柴油机尾气暴露所致DNA甲基化改变及其与微核组学指标间的关联性研究
发布时间：2018-09-18 10:03
【摘要】：近些年来,空气污染物对机体造成的健康危害越来越为公众所关注。在众多空气污染物中,颗粒物与不良健康效应的关系最为密切,而且先前的研究提示细颗粒物相较于粗颗粒物可能对人体的健康危害更大。柴油发动机尾气(Diesel engine exhaust, DEE)是空气中细颗粒物的主要来源之一。鉴于柴油发动机在市区的广泛应用,大量的城市居民已经成为DEE最主要的暴露人群。此外,许多职业人群如矿工、卡车司机也会在工作环境中暴露于DEE。流行病学研究表明长期暴露于DEE与肺癌发生风险增高间存在关联。遗传损伤被认为是癌症引发阶段的关键分子事件。胞质阻滞微核试验(Cytokinesis-block micronucleus assay, CBMN)常被用来评价暴露于遗传毒物后机体内的遗传损伤程度,它可以测定外周血淋巴细胞微核(Micronucleus, MN)、核质桥(Nucleoplasmic bridge, NPB)、核芽(Nuclear bud, NBUD)率等指标。虽然已有人群研究报道过DEE暴露可以引起MN率发生改变,但是由于存在混合暴露的问题,DEE暴露与微核组学指标改变间的关系尚不完全清楚。此外,DEE诱导遗传毒效应通路中和遗传毒效应发生后机体应答通路中的调控模式仍然不清楚。表观遗传是指不涉及DNA序列并且可以遗传的基因表达的改变。越来越多的研究表明表观遗传修饰改变作为一种新的调控基因表达的机制在环境暴露-毒效应-机体应答这一连续模式中发挥着重要的作用。一方面,特异的表观遗传修饰改变可以反映机体接触环境暴露的程度。另一方面,表观遗传修饰改变能够参与或者响应环境暴露引起的遗传毒效应。因此,表观遗传修饰不仅可能作为介导环境暴露与癌症风险间关联的生物学机制,还可能作为环境暴露和遗传损伤的生物标志物。DNA甲基化是迄今为止研究最为深入的一种表观遗传修饰,它指的是CpG位点的胞嘧啶末端共价结合甲基形成5-甲基胞嘧啶。一般情况下,DNA甲基化的发生与基因沉默有关。DNA损伤应答(DNA damage response, DDR)相关基因和重复序列的甲基化状态对于维持基因组的稳定性十分重要。有研究在肺癌患者中发现DDR相关基因和重复序列发生了异常甲基化改变。近年来,关于DEE暴露和DNA甲基化改变的人群研究逐渐增多,已有研究在哮喘患者中发现短期暴露于DEE可以引起蛋白激酶和核因子kB通路中基因的甲基化水平发生改变。然而仍然有许多关键的科学问题亟待解决,比如长期暴露于DEE能否引起DDR相关基因和重复序列的甲基化水平发生改变,这些甲基化水平改变反过来是否与DEE暴露导致的遗传毒效应间存在关联。为了解决以上科学问题,本研究选取了117名暴露于单纯DEE的柴油机试车工人和112名非DEE暴露工人,以DDR相关基因和重复序列甲基化为切入点,结合微核组学指标,探讨DEE暴露、DNA甲基化、遗传毒效应三者间的关联。一.DEE的成分解析与表征分别用扫描电迁移率粒径谱仪和碳分析仪测定了DEE颗粒相的粒径分布及元素碳和有机碳的含量,用气相色谱质谱仪测定了DEE颗粒相和工人个体暴露样本中多环芳烃(Polycyclic aromatic hydrocarbons, PAHs)的含量,用便携式气相色谱质谱仪测定了DEE气相有机物的浓度,用被动采气管测定了DEE气相二氧化氮和二氧化硫的浓度,用高效液相色谱质谱测定了工人尿中6种羟基多环芳烃(Mono-hydroxylated PAHs, OH-PAHs)的浓度。结果表明84.3%的DEE颗粒为100nm以下。元素碳和有机碳分别占细颗粒物的百分比平均为28.6±6.3%和36.2±6.5%,有机碳／元素碳平均为1.31±0.31。致癌性PAHs和非致癌性PAHs分别占总PAHs的83.3%和16.7%。DEE气相有机物主要包括链烃及其衍生物、环烷烃衍生物、苯及其衍生物。元素碳与有机碳呈显著正相关(r=0.630,P=0.002),与二氧化氮和二氧化硫呈临界显著正相关(r=0.370,P=0.090和r=0.385,P=0.077)。DEE暴露工人对萘、芴、菲、芘四种PAHs的个体暴露水平及尿中总羟基萘、二羟基芴、总羟基菲、一羟基芘的浓度均高于非DEE暴露工人,差异具有统计学意义(P0.001)。二.DEE暴露与DNA甲基化间的关联性研究用焦磷酸测序的方法测定了工人外周血淋巴细胞三个DDR相关基因(p16、RASSF1A、MGMT)和LINE-1重复序列的甲基化水平。结果表明DEE暴露工人较非DEE暴露工人p16、RASSFIA、MGMT基因甲基化水平降低,差异具有统计学意义(P0.001),并未发现LINE-1甲基化水平在两组间具有显著差异。随着尿总OH-PAHs浓度的增高,p16、RASSF1A、MGMT基因甲基化水平呈显著降低的趋势(Ptrend分别为0.018、0.006、0.001)。非吸烟工人中,在校正了混杂因素的影响后,尿总OH-PAHs浓度每上升一个四分位间距,p16、RASSF1A、 MGMT基因经logit转换后的甲基化水平平均改变分别为-0.13、-0.18、-0.19。DEE暴露工人中,DEE暴露时长与p16、RASSF1A基因甲基化水平呈显著正相关(r=0.293,P=0.001和r=0.409,P0.001)。与溶剂对照组相比,DEE提取物处理组原代淋巴细胞p16、RASSF1A、MGMT基因甲基化水平改变方向大体与人群研究结果一致。三.DEE暴露与微核组学指标间的关联性研究用CBMN法测定了工人外周血淋巴细胞MN、NPB、NBUD率并求得三者之和计算出基因组不稳定指数。用基因组不稳定指数加上本课题组先前报道的凋亡率和坏死率计算出微核组学指数。结果表明DEE暴露工人较非DEE暴露工人MN、NPB、NBUD率、基因组不稳定指数、微核组学指数升高,差异具有统计学意义(P0.001)。随着尿总OH-PAHs浓度的增高,MN、NPB、NBUD率、基因组不稳定指数、微核组学指数呈显著升高的趋势(Ptrend0.001)。非吸烟工人中,在校正了混杂因素的影响后,尿总OH-PAHs浓度每上升一个四分位间距,MN、NPB、NBUD率、基因组不稳定指数、微核组学指数平均变化百分比分别为93.48%、467.46%、160.39%、128.64%、121.22%。DEE暴露工人中,在校正了混杂因素的影响后,尿总OH-PAHs浓度每上升一个四分位间距,MN和微核组学指数平均变化百分比分别为38.13%和27.63%。