【作者】 孙迪；

【导师】 范雪云； 马骏；

【作者基本信息】 河北联合大学 ， 公共卫生与预防医学， 2011， 硕士

【摘要】 目的尘肺是在生产过程中长期吸入粉尘发生的以肺组织纤维化为主的疾病。长期吸入不同种类的粉尘可导致不同类型的尘肺或肺部疾患。尘肺是我国职业病中对工人健康危害非常严重的一类疾病,据统计,2009年新发各类职业病18128例,其中尘肺病新增14495例,在14495例尘肺病新病例中,煤工尘肺和矽肺占91.89%。尘肺的病因明确,发病过程较长,多在接触粉尘期间被确诊,脱离粉尘作业后仍可患尘肺。大量的流行病学调查发现,工作环境及接尘时间相同的个体并不都患尘肺,且患病的不同个体,其严重程度常存在差异。提示,尘肺的发生发展除取决于环境因素外,很可能与个体的易感性密切相关。粉尘刺激导致肺组织损伤,炎症因子释放,刺激网状纤维和胶原纤维增生,加之免疫复合物沉积是形成尘肺病变的基础。目前研究较多并认为与尘肺发病关系较大的细胞因子主要有:纤维粘连蛋白(FN)、肿瘤坏死因子(TNF-α、β)、转化生长因子(TGF)、白细胞介素(IL-1和IL-6)和血小板源性生长因子(PDGF)等。IL-8是一种主要由中性粒细胞产生的强有力的中性粒细胞趋化因子和活化因子,在炎症、感染、损伤、组织缺氧、酸中毒等病理状态下产生,对嗜碱性粒细胞和T细胞也有一定的趋化作用,可趋化中性粒细胞聚集,激活中性粒细胞释放超氧化物酶、溶酶体酶,产生炎症反应,而调控其表达水平的基因位点存在多态性。目前,尘肺患者的发病是否与白介素-8的基因多态性有关,国内外并未见报道。这次研究采用整群病例对照研究方法,在严格控制发病年龄、民族、接尘性质、累积接尘时间等因素的条件下,探讨白介素-8基因多态性与尘肺易感性的关系,为筛选和鉴定易感人群,防治尘肺病提供理论依据。对象与方法以唐山开滦矿业(集团)有限责任公司煤矿2568名粉尘作业工人和某金矿及耐火材料厂1265名接尘工人为研究对象,在健康检查的基础上,所有接受体检的接尘工人均由专业医务人员拍摄后前位胸片,由有资质认证的尘肺诊断专家组按照我国《尘肺病诊断标准》(GBZ270-2009)盲法诊断,以确诊的尘肺患者为病例组,共213例,均为汉族男性,其中包括101例壹期矽肺患者和112例壹期煤工尘肺患者。从受检的接尘工人中选择与病例性别、发病年龄相似,同工作场所的汉族男性无尘肺的251名接尘工人为对照。在知情同意情况下,取体检者静脉血1.5ml,加入EDTA抗凝,冰盒保存,待提取模板DNA。采用盐析法提取DNA,采用聚合酶链反应-限制性片段长度多态性(polymerase chains reaction-restriction fragment length polymorphism, PCR-RFLP)和PCR方法检测白细胞介素-8 Met31Arg、781C/T、-251A/T、RA+860位点基因型及等位基因的频率,应用χ2检验及OR值进行比较分析,得出与尘肺的相关性。结果1一般情况该研究共选择壹期、汉族、男性尘肺病人213例,平均发病年龄54.9±9.4岁,累积接尘工龄29.3±6.1年,对照组251例,平均发病年龄为54.8±6.3岁,累积接尘工龄29.5±4.9年。其中矽肺病例101例,发病年龄平均为59.1±8.9岁,累积接尘工龄平均为28.0±5.1年。对照组121例均为汉族男性,发病年龄平均为57.3±8.1岁,累积接尘工龄平均为28.1±5.2年。煤工尘肺病例112例,发病年龄平均为51.1±8.1岁,累积接尘工龄平均为29.8±6.7年。对照组130例均为汉族男性,发病年龄平均为52.4±1.9岁,累积接尘工龄平均为30.6±4.7年。矽肺病例组和煤工尘肺病例组与相应的对照比较,发病年龄、累积接尘工龄、吸烟差异均无统计学意义(P>0.05),病例组与对照组具有良好的可比性。2白介素-8 Met31Arg位点基因型及等位基因分布情况白介素-8 Met31Arg位点在尘肺病例组GG、GT和TT基因型分布频率分别为0%、14.08%和85.92%,对照组则分别为0.40%、3.59%和96.37%,经χ2检验,差异有统计学意义(P<0.05)。G等位基因在病例组和对照组的分布频率分别为7.04%和2.19%,T等位基因在病例组和对照组的分布频率分别为92.96%和97.81%,差异有统计学意义(P<0.05)。分层分析发现,矽肺组为GG、GT和TT基因型分布频率分别为0%、12.87%和87.13%;对照组则分别为0.82%、2.48%和96.69%,经χ2检验,差异有统计学意义(P<0.05)。G等位基因在病例组和对照组的分布频率分别为6.44%和2.07%,T等位基因在病例组和对照组的分布频率分别为93.56%和97.93%,差异有统计学意义(P<0.05)。煤工尘肺组为GG、GT和TT基因型分布频率分别为0%、15.18%和84.82%;对照组则分别为0%、4.62%和95.38%,经χ2检验,差异有统计学意义(P<0.05)。G等位基因在病例组和对照组的分布频率分别为7.59%和2.31%,T等位基因在病例组和对照组的分布频率分别为92.41%和97.69%,差异有统计学意义(P<0.05)。3白介素-8 -251A/T位点基因型及等位基因分布情况白介素-8 -251A/T位点在尘肺病例组TT、AT和AA基因型分布频率分别为38.97%、48.83%和12.20%;对照组则分别为17.93%、17.93%和64.14%,经χ2检验,差异有统计学意义(P<0.05)。T等位基因在病例组和对照组的分布频率分别为63.38%和53.78%,A等位基因在病例组和对照组的分布频率分别为36.62%和46.22%,差异有统计学意义(P<0.05)。分层分析表明,矽肺组组为TT、AT和AA基因型分布频率分别为40.59%、49.50%和9.90%;对照组则分别为38.84%、35.54%和25.62%,经χ2检验,差异有统计学意义(P<0.05)。T等位基因在病例组和对照组的分布频率分别为65.35%和56.61%,A等位基因在病例组和对照组的分布频率分别为34.61%和43.39%,差异没有统计学意义(P>0.05)。煤工尘肺组为TT、AT和AA基因型分布频率分别为37.50%、48.21%和14.29%;对照组则分别为33.08%、36.15%和30.77%,经χ2检验,差异有统计学意义(P<0.05)。T等位基因在病例组和对照组的分布频率分别为61.61%和51.15%,A等位基因在病例组和对照组的分布频率分别为38.39%和48.85%,差异有统计学意义(P<0.05)。4白介素-8 781C/T位点基因型及等位基因分布情况白介素-8 781C/T位点在尘肺病例组CC、CT和TT基因型分布频率分别为37.09%、41.31%和21.60%;对照组则分别为43.03%、43.03%和13.94%,经χ2检验,差异有统计学意义(P<0.05)。C等位基因在病例组和对照组的分布频率分别为57.75%和64.54%,T等位基因在病例组和对照组的分布频率分别为42.25%和35.46%,差异有统计学意义(P<0.05)。分层分析中,矽肺组为CC、CT和TT基因型分布频率分别为38.61%、40.59%和20.79%;对照组分别为46.28%、40.50%和13.22%,经χ2检验,差异无统计学意义(P>0.05)。