汽油尾气致氧化应激作用的实验研究
发布时间：2018-10-13 08:11
【摘要】： 汽油尾气已成为许多城市室外空气污染的主要来源，且其成分中含有许多已确证或可疑的人类致癌物，因此，进一步明确汽油尾气暴露与人体健康的关系就变得十分重要。现的有研究资料显示，汽油尾气暴露与许多肺及肺外疾病的发生相关。虽然，1989年国际癌症研究中心(IARC)将汽油尾气归为2B类人类致癌物，但迄今为止，对于人们最为关注的汽油尾气暴露与肺癌的发生的关系，不论流行病学、动物实验还是毒性机制研究方面的资料，都无法确认和达成共识。为了了解汽油尾气的毒作用机制及氧化损伤在其毒性机制中的作用，进一步揭示汽油尾气暴露与癌症发生之间的关系，从而为汽油尾气暴露与癌症发生之间的关系提供实验依据。本研究以汽油尾气的冷凝物、颗粒物和半挥发性有机物(Semivolatile organic compound，SVOC)三相成分的有机提取物为处理因素，探讨了汽油尾气对体外培养细胞和气管滴注染毒后的动物多器官组织细胞的氧化损伤作用。 体外试验中以人肺腺癌上皮细胞株A549细胞为研究对象，通过MTT试验，2′，7′-二氯双氢荧光素双乙酸酯(DCFH-DA)活性氧测定实验、抗氧化试验、微核试验以及彗星试验对汽油尾气影响细胞存活率、活性氧生成和脂类的氧化损伤、抗氧化酶活力以及造成细胞DNA的链断裂和染色体损伤的情况进行了观察。结果显示，汽油尾气能降低细胞存活率，诱导细胞内活性氧的生成和堆积，造成脂质的氧化损伤和抗氧化酶活力降低，并能导致DNA单链断裂的增加和染色体的损伤，表明汽油尾气具有明显的细胞毒性和遗传毒性，而氧化应激则可能是其毒作用机制之一。此外，通过还原性谷胱甘肽(Glutathion，GSH)对汽油尾气毒效应的干预实验表明GSH可以降低汽油尾气的毒性效应。表现为GSH预处理组的细胞存活率、活性氧含量和脂质过氧化程度、抗氧化酶活力的下降以及DNA单链的断裂情况明显低于未干预组。该结果从另一个侧面进一步提示氧化损伤在汽油尾气的毒效应过程中具有重要的作用。 体内实验依据美国环保局(EPA)关于新燃料毒性评价的规定，将汽油尾气的三相有机提取物通过气管滴注对SD大鼠进行染毒，然后对肺脏、脑、肝脏和睾丸等器官进行多指标的实验观察。结果表明，汽油尾气可造成大鼠肺组织中脂类、蛋白质和核酸的氧化损伤以及抗氧化酶活力的下降，此外病理组织切片观察到肺组织的炎性损伤明显加重。通过免疫组化观察肺组织中DNA氧化损伤的关键性修复酶——8-羟基鸟嘌呤DNA糖苷酶-1(OGG1)含量的变化，结果显示汽油尾气的暴露可导致肺组织8-羟基鸟嘌呤DNA糖苷酶-1的含量明显增加：电镜观察肺组织细胞器细微结构的变化，发现肺组织各型细胞中的线粒体损伤明显，板层小体结构发生病理改变，并有部分细胞核固缩。同样，汽油尾气暴露导致脑、肝脏和睾丸等器官组织中的脂质过氧化产物丙二醛(MDA)和蛋白质氧化损伤产物羰基蛋白(CP)的含量增加、抗氧化酶超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GPx)的活力下降(脑组织除外)、组织细胞DNA的链断裂增加。动物实验的结果表明，汽油尾气经呼吸道暴露后，肺是其主要的靶器官，但脑、肝、睾丸等其他脏器也会被累及，表明汽油尾气暴露后的损伤是多器官的，而氧化应激可能是其重要的毒作用机制之一。 体外体内实验的结果均表明，，汽油尾气暴露后可导致多组织多器官的脂类、核酸和蛋白质等生物大分子的的氧化损伤，并能导致DNA损伤修复酶OGG1的量发生改变，这些损伤和变化也为癌症的发生创造了可能。
[Abstract]:Gasoline tail gas has become the main source of outdoor air pollution in many cities, and its components contain many confirmed or suspected human carcinogens, so it becomes very important to further clarify the relationship between gasoline tail gas exposure and human health. The present study shows that the exposure of gasoline tail gas is related to the occurrence of many lung and extrapulmonary diseases. Although the 1989 International Cancer Research Centre (IARC) classifies gasoline off-gas as a 2B human carcinogen, Neither was able to confirm and reach consensus. In order to understand the toxic action mechanism of gasoline tail gas and the role of oxidative damage in its toxic mechanism, the relationship between the exposure of gasoline tail gas and the occurrence of cancer is further revealed, and the experimental basis is provided for the relationship between the exposure of gasoline tail gas and the occurrence of cancer. The effects of gasoline tail gas on the oxidative damage of multi-organ tissue cells in vitro cultured cells and trachea were investigated by using organic extracts of condensate, particulate matter and semi-volatile organic compound (SVOC) of gasoline tail gas as processing factors. In vitro test, human lung adenocarcinoma epithelial cell line A549 cell was used as the research object, and the tail gas shadow of gasoline was determined by MTT assay, 2-day, 7-day-dichloro-bishydrogen fluorescein diacetate (DCFH-DA) active oxygen measurement experiment, antioxidant test, micronucleus test and comet assay. In response to cell survival, the oxidative damage of active oxygen species and lipids, the activity of antioxidant enzymes, The results show that the tail gas of gasoline can reduce the cell survival rate, induce the generation and accumulation of active oxygen species in the cells, cause the oxidative damage of lipid and decrease the activity of antioxidant enzymes, and can lead to the increase of