四.DEE暴露、DNA甲基化、DNA损伤指标、微核组学指标的相互关系用因子分析、相关分析、中介效应分析、交互作用分析等方法探讨了本研究所测指标及本课题组先前报道的DNA链损伤指标[外周血淋巴细胞Olive尾距(Olive tail moment, OTM)]和DNA氧化损伤指标[尿1,N6-乙烯基脱氧腺苷(1,N6-ethenodeoxyadenosine, εdA)和3,N4-乙烯基脱氧胞苷(3,N4-ethenodeoxycytidine, εdC)]间的相互关系。结果表明p16、RASSF1A、 MGMT基因甲基化与εdA间呈显著负相关(r=-0.187,P=-0.008；r=-0.177,P=0.013;r=-0.251,P0.001),与OTM间呈显著负相关(r=-0.300,P0.001；r=-0.305,P0.001；r=-0.311,P0.001),与基因组不稳定指数间呈显著负相关(r=-0.183,P=0.007; r=-0.212, F=0.002; r=-0.244, P0.001)。p16和RASSFIA基因甲基化与核分裂指数间呈显著正相关(r=0.220,P=0.001;r=0.225, P=0.001)。OTM和基因组不稳定指数分别对核分裂指数效应的24.9%和6.1%可以被p16甲基化所介导,26.1%和6.6%可以被RASSFIA甲基化所介导。非吸烟工人中,自然对数转换后的尿总OH-PAHs每上升一个单位,LINE-1甲基化水平85.6%和≥85.6%的工人基因组不稳定指数平均变化百分比分别为64.38%和45.50%(Pinteraction=0.016)。基因组不稳定指数与εdA和OTM间呈显著正相关(r=0.204,P=0.004;r=0.353,P0.001)。用尿总OH-PAHs和εdA的第一和第四分位数作为分界分别判定DEE暴露水平高低和遗传损伤大小时,MGMT甲基化对应的曲线下面积分别为0.758和0.711。用OTM的第一分位数和第四分位数作为分界判定遗传损伤大小时,p16和RASSFIA甲基化对应的曲线下面积分别为0.705和0.724。结论1)DEE颗粒的粒径大小大多属于超细颗粒物的范畴,其上吸附的PAHs主要是致癌性PAHs。DEE气相有机组分主要包括链烃、苯及它们的衍生物。元素碳与其它DEE主要组分间呈显著或临界显著正相关。DEE暴露工人较非DEE暴露工人尿OH-PAHs浓度显著增高。随着DEE暴露工人对PAHs个体暴露水平的增高,其尿中相应PAHs代谢产物的浓度也随之增高。2)DEE暴露可以引起外周血淋巴细胞p16、RASS1F、MGMT基因发生低甲基化改变。DEE暴露与p16、RASSFIA、MGMT基因甲基化水平间呈显著负关联。DEE暴露时长与p16、RASSFIA基因甲基化水平间呈显著正相关。MGMT基因甲基化水平可以作为DEE暴露水平的生物标志物。3)DEE暴露可以导致外周血淋巴细胞MN、NPB、NBUD率、基因组不稳定指数、微核组学指数增高。DEE暴露与MN、NPB.NBUD率、基因组不稳定指数、微核组学指数呈显著正关联。DEE暴露水平与MN率和微核组学指数间呈显著正关联。4) εdA、OTM、基因组不稳定指数与p16、RASSF1A、MGMT基因甲基化水平间呈显著负相关。εdA和OTM与基因组不稳定指数间呈显著正相关。p16和RASSFIA以及MGMT基因甲基化水平可以分别作为DEE暴露所致OTM、εdA水平的生物标志物。5)p16和RASSF1A基因甲基化可以介导OTM和基因组不稳定指数与核分裂指数间的关联,LINE-1甲基化可以修饰DEE暴露与基因组不稳定指数间的关联。
[Abstract]:In recent years, more and more attention has been paid to the health hazards caused by air pollutants. Among many air pollutants, particulate matter is the most closely related to adverse health effects. Previous studies have suggested that fine particulate matter may be more harmful to human health than coarse particulate matter. Exhaust (DEE) is one of the main sources of fine particulate matter in the air. In view of the wide use of diesel engines in urban areas, a large number of urban residents have become the main exposures to DEE. In addition, many occupational groups, such as miners and truck drivers, are also exposed to DEE in the working environment. Epidemiological studies have shown that long-term exposure to DEE and lung. Cytokinesis-block micronucleus assay (CBMN) is often used to assess the degree of genetic damage in the body after exposure to genetic toxicants. It can be used to measure micronucleus in peripheral blood lymphocytes. MN, NPB, NBUD and other indicators. Although there have been population studies reported that DEE exposure can cause changes in MN rates, the relationship between DEE exposure and changes in micronucleomic indicators is not completely clear due to the problem of mixed exposure. Epigenetics refers to changes in gene expression that do not involve DNA sequences and can be inherited. A growing number of studies have shown that epigenetic modifications, as a new