应答蛋白基因多态性对矽肺发生的修饰作用
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摘要
矽肺是我国最为严重的职业卫生问题之一，其发病机制相当复杂至今尚未明确，可能涉及：矽尘诱导肺泡巨噬细胞产生各种细胞调节因子、诱发氧化应激损伤等，从而引起肺成纤维细胞增殖及细胞外基质的沉积。机体在应激状态下大量诱导合成应答蛋白，对细胞防御具有重要意义。目前关于应答蛋白与矽肺的相关研究较少，本课题采用离体实验和人群分子流行病学方法探讨应答蛋白与矽肺发生的关系。
离体实验：大鼠肺泡巨噬细胞(AMs)和人胚肺成纤维细胞(HELF)组成体外模型。采用逆转录多聚酶链反应(RT-PCR)技术测定二氧化硅粉尘处理AMs的培养上清刺激HELF应答蛋白如热休克蛋白70(HSP70)各型(HSP70-1、HSP70-2和HSP70-hom)、热休克蛋白27(HSP27)、金属硫蛋白(MT)亚型1A、2A和血红素氧合酶-1(HO-1)以及与ECM合成、降解有关的基质金属蛋白酶-9(MMP9)及其组织抑制剂1(TIMP1)mRNA表达，观察其诱导表达水平的变化规律。同时，利用蛋白质组学的双向凝胶电泳分离技术与质谱鉴定技术(2-DE-MS)对模型中HELF蛋白表达进行分析并采用免疫印迹法(western blot)对其中的HSP27进行验证。
一定范围内，HELF的HSP70-1、HSP70-2、HSP70-hom以及HO-1、HSP27等应答蛋白mRNA表达与二氧化硅(SiO_2)粉尘浓度呈现良好的剂量-效应关系和时间效应关系(P＜0.05)，但不同的应答蛋白有不同的诱导动力学特征。各种应答蛋白被诱导表达达到峰值时间各有不同。MT1A、MT2A的mRNA表达虽能被染尘AMs上清诱导，但无明显的剂量-效应关系(P＞0.05)。MMP9、TIMP1能被明显诱导表达，但MMP9、TIMP1mRNA表达存在一定的时差，早期以MMP9增高为主，随着时间的延长，以TIMP1表达增加更为显著。
2-DE-MS初步鉴定了经诱导的HELF的数个蛋白：HSP27、PeroxiredoxinⅡ亚型b、钙网蛋白-3和钙网蛋白-5类似物等。其中经western blot检测证实HSP27蛋白可被染尘AMs上清诱导表达，其表达水平随染尘剂量的增加而增加，与HSP27mRNA表达趋势基本一致。
人群分子流行病学：采用病例-对照方法。选择中国西南某两大煤矿519例矿工为研究对象，其中接尘对照294例，确诊的矽肺患者225例(Ⅰ期134例、Ⅱ期73例、Ⅲ期18例)。采用工种接触数值估测(JEM)对累积粉尘暴露进行接触评估；对矿工的吸烟、饮酒等生活习惯、职业卫生习惯、职业危害认知状况、心理状况及营养状况等进行详细的问卷调查；进行肺功能检查并调查呼吸困难情况等对接尘矿工的健康效应进行评价；采用采用酶联免疫吸附测定(ELISA)方法对174例非吸烟人群(41名健康对照、133名矿工)血清中的MMP9／TIMP1进行测定；采用聚合酶链式反应-限制性内切酶片段长度多态性(PCR-RFLP)基因分型技术对519例矿工的MMP9(C-1562T)、HSP70-1(G190C)、HSP70-2(A1267G)和HSP70-hom(T2437C)基因进行基因分型。
粉尘接触评估经单因素方差分析：JEM在病例组显著高于对照(P＜0.01)。病例组中Ⅰ期矽肺平均JEM：105.4±46.9，Ⅱ期矽肺平均JEM：114.3±40.3，Ⅲ期矽肺平均JEM：92.3±34.3，其别间差异有统计学差异(P＜0.01)。相关分析显示：JEM与矽肺分期呈明显的正相关(P＜0.01)。研究现场粉尘现况为：各项指标基本达标，但是个别工种平均浓度还有超标现象，总粉尘浓度甚至达到30％。提示还存在一定的隐患。
对各期矽肺患者的肺通气功能测定并结合详细的个体调查进行分析：矽肺组肺功能参数中用力肺活量(FVC)、一秒钟用力呼气量(FEV1.0)、FEV1.0／FVC的实测值、实测值与预测值百分比均显著低于对照组(P＜0.05)；各项肺功能参数随着JEM分级的增加而呈下降的趋势(P＜0.05)；趋势卡方检验：肺功能异常类型发生率从高到低依次为限制性通气功能障碍、阻塞性通气功能障碍、混合性通气功能障碍。两分类Logistic回归分析，矿工肺功能障碍有统计学差异的影响因素按OR值从大到小依次排列为：Ⅱ+Ⅲ期矽肺(OR=12.557)、Ⅰ期矽肺(4.863)、BI＜400(2.971)、BI600～(2.504)、JEM150～(2.390)、BI400～(2.300)、慢支(1.715)。
