【摘要】： “吸煙有害健康”，近乎家喻戶曉，人人皆知。香煙煙霧中含有多種有害的化學(xué)物質(zhì)，已被確認(rèn)是多種疾病的主要誘因之一，并且在流行病學(xué)、細(xì)胞遺傳學(xué)和分子遺傳等方面得到了廣泛證實(shí)。本文以健康人紅細(xì)胞為模型，研究香煙煙霧提取物(Cigarette smoke extract，CSE)導(dǎo)致的細(xì)胞滲透脆性、細(xì)胞膜蛋白、脂質(zhì)過(guò)氧化水平和谷胱甘肽含量改變，以及對(duì)膜磷脂和脂肪酸的作用，以評(píng)估香 煙煙霧對(duì)細(xì)胞膜的氧化損害作用，同時(shí)對(duì)茶多酚類物質(zhì)對(duì)這些損害的抑制作用進(jìn)行分析，為香煙煙霧對(duì)機(jī)體的損害機(jī)制及天然多酚化合物的防護(hù)作用提供分子水平上的依據(jù)。 紅細(xì)胞的正常生理功能與細(xì)胞膜結(jié)構(gòu)的完整性有關(guān)，其中膜蛋白的組成和結(jié)構(gòu)變化均會(huì)對(duì)膜的通透性、變形性、物質(zhì)運(yùn)輸、細(xì)胞識(shí)別以及酶的活性產(chǎn)生影響。在本實(shí)驗(yàn)中，紅細(xì)胞在經(jīng)CSE處理后，其滲透脆性減弱，溶血率和MDA含量升高，GSH含量降低，，這說(shuō)明CSE對(duì)紅細(xì)胞的抗氧化體系具有損害作用，這種損害與香煙煙霧劑量增高呈正相關(guān)。進(jìn)一步的研究發(fā)現(xiàn)，一定劑量的表沒(méi)食子兒茶素沒(méi)食子酸酯(Epigallocatechin gallate，EGCG)對(duì)CSE誘導(dǎo)的紅細(xì)胞氧化損傷有一定的防護(hù)作用。 對(duì)生物膜的作用是香煙煙霧產(chǎn)生機(jī)體損害的首要環(huán)節(jié)。紅細(xì)胞在經(jīng)CSE處理后，其膜蛋白的SDS聚丙稀酰胺凝膠電泳圖譜發(fā)生明顯改變。與空白對(duì)照比較，高濃度的CSE可以導(dǎo)致一些膜蛋白聚集，生成高分子量聚集體，這些高分子量聚集體主要來(lái)源于帶2.6，帶3，帶4.3和帶5蛋白。這些膜蛋白的聚集與CSE誘導(dǎo)的蛋白氧化和二硫鍵重組，進(jìn)而引起膜蛋白肽鏈交聯(lián)密切相關(guān)。CSE處理的膜蛋白經(jīng)DTT還原后，電泳條帶中無(wú)高分子量聚集體出現(xiàn)。EGCG對(duì)由CSE導(dǎo)致的蛋白條帶的消減有一定的抑制作用，但對(duì)由CSE引起的蛋白聚集無(wú)明顯影響。 香煙煙霧對(duì)生物膜損害的另一方面是對(duì)膜脂質(zhì)的作用。成熟紅細(xì)胞的脂類主要包含在膜內(nèi)，脂類約占膜干重的40％，其中50％為磷脂。在磷脂中，磷脂酰膽堿(PC)和磷脂酰乙醇胺(PE)為主要組分。膜磷脂的組成及含量變化可反映膜的結(jié)構(gòu)和功能的改變。本研究采用高效液相色譜(HPLC)和氣相色譜-質(zhì)譜(GC-MS)分析CSE作用后的膜脂質(zhì)成分的變化，結(jié)果顯示，紅細(xì)胞膜中最主要的兩種磷脂PE和PC在CSE的作用下含量降低。在206nm處檢測(cè)色譜分離的膜磷脂時(shí)，其信號(hào)主要與磷脂分子中不飽和脂肪酸雙鍵的含量有關(guān)，所以，PE和PC的分析值下降反映了膜磷脂分子中不飽和脂肪酸的含量減少。另外，CSE中的一些組分導(dǎo)致磷脂分子脂肪酸鏈水解也可能是細(xì)胞膜中磷脂含量減少的原因之一。 CSE可以導(dǎo)致膜脂質(zhì)中不飽和脂肪酸含量降低。GC-MS結(jié)果顯示，50mL／LCSE可使紅細(xì)胞膜中的的亞油酸和花生四烯酸消失，100mL／L CSE可使亞油酸、花生四烯酸以及二十四碳一烯酸消失，300mL／L CSE處理紅細(xì)胞后，膜脂質(zhì)中僅有飽和脂酸存在。50μg／mL EGCG則對(duì)此具有一定的防護(hù)作用。 綜上所述，香煙煙霧的水溶物對(duì)細(xì)胞的損害主要源于其中的氧化性物質(zhì)和自由基。紅細(xì)胞在CSE的作用下，抗氧化能力降低，細(xì)胞膜的性質(zhì)發(fā)生改變。這些變化將影響整個(gè)機(jī)體的抗氧化能力。EGCG及其它類型的植物多酚化合物是目前臨床上應(yīng)用最為廣泛的植物藥提取物。該類提取物已被證實(shí)對(duì)人的身體機(jī)能具有廣泛的有益作用，并對(duì)一些慢性疾病有輔助治療作用。近年來(lái)，植物來(lái)源的藥物倍受青睞。本研究采用人紅細(xì)胞為實(shí)驗(yàn)?zāi)Ｐ�，發(fā)現(xiàn)EGCG可以部分消除CSE誘導(dǎo)的細(xì)胞氧化損害，為茶多酚類物質(zhì)作為抗氧化藥物和在食品和保健品方面的應(yīng)用提供了理論上的依據(jù)。
[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.
【學(xué)位授予單位】：陜西師范大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2007
【分類號(hào)】：R285.5;R363

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