mechanism for regulating gene expression, are involved in environmental exposure-toxicity-response. Continuous patterns play an important role. On the one hand, specific epigenetic modifications can reflect the degree of exposure to the environment. On the other hand, epigenetic modifications can participate in or respond to the genetic toxicity caused by environmental exposure. DNA methylation is one of the most well-studied epigenetic modifications to date. It refers to the formation of 5-methylcytosine by covalently binding methyl at the cytosine end of the CpG site to form 5-methylcytosine. The methylation of DNA damage response (DDR) related genes and repetitive sequences is important for maintaining genomic stability. Abnormal methylation of DDR related genes and repetitive sequences has been found in lung cancer patients. Population studies on DEE exposure and DNA methylation have been conducted in recent years. Increasing numbers of studies have found that short-term exposure to DEE can cause changes in the methylation levels of genes in protein kinase and nuclear factor kB pathways in asthmatic patients. However, there are still many key scientific issues to be resolved, such as whether long-term exposure to DEE can cause changes in the methylation levels of DDR-related genes and repetitive sequences. In order to solve the above scientific problems, 117 diesel engine test workers and 112 non-DEE exposed workers were selected in this study. DDR-related genes and repetitive sequence methylation were used as the breakthrough point, and micronucleomic finger was combined with DNA methylation. The relationship among DEE exposure, DNA methylation and genetic toxicity was investigated. 1. DEE decomposition and characterization were performed by scanning electromobility particle size spectrometer and carbon analyzer respectively to determine the particle size distribution of DEE and the contents of elemental carbon and organic carbon, and to determine DEE particle phase and individual exposure samples by gas chromatography-mass spectrometry. The contents of polycyclic aromatic hydrocarbons (PAHs) were determined by portable gas chromatography-mass spectrometry (GC-MS). The concentrations of DEE vapor-phase organic compounds (VOCs) were determined by passive recovery pipe. Six kinds of hydroxyl polycyclic aromatic hydrocarbons (PAHs) in workers'urine were determined by high performance liquid chromatography-mass spectrometry (HPLC-MS). OH-PAHs concentration. The results showed that 84.3% of DEE particles were below 100 nm. The average percentage of elemental carbon and organic carbon in fine particles was 28.6 (+ 6.3%) and 36.2 (+ 6.5%) respectively. The average percentage of organic carbon/elemental carbon was 1.31 (+ 0.31%). Carcinogenic PAHs and non-carcinogenic PAHs accounted for 83.3% and 16.7% of total