174例非吸烟人群(健康对照41例，接尘工人64例，矽肺患者69例)血清MMP9及TIMP1检测结果经秩和检验：与健康对照组比较，接尘工人血清中MMP9增加较为明显(P＜0.05)，而TIMP1增加的差异没有统计学意义(P＞0.05)，各期矽肺的MMP9、TIMP1浓度呈增加趋势，而MMP9／TIMP1比值呈下降趋势，且与健康对照组相比，差异有统计学意义(P＜0.05)。经JEM分级与年龄分级校正，与接尘对照组比较，Ⅰ期矽肺患者的TIMP1升高(P＜0.05)，Ⅱ期矽肺患者MMP9／TIMP1比值增加(P＜0.05)以及Ⅲ期矽肺患者的MMP9、TIMP1升高(P＜0.05)，而MMP9／TIMP1呈下降趋势(P＜0.01)。
基因分型结果经卡方检验：HSP70-2(A1267G)等位基因频率在矽肺各期间分布差异有边缘统计学意义(P=0.046)，G位点的携带率在Ⅱ+Ⅲ期矽肺中明显高于Ⅰ期矽肺和接尘对照组(P＜0.05)。Ⅱ+Ⅲ期矽肺的HSP70-hom(T2437C)的突变纯合子(C／C)携带率高于接尘对照组和Ⅰ期矽肺(8.8％vs2.4％和3％)，在相同JEM分级和相同的布莱克曼吸烟指数(BI)分级，携带突变纯合基因型的个体发生矽肺的风险明显高于杂合子携带个体与野生纯合子携带个体(P＜0.05)。
MMP9-1562C／C野生纯合子基因型与HSP70-2G位点携带者在Ⅱ+Ⅲ期矽肺中的分布与对照和Ⅰ期矽肺组比较差异有临界差异(P=0.087)；JEM(150～)水平下，HSP70-hom2437C突变位点的携带者中，其分布在矽肺与对照组比较也表现出临界差异(P=0.094)。吸烟和基因间的交互作用分析没有发现统计学差异(P＞0.05)。LDA分析显示HSP70-1(190)、HSP70-2(1267)和HSP70-hom(2437)三对等位基因处于高度连锁不平衡；PHASE软件进行单倍型分析：HSP70-1，HSP70-2和HSP70-hom的190，1267和2437等位基因构成的G-A-T和G-G-T单倍型在Ⅱ+Ⅲ期矽肺中明显增加，与Ⅰ期矽肺和接尘对照相比差异均有统计学意义(P＜0.05)。
综上所述，HELF应答蛋白HSP70各型(HSP70-1、HSP70-2和HSP70-hom)、HSP27、HO-1以及MMP9／TIMP1mRNA的表达能被染SiO_2粉尘的AMs上清诱导并呈现一定的剂量-反应和时间-效应关系，但各自存在不同的诱导动力学特征，提示可能是因为被研究基因表达的调控序列存在差异；不同应答蛋白表达存在时相差，交错对细胞起到保护作用。MMP9／TIMP1mRNA表达时相差异提示早期以MMP9表达及活性增加，基质降解为主；TIMP1出现对MMP9起拮抗作用，随着MMP9／TIMP平衡打破，胶原沉积。HSP27、PeroxiredoxinⅡ及钙网蛋白等与SiO_2引起的氧化应激密切相关。
预防矽肺的发生除了通过控制作业场所的粉尘浓度，同时做好个体防护，减少粉尘暴露；加强矿工职业卫生教育和健康促进，识别易感人群(如HSP70-hom2437C的C／C基因型、以及HSP70-1，HSP70-2和HSP70-hom的G-G-T、G-A-T单倍型携带者)。为达到WHO消除矽肺的目标而奋斗。
Silicosis has long been one of the most serious and prevalent occupational healthproblems in China for its high morbidity and mortality. Although their pathogenesis remainsincompletely understood, some evidence suggest the participation of cytokines produced byAMs and oxidative stress induced by silica leading to the fibroblast proliferation andextracellular matrix(ECM) deposition. The heat stress proteins (HSPs) can be induced andplay important role for the cell protection as a result of various mechanical or chemicalstresses. A little is known about relation between HSPs and silicosis,the aim of this study is toexplore the roles of response proteins on occurrence of silicosis by in vitro experiments_andmolecular epidemiology.