C等位基因在病例组和对照组的分布频率分别为58.91%和66.53%,T等位基因在病例组和对照组的分布频率分别为41.09%和33.47%,差异无统计学意义(P>0.05)。煤工尘肺组为CC、CT和TT基因型分布频率分别为:35.71%、41.96%和22.33%;对照组则分别为40.00%、45.38%和14.62%,经χ2检验,差异没有统计学意义(P>0.05)。C等位基因在病例组和对照组的分布频率分别为56.70%和62.69%,T等位基因在病例组和对照组的分布频率分别为43.30%和37.31%,差异无统计学意义(P>0.05)。5白介素-8 RA+860位点基因型及等位基因分布情况白介素-8 RA+860位点在尘肺病例组GG、GC和CC基因型分布频率在病例组分别为74.24%、23.47%和3.29%;对照组则分别为73.31%、20.32%和6.37%,经χ2检验,差异没有统计学意义(P>0.05)。G等位基因在病例组和对照组的分布频率分别为84.98%和83.47%,C等位基因在病例组和对照组的分布频率分别为15.02%和16.53%,差异没有统计学意义(P>0.05)。分层分析发现,白介素-8 RA+860位点GG、GC和CC基因型分布频率在矽肺病例组分别为75.25%、21.78%和2.97%;对照组则分别为80.17%、14.88%和4.96%,经χ2检验,差异没有统计学意义(P>0.05)。G等位基因在病例组和对照组的分布频率分别为86.14%和87.60%,C等位基因在病例组和对照组的分布频率分别为13.86%和12.40%,差异没有统计学意义(P>0.05)。白介素-8 RA+860位点GG、GC和CC基因型分布频率在煤工尘肺病例组分别为:71.43%、25.00%和3.57%;对照组则分别为66.92%、25.38%和7.70%,经χ2检验,差异没有统计学意义(P>0.05)。G等位基因在病例组和对照组的分布频率分别为83.93%和79.62%,C等位基因在病例组和对照组的分布频率分别为16.07%和20.38%,差异没有统计学意义(P>0.05)。结论1研究发现白介素-8(Met31Arg)位点基因多态性与矽肺及煤工尘肺的易感性均有关联,携带IL-8(Met31Arg)GT基因型的接尘工人患尘肺的危险性增加。2研究发现白介素-8(-251A/T)位点的基因多态性与矽肺及煤工尘肺的易感性均有关联;携带IL-8(-251)AA基因型接尘工人患尘肺危险性降低。3本次研究未发现RA(+860)位点的基因多态性与尘肺易感性有关。4研究发现IL-8(781C/T)TT基因型的接尘工人患尘肺的危险性增加,但分层分析后差异无统计学意义。更多还原

【Abstract】 Objectives Pneumoconiosis is a chronic lung disease due to the inhalation of various forms of dust particles.Different types of dust can resut in different types of pneumoconiosis.Pneumoconiosis is the most common and serious occupational disease in China.According to the end of 2009 year`s data, 18128 new occupational disease had been diagnosed, 14495 of these were pneumoconiosis, coal worker’s pneumoconiosis and silicosis account 91.89%.Pneumoconiosis have clear cause and long pathogenesis,and can be diagnosed during exposure to dust. Workers can suffer from pneumoconiosis after departed dust. According to a great deal epidemiological studies of pneumoconiosis, only a few persons suffered from it even they had the same situation of exposure. They had the different ponderance in the group of patients. This fact indicated that the morbidity of pneumoconiosis had the relationship not only with environmental factor but also with individual factor. Dust stimulation can lead lung injury, release inflammatory, stimulate reticular fibers and collagen’s hyperplasia.The deposition of immune complex deposition is the basis of changes of pneumoconiosis.Now study more and repute cytokines that have close relationship with the incidence of pneumoconiosis have fibronectin(FN),factor-α,β(TNF-α,β),transforming growth factor (TGF),Interleukin-1(IL-1),Interleukin-6(IL-6) and platelet-derived growth factor(PDGF). IL-8 is a powerful neutrophil chemokine chemotactic factor and activating factor that mainly produced from neutrophils and produced in pathological conditions that including in inflammation, infection, injury, hypoxia, acidosis. IL-8 have a certain degree of chemotaxis on basophils and T cells and it can chemotaxis neutrophils gathered, activate neutrophils release super oxide enzyme and lysosomal enzymes,and produce inflammation,then control the polymorphisms of Interleukin-8. Now we don’t find the studies of relationship between the genetic polymorphisms of IL-8 and pneumoconiosis.After strictly matching age of first