PAHs, respectively. Derivatives, cycloalkane derivatives, benzene and its derivatives. Elemental carbon was positively correlated with organic carbon (r = 0.630, P = 0.002), and positively correlated with nitrogen dioxide and sulfur dioxide (r = 0.370, P = 0.090 and R = 0.385, P = 0.077). DEE exposed workers to naphthalene, fluorene, phenanthrene, pyrene, total hydroxy naphthalene, dihydroxy fluorene and total urine The methylation levels of three DDR-related genes (p16, RASSF1A, MGMT) and LINE-1 repeats in peripheral blood lymphocytes of workers were determined by pyrophosphatic acid sequencing. The methylation levels of p16, RASSFIA and MGMT genes in EE exposed workers were significantly lower than those in non-DEE exposed workers (P 0.001). There was no significant difference in the methylation levels of LINE-1 between the two groups. With the increase of urinary total OH-PAHs concentration, the methylation levels of p16, RASSF1A and MGMT genes were significantly decreased (P tree was 0.018, 0.001, respectively). Among non-smoking workers, the methylation levels of p16, RASSF1A, MGMT genes were - 0.13, - 0.18, - 0.19. The methylation levels of p16, RASSF1A genes were significantly positively correlated with the exposure time of DEE in non-smoking workers after adjusting for the influence of confounding factors. Guan (r = 0.293, P = 0.001 and R = 0.409, P 0.001). Compared with the solvent control group, the methylation levels of p16, RASSF1A and MGMT genes in primary lymphocytes treated with DEE extract were generally consistent with the results of the population study. 3. Correlation between DEE exposure and micronucleomic indices. MN, NPB, NBUD in peripheral blood lymphocytes of workers were measured by CBMN method. The results showed that the MN, NPB, NBUD, genomic instability index and micronucleomic index of DEE exposed workers were higher than those of non-DEE exposed workers. Significance (P 0.001). With the increase of urinary total OH-PAHs concentration, MN, NPB, NBUD rate, genomic instability index, micronucleomic index showed a significant increase trend (Ptrend 0.001). Among non-smokers, after adjusting for the influence of confounding factors, the urinary total OH-PAHs concentration increased by a quartile interval, MN, NPB, NBUD rate, genomic instability index, and micronucleomic index. Among DEE exposed workers, the average change percentages of MN and MN and MN were 38.13% and 27.63% for each quartile increase in urinary total OH-PAHs concentration after adjusting for confounding factors. The correlations of micronucleomic indices were analyzed by factor analysis, correlation analysis, mediating effect analysis and interaction analysis. The indexes measured in this study and the DNA strand damage indices [Olive tail moment (OTM) of peripheral blood lymphocytes] and DNA oxidative damage indices [urine 1, N6-vinyl deoxygland] previously