Sprague-Dawley (SD) rat alveolar macrophages (AMs) and human embryonic lungfibroblasts (HELF) was formed into an in vitro model. The mRNA levels of heat stressproteins(HSPs), such as HSP70-1, HSP70-2,HSP70-hom, HSP27, metallothionein (MT)1A、MT2A, haemoxygenase-1(HO-1) and the reported factors related to the synthesis ordegradation of ECM, metalloproteinases-9(MMP9) and tissue inhibitor ofmetalloproteinases-1(TIMP1) of HELF stimulated by supernatants of AMs exposed to silicawere measured by RT-PCR technology. At the same time, identification of proteins of treatedHELF was conducted by two-dimensional polyacrylamide gel electrophoresis(2-DE) andmatrix assisted laser desorption-ionization-time of flight massspectrometry(MALDI-TOF-MS). One of identified proteins,HSP27,was further validated byWestern blot assay.
It was shown that there is good dose-effect and time-effect relationships betweenHSP70-1, HSP70-2, HSP70-hom, HSP27, HO-1, MMP9 and TIMP1 mRNA levels andconcentration of silica and supernatants exposure time (P＜0.05). Different response proteinshad their own induction dynamic characters, and the time they reach the peak expression wasdiversity. It is not found dose-effect relationship between the MT1A, MT2A mRNA levels andthe concentration of silica, while they can be induced by the supernatants(P＞0.05). MMP9,TIMP1 mRNA can be obviously induced and MMP9 mRNA levels increased earlier thanthose of TIMP1, but TIMP1 increased more obviously than MMP9 with time continuing.
Several proteins in induced HELF were identified by 2-DE-MS: PeroxiredoxinⅡ isoform b, heat shock protein 27, a protein similar to calreticulin isoform 3, a protein similar tocalreticulin isoform 5. HSP27 was further validated by western blot and the results showedthat it increased with the concentration of silica increasing. The change tendency ofHSP27mRNA and protein expression was consistent.