diagnosis,nationality, the character of exposed to dust and cumulative length of service, we studied the relationship between genetic polymorphism of IL-8 and pneumoconiosis in order to offer the biomarker for susceptibility group of exposure to dust. Subjects and methods 1265 gold mine workers and 2568 coal workers were investigated. All the workers exposed to dust were examined by dorsaventral chest X-ray base on the health check-up. The pneumoconiosis was diagnosed by the Pneumoconiosis Diagnosis Expert Panel, based on the Chinese National Diagnosis Criteria of Pneumoconiosis (GBZ70-2009).From them, pneumoconiosis diagnosed were selected as cases, workers exposed dust were selected as controls whose age,the cumulative length of service same as control groups. The cases and control worked in the same jop place.Finally, 213 pneumoconiosis patients were selected as cases,including 101 silicosis patients and 112 coal worker`s pneumoconiosis patients, all them were male and Han nationality. Workers exposed dust were selected as controls.The sex, age had no different between the cases and control groups. The cases and control worked in the same jop place. In the case of informed consent, 1.5 ml venous blood was drawn and anticoagulated with EDTA. The polymorphisms of IL-8 (Met31Arg,781C/T,-251A/T,RA+860) were detected using the method of polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) techniques and PCR. Based on many preliminary experiments, the most adaptive PCR reaction, the restriction enzyme digest and electrophoresis system were determined. We analysed the genotypes with pneumoconiosis by using Pearson′sχ2 test and OR.Results1 General informations213 cases were selected, who were male, Han nationality, stageⅠof pneumoconiosis. Their average age was 54.9±9.4 years, average cumulative length of service was 29.3±6.1 years. 251 controls were male and Han nationality too. Their average age was 54.8±6.3 years, average cumulative length of service was 29.5±4.9 years. There are 101 silicosis patients, their average age was 59.1±8.9 year, average cumulative length of service was 28.0±5.1 years.121 controls were male and Han nationality, their average age was 57.3±8.1 year, average cumulative length of service was 28.1±5.2 years. There are 112 coal worker`s pneumoconiosis patients, their age of first diagnosis was 51.1±8.1 year, average cumulative length of service was 29.8±6.7 years. 130 controls were male and Han nationality, their age was 52.4±1.9 year, average cumulative length of service was 30.6±4.7 years. Differences of age, the beginning year of exposure to dust, cumulative length of service between cases and controls were no significance(P>0.05). The two groups had good comparability.2 The distribution of genotype and allele for IL-8 Met31ArgThe frequencies of GG, GT and TT in IL-8 Met31Arg locus in the pneumoconiosis cases were 0%,14.08% and 85.92% and in the controls were 0.40%,3.59% and 96.37%. The difference were significant (P<0.05). The frequencies of G allele were 7.04% and 92.96% in case and control, the frequencies of T allele were 2.19% and 97.81% in case and control. The difference were