reported by our group were discussed. The results showed that p16, RASSF1A, MGMT gene methylation was significantly negatively correlated with epdA (r = - 0.187, P = - 0.187, P = - 0.008; r = - 0.177, P = 0.013; r = - 0.251, P 0.251, P 0.001), and was negatively correlwith OTM (r = - 300, P = - 300, P = - 300, P = - 0.300, P = - 300, P = - 0.001, P = - 0.001; r = - 0.30, r = - 0.30; r = - 0.30; r = - 0.30; 0.0.0.001, P = - 0.001, P = - 0.001; 0.001, P = - 0.001; 5. P16 and RASSFIA gene methylation were positively correlated with mitotic index (r = 0.220, P = 0.001; r = 0.183, P = 0.007; r = - 0.212, F = 0.002; r = - 0.244, P 0.001). OTM and genomic instability index were positively correlated with mitotic index (r = 0.220, P = 0.001; r = 0.225, P = 0.001), respectively. In non-smoking workers, the average percentage of changes in the genomic instability index of LINE-1 methylation levels of 85.6% and (>85.6%) were 64.38% and 45.50% respectively (Pinteraction = 0.016) for each unit increase in urinary total OH-PAHs after natural logarithmic conversion. Genome instability index was positively correlated with epsilon dA and OTM (r = 0.204, P = 0.004; r = 0.353, P 0.001). When the first and fourth quantiles of urinary total OH-PAHs and epsilon dA were used as the dividing line to determine the level of DEE exposure and the extent of genetic damage, the sub-curves of MGMT methylation were 0.758 and 0.711, respectively. Conclusion 1) The size of DEE particles belongs to the category of ultrafine particles, and the PAHs adsorbed on them are mainly carcinogenic PAHs. DEE vapor phase organic components mainly include chain hydrocarbons, benzene and their derivatives. The concentration of urinary OH-PAHs in DEE exposed workers was significantly higher than that in non-DEE exposed workers. With the increase of individual exposure to PAHs, the concentration of urinary PAHs metabolites in DEE exposed workers also increased. 2) DEE exposure could induce p16 and RAS in peripheral blood lymphocytes. There was a significant negative correlation between DEE exposure and methylation levels of p16, RASSFIA, and MGMT genes. DEE exposure was positively correlated with MN, NPB. NBUD, genomic instability index, genomic instability index and micronucleomic index. There was a significant positive correlation between DEE exposure and MN and micronucleomic index. 4) There was a significant positive correlation between epsilon dA, OTM, genomic instability index and methylation levels of p16, RASSF1A and MGMT genes. The methylation levels of p16, RASSFIA and MGMT genes could be used as biomarkers of OTM, and the methylation levels of epsilon dA and RASSF1A genes could mediate the association between OTM and genomic instability index and mitotic index, respectively. Methylation can modify the association between DEE exposure and genomic instability index.
