香烟烟雾提取物诱导人红细胞膜损伤及茶多酚的保护作用

发布时间：2018-10-20 08:59

【摘要】： “吸烟有害健康”，近乎家喻户晓，人人皆知。香烟烟雾中含有多种有害的化学物质，已被确认是多种疾病的主要诱因之一，并且在流行病学、细胞遗传学和分子遗传等方面得到了广泛证实。本文以健康人红细胞为模型，研究香烟烟雾提取物(Cigarette smoke extract，CSE)导致的细胞渗透脆性、细胞膜蛋白、脂质过氧化水平和谷胱甘肽含量改变，以及对膜磷脂和脂肪酸的作用，以评估香烟烟雾对细胞膜的氧化损害作用，同时对茶多酚类物质对这些损害的抑制作用进行分析，为香烟烟雾对机体的损害机制及天然多酚化合物的防护作用提供分子水平上的依据。红细胞的正常生理功能与细胞膜结构的完整性有关，其中膜蛋白的组成和结构变化均会对膜的通透性、变形性、物质运输、细胞识别以及酶的活性产生影响。在本实验中，红细胞在经CSE处理后，其渗透脆性减弱，溶血率和MDA含量升高，GSH含量降低，，这说明CSE对红细胞的抗氧化体系具有损害作用，这种损害与香烟烟雾剂量增高呈正相关。进一步的研究发现，一定剂量的表没食子儿茶素没食子酸酯(Epigallocatechin gallate，EGCG)对CSE诱导的红细胞氧化损伤有一定的防护作用。对生物膜的作用是香烟烟雾产生机体损害的首要环节。红细胞在经CSE处理后，其膜蛋白的SDS聚丙稀酰胺凝胶电泳图谱发生明显改变。与空白对照比较，高浓度的CSE可以导致一些膜蛋白聚集，生成高分子量聚集体，这些高分子量聚集体主要来源于带2.6，带3，带4.3和带5蛋白。这些膜蛋白的聚集与CSE诱导的蛋白氧化和二硫键重组，进而引起膜蛋白肽链交联密切相关。CSE处理的膜蛋白经DTT还原后，电泳条带中无高分子量聚集体出现。EGCG对由CSE导致的蛋白条带的消减有一定的抑制作用，但对由CSE引起的蛋白聚集无明显影响。香烟烟雾对生物膜损害的另一方面是对膜脂质的作用。成熟红细胞的脂类主要包含在膜内，脂类约占膜干重的40％，其中50％为磷脂。在磷脂中，磷脂酰胆碱(PC)和磷脂酰乙醇胺(PE)为主要组分。膜磷脂的组成及含量变化可反映膜的结构和功能的改变。本研究采用高效液相色谱(HPLC)和气相色谱-质谱(GC-MS)分析CSE作用后的膜脂质成分的变化，结果显示，红细胞膜中最主要的两种磷脂PE和PC在CSE的作用下含量降低。在206nm处检测色谱分离的膜磷脂时，其信号主要与磷脂分子中不饱和脂肪酸双键的含量有关，所以，PE和PC的分析值下降反映了膜磷脂分子中不饱和脂肪酸的含量减少。另外，CSE中的一些组分导致磷脂分子脂肪酸链水解也可能是细胞膜中磷脂含量减少的原因之一。 CSE可以导致膜脂质中不饱和脂肪酸含量降低。GC-MS结果显示，50mL／LCSE可使红细胞膜中的的亚油酸和花生四烯酸消失，100mL／L CSE可使亚油酸、花生四烯酸以及二十四碳一烯酸消失，300mL／L CSE处理红细胞后，膜脂质中仅有饱和脂酸存在。50μg／mL EGCG则对此具有一定的防护作用。综上所述，香烟烟雾的水溶物对细胞的损害主要源于其中的氧化性物质和自由基。红细胞在CSE的作用下，抗氧化能力降低，细胞膜的性质发生改变。这些变化将影响整个机体的抗氧化能力。EGCG及其它类型的植物多酚化合物是目前临床上应用最为广泛的植物药提取物。该类提取物已被证实对人的身体机能具有广泛的有益作用，并对一些慢性疾病有辅助治疗作用。近年来，植物来源的药物倍受青睐。本研究采用人红细胞为实验模型，发现EGCG可以部分消除CSE诱导的细胞氧化损害，为茶多酚类物质作为抗氧化药物和在食品和保健品方面的应用提供了理论上的依据。
[Abstract]:"Smoking is harmful to health" It's almost a household name. It's well known. Cigarette smoke contains a variety of harmful chemicals and has been identified as one of the main causes of a variety of diseases, and has been widely confirmed in epidemiology, cytogenetics and molecular genetics. In this paper, the cell osmotic fragility, cell membrane protein, lipid peroxidation level and glutathione content of cigarette smoke extract (CSE) induced by cigarette smoke extract (CSE), as well as the effect of membrane phospholipids and fatty acids, were studied. to evaluate the aroma The effects of smoke smoke on the oxidative damage of membrane were analyzed, and the inhibition effect of tea polyphenols on these lesions was analyzed. The damage mechanism of cigarette smoke to organism and the protective effect of natural polyphenol compounds were provided. The normal physiological function of red blood cells is related to the integrity of membrane structure, and the composition and structural changes of membrane proteins are related to the permeability, deformability and substance transport of membrane. In this experiment, after CSE treatment, the osmotic fragility of red blood cells decreased, the hemolysis rate and MDA content increased, GSH content decreased, which indicated that CSE had damage to the anti-oxidation system of red blood cells. The damage was positively correlated with the increase of cigarette smoke dose. Red blood cell oxidative damage has certain protective effect. The effect of biofilm on biofilm is the primary link to the damage of cigarette smoke. The red blood cells are treated by CSE, the membrane Compared with the blank control, the high concentration of CSE can cause some membrane proteins to accumulate and produce high molecular weight aggregates, and these high molecular weight aggregates The body mainly comes from band 2. 