DNA single-strand breaks. The damage of the chromosome shows that the tail gas of gasoline has obvious cytotoxicity and genotoxicity, and oxidative stress may be its poison. In addition, the effect of reducing glutathione (GSH) on the tail gas toxicity of gasoline indicates that GSH can lower the tail of gasoline The results showed that the cell survival rate, reactive oxygen content and lipid peroxidation, the decrease of antioxidant enzyme activity and the fragmentation of DNA single strand in GSH pretreatment group were observed. lower than the non-intervention group. The result further suggests that oxidative damage occurs in the poison effect of gasoline tail gas from the other side In vivo, the three-phase organic extracts of gasoline tail gas were exposed to SD rats by air-gas instillation, and then lung, brain, liver and testis were treated according to EPA's regulations on new fuel toxicity evaluation. The results showed that the oxidative damage of lipid, protein and nucleic acid in lung tissue of rats and the decrease of antioxidant enzyme activity could be caused by the tail gas of gasoline, and the pathological tissue sections were observed. The results show that the exposure of gasoline tail gas can lead to 8-hydroxyproline DN in lung tissues. The changes of cell organelle structure of lung tissue were observed by electron microscope, and the damage of mitochondria in various cells of lung tissue was found to be obvious. In the same vein, the exposure of gasoline tail gas leads to lipid peroxidation products (MDA) and protein oxidative damage products in organs such as brain, liver and testis. The content of zinc finger protein (CP) increased, the activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx) decreased (except brain tissue). The results of animal experiments indicate that the lung is its main target organ after exposure to respiratory tract, but other organs, such as brain, liver and testis, can also be involved, indicating that the injury after exposure of gasoline tail gas is multi-organ, and oxidative stress can The results of in vitro experiments show that the oxidative damage of biological macromolecules such as lipids, nucleic acids and proteins of multiple organs can be caused after exposure of gasoline tail gas, and can resulting in a change in the amount of DNA damage repair enzyme OGG1
【学位授予单位】：四川大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R363
本文编号：2267894
[Abstract]:Gasoline tail gas has become the main source of outdoor air pollution in many cities, and its components contain many confirmed or suspected human carcinogens, so it becomes very important to further clarify the relationship between gasoline tail gas exposure and human health. The present study shows that the exposure of gasoline tail gas is related to the occurrence of many lung and extrapulmonary diseases. Although the 1989 International Cancer Research Centre (IARC) classifies gasoline off-gas as a 2B human carcinogen, Neither was able to confirm and reach consensus. In order to understand the toxic action mechanism of gasoline tail gas and the role of oxidative damage in its toxic mechanism, the relationship between the exposure of gasoline tail gas and the occurrence of cancer is further revealed, and the experimental basis is provided for the relationship between the exposure of gasoline tail gas and the occurrence of cancer. The effects of gasoline tail gas on the oxidative damage of multi-organ tissue