Epidemiologic investigation: Case-control study was applied in our study. Weinvestigated 519 workers from two large coal plants in Southwestern China, including 225patients with silicosis (Ⅰ: 134;Ⅱ, 73;Ⅲ, 18) and 294 control miners(self-reported to beHan descent, the primary ethnic group in China) with similar coal mine dust-exposurehistories but without apparent pulmonary disease or inflammation. Job exposure matrix(JEM)was used to assess the exposure to accumulated dust. Uniform Questionnaire was applied toevery subject for collecting the personal information including cigarette smoking, alcoholconsumption, occupational health habit, cognition of occupational hazard, psychological statusand nutrition status. ELISA method was used to determine Serum MMP9/TIMP1 in 174nonsmokers (41 healthy controls and 133 workers). PCR-RFLP-was used to genotype 519workers by MMP9(C-1562T)、HSP70-1(G190C)、HSP70-2(A1267G) andHSP70-hom(T2437C).
It was found that JEM (Arithmetic mean, AM) in the cases was significantly higher thanthat in the control miners by one-way ANOVA (Ⅰ: 105.4±46.9;Ⅱ: 114.3±40.3;Ⅲ:92.3±34.3; p＜0.01). Silicosis category has positive correlation to JEM (P＜0.01). Althougheach index of dust exposure was under the occupational exposure limit (OEL), the mean dustconcentration of some job categories were over it. At some workplace, the total dustconcentrations reached 30%.
The measurement of pulmonary ventilation function showed that each lung function index,including FVC, FEV1.0 and FEV1.0/FVC (measured value and predicted value), weresignificantly lower in the silicosis group than in the control miners (p＜0.05) and degradedfollowing the increase of JEM category (p＜0.05). The prevalence of lung dysfunction categorywas trended as follows by Chi-square test: Restricted ventilation dysfunction＞Obstructedventilation dysfunction＞Mixed ventilation dysfunction. The odds ratio (OR) of lungdysfunction in the workers was obtained by Logistic regression analysis and listed as follows,Ⅱ+Ⅲsilicosis (12.557),Ⅰcategory silicosis (4.863), BI＜400(2.971), BI 600～(2.504),JEM150～(2.390), BI 400～(2.300), chronic bronchitis (1.715).
Serum MMP9 in the 64 control miners was significantly higher than that in the 41 healthycontrols by Rank test (p＜0.05).There was no such relationship in serum TIMP1 (P＞0.05). Asthe silicosis category increased, the concentrations of serum MMP9 and TIMP1 were also increased, but the ratio of MMP9/TIMP1 was decreased, both of these end points weresignificantly higher in each silicosis category than that in the healthy controls (p＜0.05).Compared with control miners in the adjusted model considering the JEM category and agecategory, TIMP1 level in I silicosis patients was significantly higher (p＜0.05), MMP9/TIMP1ratio in theⅡsilicosis patients was significantly higher (p<0.05), MMP9 and TIMP1 level intheⅢsilicosis patients were significantly higher (p＜0.05), but MMP9/TIMP1 ratio in theⅢsilicosis patients were significantly lower (p＜0.01)
The frequency of HSPT0-2 1267G was significant different in each silicosis category(P=0.046), moreⅡandⅢpatients carried G allele than that those with I silicosis category andcontrol miners ((P＜0.05). The frequency of HSP70-hom(T2437C) C/C carriers in theⅡandⅢsilicosis patients was higher than that in control miners andⅠsilicosis patients (8.8% vs2.4%和3%). Adjusted by JEM category and BI category, silicosis risk in the homozygous major allelecarriers was significantly higher than that in the heterozygous allele carriers and homozygousminor allele carriers(P＜0.05).
The distribution of MMP9-1562C/C carriers and HSP70-2G carriers were marginallydifferent betweenⅡandⅢsilicosis patients andⅠsilicosis and control miners (P=0.087). Inthe subgroup with JEM more than 150, the frequency of HSP70-hom2437C carriers was alsomarginally different in the silicosis group compared with the control miners (P=0.094). Nointeraction was found between cigarette smoking and genotypes. HSP70-1(190)、HSP70-2(1267) and HSP70-hom(2437) were highly linked with each other by LDA software.Haplotype analysis by PHASE analysis soft found that G-A-T and G-G-T haplotypes of thesethree SNPs was more frequent in theⅡandⅡcategory silicosis compared to theⅠsilicosisand controls(P＜0.05).