6, band 3, band 4. 3 and band 5 protein. The aggregation of these membrane proteins and the protein oxygen induced by CSE The recombination of disulfide bonds and disulfide bonds, in turn, causes the cross-linking of membrane proteins to be closely related to the membrane proteins of CSE treatment. There was no high molecular weight aggregates in the electrophoretic bands after the reduction of the protein. The results showed that the reduction of the protein bands caused by CSE was certain. Inhibition, but no significant effect on protein aggregation due to CSE. Cigarette smoke, on the other hand, plays an important role in membrane lipid peroxidation. The mature red blood cells The lipid is mainly contained in the membrane, the lipid accounts for about 40% of the dry weight of the membrane, In phospholipids, phospholipids are choline (PC) and phospholipids and ethanolamine (PE The composition and content of membrane phospholipids could reflect the changes of membrane structure and function. The results showed that the changes of membrane lipid component after CSE were analyzed by high performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS). and at 206nm, the signal is mainly related to the content of unsaturated fatty acid double bonds in the phospholipid molecules, therefore, The decrease of the analysis value of PE and PC reflects the decrease of unsaturated fatty acid content in membrane phospholipid molecules. In addition, some components in CSE The hydrolysis of the fatty acid chain of the phospholipid may also be a phospholipid in the cell membrane The results of GC-MS show that 50mL/ LCSE can cause the disappearance of linoleic acid and arachidonic acid in erythrocyte membrane, 100mL/ L CSE. Linoleic acid, arachidonic acid and 24-carbon monoenoic acid disappear and 300mL/ L CSE deals with red fine After the cell, only the saturated acid is present in the membrane lipid. Protective effects. To sum up, the water-soluble substance of cigarette smoke damage the cells mainly derived from oxidizing substances and free radicals therein. Under the action of CSE, the anti-oxidation ability of cells decreased and the properties of cell membranes changed. The anti-oxidation ability of the whole organism is the most widely used in clinic. An extract of the extract. The extract has been proved to be Human body function has a wide range of beneficial effects and has an auxiliary therapeutic effect on some chronic diseases. In recent years, the drugs of plant origin have been favored. Human red blood cells are used as experimental models, which can partially eliminate CS.
【学位授予单位】：陕西师范大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R285.5;R363

【相似文献】