/lipid external interface in DPPC and DPPC-Chol liposomes. This fraction increases in the presence of cardiolipin (up to 30% of the overall fluorescence). The possible implications of these findings for the nature of the targets of photosensitization in cell membranes are discussed.
在旨在确定特定参数对选定的疏水性光敏剂（即血卟啉（HP）和原卟啉（PP））在正常组织和肿瘤组织中分布特性影响的研究中，二棕榈酰磷脂酰胆碱（DPPC）的单层脂质体已被选为合适的细胞膜模型。为了更好地模拟体内情况，DPPC脂质体有时会与心磷脂（Card）或胆固醇（Chol）混合。主要使用了两种技术：通过甲基紫精淬灭卟啉荧光，其可以区分囊泡内不同的染料群体以及它们对外部介质的可及程度；以及卟啉荧光的偏振，其通过染料的旋转自由度给出关于染料微环境的信息。发现每个磷脂单层中卟啉结合位点的性质是疏水性程度、染料浓度以及脂质体化学成分的函数。在DPPC和DPPC-Chol脂质体中，所有PP分子都深深嵌入内脂质单层的非常刚性的疏水区域。仅在心磷脂存在下，当[PP]大于2.5 microM时，观察到染料分子部分向外脂质单层转移。在所有脂质体中，HP大多位于内脂质/水界面：在非常低的浓度（[HP]约等于0.5 microM）下，染料通过其羧基与脂质的极性头部结合，分子的其余部分溶解在内水相中。在较高浓度下，HP分子改变其取向：离子化的丙酸链仍与极性头部相互作用，而疏水核心位于DPPC和DPPC-Card囊泡的脂质相中。在我们研究的所有浓度范围内，掺入DPPC-Chol混合脂质体中的HP从内脂质相突出到水相中。在DPPC和DPPC-Chol脂质体中，非常小部分的HP群体（对应于总荧光的5-10%）位于水/脂质外界面。在心磷脂存在下，这一比例增加（高达总荧光的30%）。讨论了这些发现对细胞膜中光致敏靶点性质的可能影响。