【学位授予单位】：中国疾病预防控制中心
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R114
本文编号：2247561
[Abstract]:In recent years, more and more attention has been paid to the health hazards caused by air pollutants. Among many air pollutants, particulate matter is the most closely related to adverse health effects. Previous studies have suggested that fine particulate matter may be more harmful to human health than coarse particulate matter. Exhaust (DEE) is one of the main sources of fine particulate matter in the air. In view of the wide use of diesel engines in urban areas, a large number of urban residents have become the main exposures to DEE. In addition, many occupational groups, such as miners and truck drivers, are also exposed to DEE in the working environment. Epidemiological studies have shown that long-term exposure to DEE and lung. Cytokinesis-block micronucleus assay (CBMN) is often used to assess the degree of genetic damage in the body after exposure to genetic toxicants. It can be used to measure micronucleus in peripheral blood lymphocytes. MN, NPB, NBUD and other indicators. Although there have been population studies reported that DEE exposure can cause changes in MN rates, the relationship between DEE exposure and changes in micronucleomic indicators is not completely clear due to the problem of mixed exposure. Epigenetics refers to changes in gene expression that do not involve DNA sequences and can be inherited. A growing number of studies have shown that epigenetic modifications, as a new mechanism for regulating gene expression, are involved in environmental exposure-toxicity-response. Continuous patterns play an important role. On the one hand, specific epigenetic modifications can reflect the degree of exposure to the environment. On the other hand, epigenetic modifications can participate in or respond to the genetic toxicity caused by environmental exposure. DNA methylation is one of the most well-studied epigenetic modifications to date. It refers to the formation of 5-methylcytosine by covalently binding methyl at the cytosine end of the CpG site to form 5-methylcytosine. The methylation of DNA damage response (DDR) related genes and repetitive sequences is important for maintaining genomic stability. Abnormal methylation of DDR related genes and repetitive sequences has been found in lung cancer patients. Population studies on DEE exposure and DNA methylation have been conducted in recent years. Increasing numbers of studies have found that short-term exposure to DEE can cause changes in the methylation levels of genes in protein kinase and nuclear factor kB pathways in asthmatic patients. However, there are still many key scientific issues to be resolved, such as whether long-term exposure to DEE can cause changes in the methylation levels of DDR-related genes and repetitive sequences. In order to solve the above scientific problems, 117 diesel engine test workers and 112 non-DEE exposed workers were selected in this study. DDR-related genes and repetitive sequence methylation were used as the breakthrough point, and micronucleomic finger was combined with DNA methylation. The relationship among DEE exposure, DNA methylation and genetic toxicity was investigated. 1. DEE decomposition and characterization were performed by scanning electromobility particle size spectrometer and carbon analyzer respectively to determine the particle size distribution of DEE and the contents of elemental carbon and organic carbon, and to determine DEE particle phase and individual exposure samples by gas chromatography-mass spectrometry. The contents of polycyclic aromatic hydrocarbons (PAHs) were determined by portable gas chromatography-mass spectrometry (GC-MS). The concentrations of DEE vapor-phase organic compounds (VOCs) were determined by passive recovery pipe. Six kinds of hydroxyl polycyclic aromatic hydrocarbons (PAHs) in workers'urine were determined by high performance liquid chromatography-mass spectrometry (HPLC-MS). OH-PAHs concentration. The results showed that 84.3% of DEE particles were below 100 nm. The average percentage of elemental carbon and organic carbon in fine particles was 28.6 (+ 6.3%) and 36.2 (+ 6.5%) respectively. The average percentage of organic carbon/elemental carbon was 1.31 (+ 0.31%). Carcinogenic PAHs and non-carcinogenic PAHs accounted for 83.3% and 16.7% of total PAHs, respectively. Derivatives, cycloalkane derivatives, benzene and its derivatives. Elemental carbon was positively correlated with organic carbon (r = 0.630, P = 0.002), and positively correlated with nitrogen dioxide and sulfur dioxide (r = 0.370, P = 0.090 and R = 0.385, P = 0.077). DEE exposed workers to naphthalene, fluorene, phenanthrene, pyrene, total hydroxy naphthalene, dihydroxy fluorene and total urine The methylation levels of three DDR-related genes (p16, RASSF1A, MGMT) and LINE-1 repeats in peripheral blood lymphocytes of workers were determined by pyrophosphatic acid sequencing. The methylation levels of p16, RASSFIA and MGMT genes in EE exposed workers were significantly lower than those in non-DEE exposed workers (P 0.001). There was