竹红菌甲素对红细胞膜蛋白及膜磷脂的光敏损伤

本文以红细胞膜为材料,通过测量磷脂过氧化荧光产物的产生,磷脂组分的损伤以及膜蛋白二级结构的破坏,内源荧光的下降和蛋白SDS-凝胶电泳分析,探讨了甲素光敏作用中蛋白和磷脂的损伤。结果表明:甲素存在时红细胞膜样品照光,使磷脂产生过氧化荧光产物和丙二醛,磷脂组分受到破坏(其中PE与PS较敏感),在膜状态的磷脂比提取的磷脂脂质体中的损伤来得剧烈。同时,膜蛋白二级结构遭到破坏,内源荧光下降。在膜状态中spectrin的损伤比提取出的spectrin的损伤来得严重。据此,我们认为,在甲素光敏作用产生的蛋白和磷脂的损伤过程中,蛋白与磷脂间存在着相互作用和相互影响,使损伤加剧。     

竹红菌甲素对红细胞膜和几种磷脂脂质体膜的流动...

In this paper, the photodamage of Hypocrellin A to the fluidity of human erythrocyte membranes and some kinds of membranes of phospholipid liposomes was investigated by measuring the changes in fluorescence polarization of the membranes. The results showed that the photosensitization effect of HA caused the decrease of membrane fluidity of the phospholipid (DPPC, DPPC/DPPE, phospholipid of erythrocyte membranes) liposomes. The DPPC and DPPC/DPPE liposomes were more sensitive to the damage than the phospholipid liposomes of erythrocyte membranes. To human erythrocyte membranes, the photodamage effect of HA caused its fluidity first increased and then, with the increment of illumination time, decreased. To spectrin-depleted and trypsin-treated erythrocyte membranes, this kind of change in fluidity was inhibited. All of the results indicated that phospholipids and proteins play different roles in the photodamage of HA to the fluidity of membranes. Membrane proteins, especially spectrin, were the key factor involved in the changes of the fluidity.     

竹红菌甲素对红细胞膜和几种磷脂脂质体膜的流动性的光敏损伤

本文以荧光探针为手段,通过测量膜偏振度的变化,探讨了竹红菌甲素光敏作用对红细胞膜和几种磷脂脂质体膜的流动性的损伤。结果表明,甲素光敏作用使不同种类的磷脂(DPPC,DPPC/DPPE,红细胞膜磷脂)脂质体的流动性增加,其对光敏作用的敏感程度为红细胞膜磷脂脂质体显著小于DPPC/DPPE脂质体及DPPC脂质体。对红细胞膜来说,甲素光敏作用使其流动性呈现先降低而后增加的现象。去除膜上的spectrin以及用胰蛋白酶处理可使这种流动性变化的幅度受到抑制。据此,我们认为,膜磷脂,膜蛋白对甲素光敏作用中膜流动性的变化有着不同的影响,膜蛋白,特别是spectrin,是其中极重要的因素。     

竹红菌甲素对红细胞膜上几种酶光敏失活作用的研究

Hypocrellin A (HA)-sensitized photoinactivation of enzymes in human erythrocyte membrane, including AchE, GPDH, Na(+)-K+ ATPase, Ca2(+)-Mg2+ ATPase were studied in this paper. The sensitivity of these four enzymes inactivated by HA and light are as following order: Ca2(+)-Mg2+ ATPase greater than Na(+)-K+ ATPase greater than GPDH greater than AchE. The relationship among ATPase inactivation, sulfhydryl photoinactivation and lipid peroxidation was also investigated. Results show that SH group photooxidation probably is one of the major reasons of enzyme inactivation whereas lipid peroxidation has little effect. The isolated GPDH was less sensitive than that membrane-bound, GSH, NAD acted protectively on GPDH and ATPase respectively. The evidence of electrophoresis and protein intrinsic fluorescence showed that protein structure did not change significantly even though most activity had lost in case of GPDH.     

竹红菌甲素对红细胞膜内脂双层的微扰

In this paper, using human erythrocyte membrane, the effect of Hypocrellin A on the lipid bilayer of the membrane was studied by measuring the change of the fluidity of the membrane, the energy transfer of the fluorescent probes, the shift of the fluorescent emission peaks, and the split of band-a of Hypocrellin A. The results showed that in the presence of HA, the fluidity of erythrocyte membrane was increased, the fluorescence intensity of the probes was decreased, and the fluorescence peaks shifted blue. These phenomena took place more seriously with the increment of HA concentration. Meanwhile, the band-a of HA excitation spectra was splitted. It was suggested from all of the results that HA could significantly perturb the lipid bilayer of erythrocyte membrane, there were interactions existing between the Hypocrellin A and the membrane. The HA was mainly located in the middle range of the membrane lipid bilayer when in high concentration (mainly to the 12-16 positions of the long chain fatty acid).     