significant (P<0.05).Using hierarchy analysis, it was found that there were significant difference in the frequencies of IL-8 Met31Arg genotype between the silicosis cases and control groups (P<0.05). The frequencies of GG, GT and TT in IL-8 Met31Arg locus in the silicosis cases were 0%, 12.87% and 87.13% and in the controls were 0.82%,2.48% and 96.69%. The frequencies of G allele were6.44% and 93.56% in case and control, the frequencies of T allele were 2.07%and 97.93% in case and control. The difference were significant (P<0.05).It was found that there were significant difference in the frequencies of IL-8 Met31Arg genotype between the coal worker`s pneumoconiosis cases and control groups (P<0.05). The frequencies of GG, GT and TT in IL-8 Met31Arg locus in the coal worker`s pneumoconiosis cases were 0%, 15.18% and 84.82% and in the controls were 0%,4.62% and 95.38%. The frequencies of G allele were7.59% and 92.41% in case and control, the frequencies of T allele were 2.31% and 97.69% in case and control. The difference were significant (P<0.05).3 The distribution of genotype and allele for IL-8 -251A/TThe frequencies of TT, AT and AA in IL-8 -251A/T locus in the pneumoconiosis cases were 38.97%,48.83% and 12.20% and in the controls were17.93%,17.93% and 64.14%. The difference were significant (P<0.05). The frequencies of T allele were 63.38%and 53.78% in case and control, the frequencies of A allele were 36.62% and 46.22% in case and control. The difference were significant (P<0.05).Using hierarchy analysis, it was found that there were significant difference in the frequencies of IL-8 -251A/T genotype between the silicosis cases and control groups (P<0.05). The frequencies of TT, AT and AA in IL-8 -251A/T locus in the silicosis cases were 0.59%,49.50% and 9.90% and in the controls were 38.84%,35.54% and 25.62%. The frequencies of T allele were 65.35% and 56.61% in case and control, the frequencies of A allele were 34.61% and 43.39% in case and control. There were not significant difference between cases and controls (P>0.05).It was found that there were significant difference in the frequencies of IL-8 -251A/T genotype between the coal worker`s pneumoconiosis cases and control groups (P<0.05). The frequencies of TT, AT and AA in IL-8 -251A/T locus in the coal worker`s pneumoconiosis cases were37.50%,48.21% and 14.29% and in the controls were33.08%,36.15% and 30.77%. The frequencies of T allele were 61.61% and 51.15% in case and control, the frequencies of A allele were 38.39% and 48.85% in case and control. The difference were significant (P<0.05). 4 The distribution of genotype and allele for IL-8 781C/TThe frequencies of CC,CT and TT in IL-8 781C/T locus in the pneumoconiosis cases were 37.09%,41.31% and 21.60% and in the controls were43.03%,43.03% and 13.94%. The difference were significant (P<0.05).The frequencies of C allele were 57.75% and 64.54% in case and control, the frequencies of T allele were 42.25% and 35.46% in case and control. The difference were significant (P<0.05).Using hierarchy analysis, it was not found that there were significant difference in the frequencies of IL-8 781C/T genotype