no significant difference in the methylation levels of LINE-1 between the two groups. With the increase of urinary total OH-PAHs concentration, the methylation levels of p16, RASSF1A and MGMT genes were significantly decreased (P tree was 0.018, 0.001, respectively). Among non-smoking workers, the methylation levels of p16, RASSF1A, MGMT genes were - 0.13, - 0.18, - 0.19. The methylation levels of p16, RASSF1A genes were significantly positively correlated with the exposure time of DEE in non-smoking workers after adjusting for the influence of confounding factors. Guan (r = 0.293, P = 0.001 and R = 0.409, P 0.001). Compared with the solvent control group, the methylation levels of p16, RASSF1A and MGMT genes in primary lymphocytes treated with DEE extract were generally consistent with the results of the population study. 3. Correlation between DEE exposure and micronucleomic indices. MN, NPB, NBUD in peripheral blood lymphocytes of workers were measured by CBMN method. The results showed that the MN, NPB, NBUD, genomic instability index and micronucleomic index of DEE exposed workers were higher than those of non-DEE exposed workers. Significance (P 0.001). With the increase of urinary total OH-PAHs concentration, MN, NPB, NBUD rate, genomic instability index, micronucleomic index showed a significant increase trend (Ptrend 0.001). Among non-smokers, after adjusting for the influence of confounding factors, the urinary total OH-PAHs concentration increased by a quartile interval, MN, NPB, NBUD rate, genomic instability index, and micronucleomic index. Among DEE exposed workers, the average change percentages of MN and MN and MN were 38.13% and 27.63% for each quartile increase in urinary total OH-PAHs concentration after adjusting for confounding factors. The correlations of micronucleomic indices were analyzed by factor analysis, correlation analysis, mediating effect analysis and interaction analysis. The indexes measured in this study and the DNA strand damage indices [Olive tail moment (OTM) of peripheral blood lymphocytes] and DNA oxidative damage indices [urine 1, N6-vinyl deoxygland] previously reported by our group were discussed. The results showed that p16, RASSF1A, MGMT gene methylation was significantly negatively correlated with epdA (r = - 0.187, P = - 0.187, P = - 0.008; r = - 0.177, P = 0.013; r = - 0.251, P 0.251, P 0.001), and was negatively correlwith OTM (r = - 300, P = - 300, P = - 300, P = - 0.300, P = - 300, P = - 0.001, P = - 0.001; r = - 0.30, r = - 0.30; r = - 0.30; r = - 0.30; 0.0.0.001, P = - 0.001, P = - 0.001; 0.001, P = - 0.001; 5. P16 and RASSFIA gene methylation were positively correlated with mitotic index (r = 0.220, P = 0.001; r = 0.183, P = 0.007; r = - 0.212, F = 0.002; r = - 0.244, P 0.001). OTM and genomic instability index were positively correlated with mitotic index (r = 0.220, P = 0.001; r = 0.225, P = 0.001), respectively. In non-smoking workers, the average percentage of changes in the genomic instability index of LINE-1 methylation levels of 85.6% and (>85.6%) were 64.38% and 45.50% respectively (Pinteraction = 0.016) for each unit increase in urinary total OH-PAHs after natural logarithmic conversion. Genome instability index was positively correlated with epsilon dA and OTM (r = 0.204, P = 0.004; r = 0.353, P 0.001). When the first and fourth quantiles of urinary total OH-PAHs and epsilon dA were used as the dividing line to determine the level of DEE exposure and the extent of genetic damage, the sub-curves of MGMT methylation were 0.758 and 0.711, respectively. Conclusion 1) The size of DEE particles belongs to the category of ultrafine particles, and the PAHs adsorbed on them are mainly carcinogenic PAHs. DEE vapor phase organic components mainly include chain hydrocarbons, benzene and their derivatives. The concentration of urinary OH-PAHs in DEE exposed workers was significantly higher than that in non-DEE exposed workers. With the increase of individual exposure to PAHs, the concentration of urinary PAHs metabolites in DEE exposed workers also increased. 2) DEE exposure could induce p16 and RAS in peripheral blood lymphocytes. There was a significant negative correlation between DEE exposure and methylation levels of p16, RASSFIA, and MGMT genes. DEE exposure was positively correlated with MN, NPB. NBUD, genomic instability index, genomic instability index and micronucleomic index. There was a significant positive correlation between DEE exposure and MN and micronucleomic index. 4) There was a significant positive correlation between epsilon dA, OTM, genomic instability index and methylation levels of p16, RASSF1A and MGMT genes. The methylation levels of p16, RASSFIA and MGMT genes could be used as biomarkers of OTM, and the methylation levels of epsilon dA and RASSF1A genes could mediate the association between OTM and genomic instability index and mitotic index, respectively. Methylation can modify the association between DEE exposure and genomic instability index.
【学位授予单位】：中国疾病预防控制中心
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R114
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