In conclusion, the expression of HSP70-1, HSP70-2,HSP70-hom, HSP27, HO-1 andMMP9/TIMP1 mRNA of HELF could be induced by AMs supernatants exposed to SiO_2,which displayed a clear dose-effect and time-effect relationships. They had differentcharacteristics of induction, which might be caused by the differences of the expression ofregular sequence in the genes. There were phase differences in the expression of differentresponse protein, which could protect the cells in turn. The phase difference of the expressionof MMP9/TIMP1 mRNA indicated MMP9 expression and activity increased in early phase,and ECM degraded. The appearance of TIMP1 inhibited the activity of MMP9, and theMMP9/TIMP imbalance appeared, which prompted collagen deposition. HSP27,PeroxiredoxinⅡand calreticulin had a closed correlation with oxidative-stress induced bySiO_2.
In order to prevent silicosis, we not only control the dust concentration in the workplace,but also improve personal protection and decrease the dust exposure at the same time. We alsoneed to enhance occupational health education and health promotion in miners, and identifysusceptible population (HSP70-hom2437C C/C genotype, G-A-T or G-G-T haplotype carriersof HSP70-1, HSP70-2, HSP70-hom). Struggle for the WHO goal of eliminating silicosis.
离体实验：大鼠肺泡巨噬细胞(AMs)和人胚肺成纤维细胞(HELF)组成体外模型。采用逆转录多聚酶链反应(RT-PCR)技术测定二氧化硅粉尘处理AMs的培养上清刺激HELF应答蛋白如热休克蛋白70(HSP70)各型(HSP70-1、HSP70-2和HSP70-hom)、热休克蛋白27(HSP27)、金属硫蛋白(MT)亚型1A、2A和血红素氧合酶-1(HO-1)以及与ECM合成、降解有关的基质金属蛋白酶-9(MMP9)及其组织抑制剂1(TIMP1)mRNA表达，观察其诱导表达水平的变化规律。同时，利用蛋白质组学的双向凝胶电泳分离技术与质谱鉴定技术(2-DE-MS)对模型中HELF蛋白表达进行分析并采用免疫印迹法(western blot)对其中的HSP27进行验证。
一定范围内，HELF的HSP70-1、HSP70-2、HSP70-hom以及HO-1、HSP27等应答蛋白mRNA表达与二氧化硅(SiO_2)粉尘浓度呈现良好的剂量-效应关系和时间效应关系(P＜0.05)，但不同的应答蛋白有不同的诱导动力学特征。各种应答蛋白被诱导表达达到峰值时间各有不同。MT1A、MT2A的mRNA表达虽能被染尘AMs上清诱导，但无明显的剂量-效应关系(P＞0.05)。MMP9、TIMP1能被明显诱导表达，但MMP9、TIMP1mRNA表达存在一定的时差，早期以MMP9增高为主，随着时间的延长，以TIMP1表达增加更为显著。
2-DE-MS初步鉴定了经诱导的HELF的数个蛋白：HSP27、PeroxiredoxinⅡ亚型b、钙网蛋白-3和钙网蛋白-5类似物等。其中经western blot检测证实HSP27蛋白可被染尘AMs上清诱导表达，其表达水平随染尘剂量的增加而增加，与HSP27mRNA表达趋势基本一致。
