Spontaneous intermembrane transfer of various cholesterol-derived hydroperoxide species: kinetic studies with model membranes and cells.
Spontaneous intermembrane transfer of various cholesterol-derived hydroperoxide species: kinetic studies with model membranes and cells.
复制标题
各种胆固醇衍生的氢过氧化物种类的自发膜间转移：模型膜和细胞的动力学研究。
DOI：
10.1021/bi011408r
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发表时间：
2001
期刊：
影响因子：
2.9
通讯作者：
Girotti,AW
中科院分区：
文献类型：
--
作者：
Vila,A;Korytowski,W;Girotti,AW
虽然自发的和蛋白质介导的转移/交换的胆固醇（Ch）之间的膜已被广泛研究，相对较少的是已知的易位的Ch氧化产物，特别是氢过氧化物物种（ChOOHs），它可以作为细胞毒性促氧化剂。本研究的主要目的是检查和比较几种生物相关的ChOOH异构体的膜间转移特性，包括单线态氧衍生的5α-OOH、6α-OOH和6β-OOH以及自由基衍生的7α-OOH和7β-OOH。这些物质是在[14 C] Ch标记的供体膜[红细胞血影或单层DMPC/Ch（1.0：0.8 mol/mol）脂质体]中通过染料敏化的光过氧化反应产生的。在37 °C下，通过薄层色谱法和磷光成像放射性检测（HPTLC-PI）或液相色谱法和汞阴极电化学检测[HPLC-EC（Hg）]监测自发转移至非氧化受体膜（分别为DMPC脂质体或血影）。前者允许测量总（未分离）ChOOH沿着母体Ch，而后者允许测量单个ChOOH。Ghost膜中约4%的Ch被过氧化，主要产生5α-OOH，总ChOOH和Ch以表观一级方式转移到脂质体中，ChOOH的速率常数是其65倍。像Ch解吸，ChOOH解吸从供体膜被发现是速率限制，和速率与大小成反比时，使用脂质体供体。对于单个ChOOH，速率常数大小（7α/7β-OOH > 5α-OOH > 6 α-OOH> 6β-OOH）与反相HPLC保留时间呈负相关，表明更快的转移反映了更大的亲水性。脂质体携带的ChOOOH对COH-BR 1细胞表现出相同的毒性顺序，COH-BR 1细胞缺乏对这些过氧化物解毒的能力。从这些发现中可以很容易地看出，通过ChOOH和其他脂质氢过氧化物的易位传播氧化性细胞损伤的前景是显而易见的。
Whereas spontaneous and protein-mediated transfer/exchange of cholesterol (Ch) between membranes has been widely studied, relatively little is known about the translocation of Ch oxidation products, particularly hydroperoxide species (ChOOHs), which can act as cytotoxic prooxidants. A major aim of the present study was to examine and compare the intermembrane transfer characteristics of several biologically relevant ChOOH isomers, including singlet oxygen-derived 5α-OOH, 6α-OOH, and 6β-OOH and free radical-derived 7α-OOH and 7β-OOH. These species were generated in [14C]Ch-labeled donor membranes [erythrocyte ghosts or unilamellar DMPC/Ch (1.0:0.8 mol/mol) liposomes] by means of dye-sensitized photoperoxidation. Spontaneous transfer to nonoxidized acceptor membranes (DMPC liposomes or ghosts, respectively) at 37 °C was monitored by thin-layer chromatography with phosphorimaging radiodetection (HPTLC-PI) or liquid chromatography with mercury cathode electrochemical detection [HPLC-EC(Hg)]. The former allowed measurement of total (unresolved) ChOOH along with parent Ch, whereas the latter allowed measurement of individual ChOOHs. Ghost membranes in which ∼4% of the Ch had been peroxidized, giving mainly 5α-OOH, transferred total ChOOH and Ch to liposomes in apparent first-order fashion, the rate constant for ChOOH being ∼65 times greater. Like Ch desorption, ChOOH desorption from donor membranes was found to be rate limiting, and rate varied inversely with size when liposomal donors were used. For individual ChOOHs, rate constant magnitude (7α/7β-OOH > 5α-OOH > 6α-OOH > 6β-OOH) correlated inversely with reverse-phase HPLC retention time, suggesting that faster transfer reflects greater hydrophilicity. Liposome-borne ChOOHs exhibited the same order of toxicity toward COH-BR1 cells, which are deficient in ability to detoxify these peroxides. The prospect of disseminating oxidative cell injury via translocation of ChOOHs and other lipid hydroperoxides is readily apparent from these findings.