cells in vitro cultured cells and trachea were investigated by using organic extracts of condensate, particulate matter and semi-volatile organic compound (SVOC) of gasoline tail gas as processing factors. In vitro test, human lung adenocarcinoma epithelial cell line A549 cell was used as the research object, and the tail gas shadow of gasoline was determined by MTT assay, 2-day, 7-day-dichloro-bishydrogen fluorescein diacetate (DCFH-DA) active oxygen measurement experiment, antioxidant test, micronucleus test and comet assay. In response to cell survival, the oxidative damage of active oxygen species and lipids, the activity of antioxidant enzymes, The results show that the tail gas of gasoline can reduce the cell survival rate, induce the generation and accumulation of active oxygen species in the cells, cause the oxidative damage of lipid and decrease the activity of antioxidant enzymes, and can lead to the increase of DNA single-strand breaks. The damage of the chromosome shows that the tail gas of gasoline has obvious cytotoxicity and genotoxicity, and oxidative stress may be its poison. In addition, the effect of reducing glutathione (GSH) on the tail gas toxicity of gasoline indicates that GSH can lower the tail of gasoline The results showed that the cell survival rate, reactive oxygen content and lipid peroxidation, the decrease of antioxidant enzyme activity and the fragmentation of DNA single strand in GSH pretreatment group were observed. lower than the non-intervention group. The result further suggests that oxidative damage occurs in the poison effect of gasoline tail gas from the other side In vivo, the three-phase organic extracts of gasoline tail gas were exposed to SD rats by air-gas instillation, and then lung, brain, liver and testis were treated according to EPA's regulations on new fuel toxicity evaluation. The results showed that the oxidative damage of lipid, protein and nucleic acid in lung tissue of rats and the decrease of antioxidant enzyme activity could be caused by the tail gas of gasoline, and the pathological tissue sections were observed. The results show that the exposure of gasoline tail gas can lead to 8-hydroxyproline DN in lung tissues. The changes of cell organelle structure of lung tissue were observed by electron microscope, and the damage of mitochondria in various cells of lung tissue was found to be obvious. In the same vein, the exposure of gasoline tail gas leads to lipid peroxidation products (MDA) and protein oxidative damage products in organs such as brain, liver and testis. The content of zinc finger protein (CP) increased, the activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx) decreased (except brain tissue). The results of animal experiments indicate that the lung is its main target organ after exposure to respiratory tract, but other organs, such as brain, liver and testis, can also be involved, indicating that the injury after exposure of gasoline tail gas is multi-organ, and oxidative stress can The results of in vitro experiments show that the oxidative damage of biological macromolecules such as lipids, nucleic acids and proteins of multiple organs can be caused after exposure of gasoline tail gas, and can resulting in a change in the amount of DNA damage repair enzyme OGG1
【学位授予单位】：四川大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R363
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本文编号：2267894
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