人群分子流行病学：采用病例-对照方法。选择中国西南某两大煤矿519例矿工为研究对象，其中接尘对照294例，确诊的矽肺患者225例(Ⅰ期134例、Ⅱ期73例、Ⅲ期18例)。采用工种接触数值估测(JEM)对累积粉尘暴露进行接触评估；对矿工的吸烟、饮酒等生活习惯、职业卫生习惯、职业危害认知状况、心理状况及营养状况等进行详细的问卷调查；进行肺功能检查并调查呼吸困难情况等对接尘矿工的健康效应进行评价；采用采用酶联免疫吸附测定(ELISA)方法对174例非吸烟人群(41名健康对照、133名矿工)血清中的MMP9／TIMP1进行测定；采用聚合酶链式反应-限制性内切酶片段长度多态性(PCR-RFLP)基因分型技术对519例矿工的MMP9(C-1562T)、HSP70-1(G190C)、HSP70-2(A1267G)和HSP70-hom(T2437C)基因进行基因分型。
粉尘接触评估经单因素方差分析：JEM在病例组显著高于对照(P＜0.01)。病例组中Ⅰ期矽肺平均JEM：105.4±46.9，Ⅱ期矽肺平均JEM：114.3±40.3，Ⅲ期矽肺平均JEM：92.3±34.3，其别间差异有统计学差异(P＜0.01)。相关分析显示：JEM与矽肺分期呈明显的正相关(P＜0.01)。研究现场粉尘现况为：各项指标基本达标，但是个别工种平均浓度还有超标现象，总粉尘浓度甚至达到30％。提示还存在一定的隐患。
对各期矽肺患者的肺通气功能测定并结合详细的个体调查进行分析：矽肺组肺功能参数中用力肺活量(FVC)、一秒钟用力呼气量(FEV1.0)、FEV1.0／FVC的实测值、实测值与预测值百分比均显著低于对照组(P＜0.05)；各项肺功能参数随着JEM分级的增加而呈下降的趋势(P＜0.05)；趋势卡方检验：肺功能异常类型发生率从高到低依次为限制性通气功能障碍、阻塞性通气功能障碍、混合性通气功能障碍。两分类Logistic回归分析，矿工肺功能障碍有统计学差异的影响因素按OR值从大到小依次排列为：Ⅱ+Ⅲ期矽肺(OR=12.557)、Ⅰ期矽肺(4.863)、BI＜400(2.971)、BI600～(2.504)、JEM150～(2.390)、BI400～(2.300)、慢支(1.715)。
174例非吸烟人群(健康对照41例，接尘工人64例，矽肺患者69例)血清MMP9及TIMP1检测结果经秩和检验：与健康对照组比较，接尘工人血清中MMP9增加较为明显(P＜0.05)，而TIMP1增加的差异没有统计学意义(P＞0.05)，各期矽肺的MMP9、TIMP1浓度呈增加趋势，而MMP9／TIMP1比值呈下降趋势，且与健康对照组相比，差异有统计学意义(P＜0.05)。经JEM分级与年龄分级校正，与接尘对照组比较，Ⅰ期矽肺患者的TIMP1升高(P＜0.05)，Ⅱ期矽肺患者MMP9／TIMP1比值增加(P＜0.05)以及Ⅲ期矽肺患者的MMP9、TIMP1升高(P＜0.05)，而MMP9／TIMP1呈下降趋势(P＜0.01)。
基因分型结果经卡方检验：HSP70-2(A1267G)等位基因频率在矽肺各期间分布差异有边缘统计学意义(P=0.046)，G位点的携带率在Ⅱ+Ⅲ期矽肺中明显高于Ⅰ期矽肺和接尘对照组(P＜0.05)。Ⅱ+Ⅲ期矽肺的HSP70-hom(T2437C)的突变纯合子(C／C)携带率高于接尘对照组和Ⅰ期矽肺(8.8％vs2.4％和3％)，在相同JEM分级和相同的布莱克曼吸烟指数(BI)分级，携带突变纯合基因型的个体发生矽肺的风险明显高于杂合子携带个体与野生纯合子携带个体(P＜0.05)。
MMP9-1562C／C野生纯合子基因型与HSP70-2G位点携带者在Ⅱ+Ⅲ期矽肺中的分布与对照和Ⅰ期矽肺组比较差异有临界差异(P=0.087)；JEM(150～)水平下，HSP70-hom2437C突变位点的携带者中，其分布在矽肺与对照组比较也表现出临界差异(P=0.094)。吸烟和基因间的交互作用分析没有发现统计学差异(P＞0.05)。LDA分析显示HSP70-1(190)、HSP70-2(1267)和HSP70-hom(2437)三对等位基因处于高度连锁不平衡；PHASE软件进行单倍型分析：HSP70-1，HSP70-2和HSP70-hom的190，1267和2437等位基因构成的G-A-T和G-G-T单倍型在Ⅱ+Ⅲ期矽肺中明显增加，与Ⅰ期矽肺和接尘对照相比差异均有统计学意义(P＜0.05)。
综上所述，HELF应答蛋白HSP70各型(HSP70-1、HSP70-2和HSP70-hom)、HSP27、HO-1以及MMP9／TIMP1mRNA的表达能被染SiO_2粉尘的AMs上清诱导并呈现一定的剂量-反应和时间-效应关系，但各自存在不同的诱导动力学特征，提示可能是因为被研究基因表达的调控序列存在差异；不同应答蛋白表达存在时相差，交错对细胞起到保护作用。MMP9／TIMP1mRNA表达时相差异提示早期以MMP9表达及活性增加，基质降解为主；TIMP1出现对MMP9起拮抗作用，随着MMP9／TIMP平衡打破，胶原沉积。HSP27、PeroxiredoxinⅡ及钙网蛋白等与SiO_2引起的氧化应激密切相关。
预防矽肺的发生除了通过控制作业场所的粉尘浓度，同时做好个体防护，减少粉尘暴露；加强矿工职业卫生教育和健康促进，识别易感人群(如HSP70-hom2437C的C／C基因型、以及HSP70-1，HSP70-2和HSP70-hom的G-G-T、G-A-T单倍型携带者)。为达到WHO消除矽肺的目标而奋斗。