between the silicosis cases and control groups (P>0.05). The frequencies of CC,CT and TT in IL-8 781C/T locus in the silicosis cases were 38.61%,40.59% and 20.79% and in the controls were 46.28%,40.50% and 13.22%. The frequencies of C allele were 58.91% and 66.53% in case and control, the frequencies of T allele were 41.09% and 33.47% in case and control. There were not significant difference between cases and controls (P>0.05).It was found that there were not significant difference in the frequencies of IL-8 781C/T genotype between the coal worker`s pneumoconiosis cases and control groups (P>0.05). The frequencies of CC,CT and TT in IL-8 781C/T locus in the coal worker`s pneumoconiosis cases were35.71%,41.96% and 22.33% and in the controls were 40.00%,45.38% and 14.62%. The frequencies of C allele were56.70% and 62.69% in case and control, the frequencies of T allele were 43.30% and 37.31% in case and control. There were not significant difference (P>0.05) 5 The distribution of genotype and allele for IL-8 RA+860The frequencies of GG,GC and CC in IL-8 RA+860 locus in the pneumoconiosis cases were 74.24%,23.47% and 3.29% and in the control were 73.31%,20.32% and 6.37%. There were not significant difference (P >0.05).The frequencies of G allele were 84.98% and 83.47% in case and control, the frequencies of C allele were 15.02% and 16.53% in case and control. There were not significant difference (P>0.05).Using hierarchy analysis, it was not found that there were significant difference in the frequencies of IL-8 RA+860 genotype between the silicosis cases and control groups (P>0.05). The frequencies of GG,GC and CC in IL-8 RA+860 locus in the silicosis cases were 75.25%,21.78% and 2.97% and in the control were 80.17%,14.88% and 4.96%. The frequencies of G allele were 86.14% and 87.60 in case and control, the frequencies of C allele were 13.86% and 12.40% in case and control. There were not significant difference between cases and controls (P >0.05).It was found that there were not significant difference in the frequencies of IL-8 RA+860 genotype between the coal worker`s pneumoconiosis cases and control groups (P>0.05). The frequencies of GG,GC and CC in IL-8 RA+860 locus in the coal worker`s pneumoconiosis cases were 71.43%,25.00% and 3.57% and in the control were 66.92%,25.38% and 7.70%. The frequencies of G allele were 83.93% and 79.62% in case and control, the frequencies of C allele were 16.07% and 20.38% in case and control. There were not significant difference (P >0.05)Conclusions1 The genetic polymorphism of IL-8(Met31Arg) relatied to susceptibility of pneumoconiosis.Workers of carrying IL-8(Met31Arg)GT genotype were likely to increase the risk of pneumoconiosis.2 IL-8(-251)related to the development of pneumoconiosis. Workers of carrying IL-8(-251)AA genotype were likely to reduce the risk of pneumoconiosis.3 The genetic polymorphism of IL-8(781C/T) relatied to susceptibility of pneumoconiosis.Workers of carrying IL-8(781C/T)TT genotype were likely to increase the risk of pneumoconiosis.4 The relationship between genetic polymorphism of RA+860 and pneumoconiosis hasn’t been found in the study.更多还原