竹红菌甲素对红细胞膜内脂双层的微扰

本文以荧光探针为手段,以人红细胞膜为材料,测量了膜偏振度的改变,荧光探针能量转移,荧光峰的蓝移和甲素激发峰的分裂。结果表明在有竹红菌甲素存在时,红细胞膜偏振度增加,探针荧光强度减小,荧光峰蓝移。甲素浓度增加时,上述现象更加明显,即它们之间有正的相关关系。同时,甲素激发光谱的a带发生分裂。据此,我们认为甲素对红细胞膜内脂双层产生明显微扰,甲素与红细胞膜间存在着相互作用。在甲素浓度较大时,它主要是渗入到红细胞膜脂双层的深层部位(膜脂肪酸链的12—16位)。     

Effect of asymmetric distribution of phospholipids in ghost membrane from rat blood on peroxidation induced by ferrous ion

This study used rat erythrocyte ghost membrane to investigate the effect of aminophospholipid distribution in biological membranes on oxidative susceptibility. Aminophospholipids, lipid peroxidation, and carbonyl compounds were quantified; plasma membrane structure was examined using atomic force microscopy (AFM) and SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Inside-out vesicles (IOVs) had significantly more aminophospholipids and greater lipid peroxidation than right-side-out vesicles (ROVs). Spectrin bands in IOVs disappeared obviously than in ROVs as shown in SDS-PAGE. In both systems vesicle protein size increased significantly with oxidation. Proteins aggregated much more in IOVs than ROVs at 48 h. These observations suggest that IOVs were more susceptible to ferrous ion-induced peroxidation than ROVs and that asymmetric phospholipid distribution affects biomembranes' oxidative susceptibility.     

Effects of added dietary taurine on erythrocyte lipids and oxidative stress in rabbits fed a high cholesterol diet

Lipid peroxidation leads to damage of polyunsaturated fatty acids of membrane phospholipids. The contribution of oxidative stress to hypercholesterolemia-induced hemolytic anemia and the effects of addition of taurine on erythrocyte lipid composition, oxidative stress, and hematological data were studied in rabbits fed on a high cholesterol (HC) diet (1%, w/w) for 2 months. The effects of taurine on erythrocyte hemolysis and H2O2-induced lipid peroxidation were investigated in normal rabbit erythrocytes in vitro. The HC diet resulted in increases in plasma lipids and lipid peroxide levels as well as increases in cholesterol levels and the cholesterol:phospholipid ratio in the erythrocytes. This diet caused a hemolytic anemia, but lipid peroxide levels remained unchanged in the erythrocytes of the rabbits. Taurine (2.5%, w/w) added to the food has an ameliorating effect on plasma lipids and lipid peroxide levels in rabbits fed on a HC diet. This treatment also caused decreases in elevated erythrocyte cholesterol levels and cholesterol:phospholipid ratio due to the HC diet, but it did not prevent the hemolytic anemia and did not change erythrocyte lipid peroxide levels. In addition, in an in vitro study, taurine did not protect erythrocytes against H2O2-induced hemolysis or lipid peroxidation. These results show that the HC diet causes hemolytic anemia without any changes in erythrocyte lipid peroxidation, and taurine treatment was not effective against hemolytic anemia caused by the HC diet.     

血红蛋白对人红细胞膜流动性的影响

本文报道了pH7.5时血红蛋白和红细胞膜的结合效应.在10—45℃温度范围内观察到血红蛋白对膜脂质流动性的限制作用.看来这种限制作用不是脂质过氧化所致,而是血红蛋白和红细胞膜直接作用的结果.对流动性大的膜,血红蛋白的效应也随之增大.高铁血红蛋白及红细胞膜去胆固醇皆能修饰血红蛋白和膜的相互作用.     

Lipid peroxidation and phospholipase A2 activity in liposomes composed of unsaturated phospholipids: a structural basis for enzyme activation

The effect of lipid peroxidation on membrane structure and phospholipase A2 activity was studied using liposomes composed of bovine liver phosphatidylcholine (PC) and phosphatidylethanolamine (PE). The phospholipids were mixed at set ratios and sonicated to yield small unilamellar vesicles. The liposome preparations were subjected to lipid peroxidation as induced by cumene hydroperoxide and hematin. Under these conditions, a sharp increase in lipid peroxidation was noted over a 30 min incubation period and was accompanied by loss of polyunsaturated fatty acids (PUFA). Liposomes enriched in PE were most extensively peroxidized with a preferred oxidation of this phospholipid. The extent of PC oxidation was also greater in liposomes containing the largest proportions of PE. Analysis of liposome anisotropy, via steady-state fluorescence polarization of diphenylhexatriene indicated that progressive increases in either PE content or the level of lipid peroxidation increased the apparent microviscosity of the vesicles. Moreover, lipid peroxidation increased anisotropy more effectively than variations in the ratios of PE vs. PC. Thus, peroxidation of 5-10% of the phospholipids produced the same anisotropy increase as a 20% increase in the ratio of PE vs. PC. Analysis of vesicle turbidity suggested that fusion was also more readily achieved through lipid peroxidation. When liposomes were incubated with 0.4 U/ml of snake venom phospholipase A2, a direct correlation was found between the degree of lipid peroxidation and the extent of phospholipid hydrolysis. The more unsaturated phospholipid, PE, was most extensively hydrolyzed following peroxidation. Increasing the proportion of PE also resulted in more extensive phospholipid hydrolysis. These findings indicate that lipid peroxidation produces a general increase in membrane viscosity which is associated with vesicle instability and enhanced phospholipase A2 attack. A structural basis for membrane phospholipase A2 activation as a consequence of lipid peroxidation is discussed in light of these findings.