Silicosis has long been one of the most serious and prevalent occupational healthproblems in China for its high morbidity and mortality. Although their pathogenesis remainsincompletely understood, some evidence suggest the participation of cytokines produced byAMs and oxidative stress induced by silica leading to the fibroblast proliferation andextracellular matrix(ECM) deposition. The heat stress proteins (HSPs) can be induced andplay important role for the cell protection as a result of various mechanical or chemicalstresses. A little is known about relation between HSPs and silicosis,the aim of this study is toexplore the roles of response proteins on occurrence of silicosis by in vitro experiments_andmolecular epidemiology.
Sprague-Dawley (SD) rat alveolar macrophages (AMs) and human embryonic lungfibroblasts (HELF) was formed into an in vitro model. The mRNA levels of heat stressproteins(HSPs), such as HSP70-1, HSP70-2,HSP70-hom, HSP27, metallothionein (MT)1A、MT2A, haemoxygenase-1(HO-1) and the reported factors related to the synthesis ordegradation of ECM, metalloproteinases-9(MMP9) and tissue inhibitor ofmetalloproteinases-1(TIMP1) of HELF stimulated by supernatants of AMs exposed to silicawere measured by RT-PCR technology. At the same time, identification of proteins of treatedHELF was conducted by two-dimensional polyacrylamide gel electrophoresis(2-DE) andmatrix assisted laser desorption-ionization-time of flight massspectrometry(MALDI-TOF-MS). One of identified proteins,HSP27,was further validated byWestern blot assay.
It was shown that there is good dose-effect and time-effect relationships betweenHSP70-1, HSP70-2, HSP70-hom, HSP27, HO-1, MMP9 and TIMP1 mRNA levels andconcentration of silica and supernatants exposure time (P＜0.05). Different response proteinshad their own induction dynamic characters, and the time they reach the peak expression wasdiversity. It is not found dose-effect relationship between the MT1A, MT2A mRNA levels andthe concentration of silica, while they can be induced by the supernatants(P＞0.05). MMP9,TIMP1 mRNA can be obviously induced and MMP9 mRNA levels increased earlier thanthose of TIMP1, but TIMP1 increased more obviously than MMP9 with time continuing.
Several proteins in induced HELF were identified by 2-DE-MS: PeroxiredoxinⅡ isoform b, heat shock protein 27, a protein similar to calreticulin isoform 3, a protein similar tocalreticulin isoform 5. HSP27 was further validated by western blot and the results showedthat it increased with the concentration of silica increasing. The change tendency ofHSP27mRNA and protein expression was consistent.
Epidemiologic investigation: Case-control study was applied in our study. Weinvestigated 519 workers from two large coal plants in Southwestern China, including 225patients with silicosis (Ⅰ: 134;Ⅱ, 73;Ⅲ, 18) and 294 control miners(self-reported to beHan descent, the primary ethnic group in China) with similar coal mine dust-exposurehistories but without apparent pulmonary disease or inflammation. Job exposure matrix(JEM)was used to assess the exposure to accumulated dust. Uniform Questionnaire was applied toevery subject for collecting the personal information including cigarette smoking, alcoholconsumption, occupational health habit, cognition of occupational hazard, psychological statusand nutrition status. ELISA method was used to determine Serum MMP9/TIMP1 in 174nonsmokers (41 healthy controls and 133 workers). PCR-RFLP-was used to genotype 519workers by MMP9(C-1562T)、HSP70-1(G190C)、HSP70-2(A1267G) andHSP70-hom(T2437C).
It was found that JEM (Arithmetic mean, AM) in the cases was significantly higher thanthat in the control miners by one-way ANOVA (Ⅰ: 105.4±46.9;Ⅱ: 114.3±40.3;Ⅲ:92.3±34.3; p＜0.01). Silicosis category has positive correlation to JEM (P＜0.01). Althougheach index of dust exposure was under the occupational exposure limit (OEL), the mean dustconcentration of some job categories were over it. At some workplace, the total dustconcentrations reached 30%.
The measurement of pulmonary ventilation function showed that each lung function index,including FVC, FEV1.0 and FEV1.0/FVC (measured value and predicted value), weresignificantly lower in the silicosis group than in the control miners (p＜0.05) and degradedfollowing the increase of JEM category (p＜0.05). The prevalence of lung dysfunction categorywas trended as follows by Chi-square test: Restricted ventilation dysfunction＞Obstructedventilation dysfunction＞Mixed ventilation dysfunction. The odds ratio (OR) of lungdysfunction in the workers was obtained by Logistic regression analysis and listed as follows,Ⅱ+Ⅲsilicosis (12.557),Ⅰcategory silicosis (4.863), BI＜400(2.971), BI 600～(2.504),JEM150～(2.390), BI 400～(2.300), chronic bronchitis (1.715).
Serum MMP9 in the 64 control miners was significantly higher than that in the 41 healthycontrols by Rank test (p＜0.05).There was no such relationship in serum TIMP1 (P＞0.05). Asthe silicosis category increased, the concentrations of serum MMP9 and TIMP1 were also increased, but the ratio of MMP9/TIMP1 was decreased, both of these end points weresignificantly higher in each silicosis category than that in the healthy controls (p＜0.05).Compared with control miners in the adjusted model considering the JEM category and agecategory, TIMP1 level in I silicosis patients was significantly higher (p＜0.05), MMP9/TIMP1ratio in theⅡsilicosis patients was significantly higher (p<0.05), MMP9 and TIMP1 level intheⅢsilicosis patients were significantly higher (p＜0.05), but MMP9/TIMP1 ratio in theⅢsilicosis patients were significantly lower (p＜0.01)
The frequency of HSPT0-2 1267G was significant different in each silicosis category(P=0.046), moreⅡandⅢpatients carried G allele than that those with I silicosis category andcontrol miners ((P＜0.05). The frequency of HSP70-hom(T2437C) C/C carriers in theⅡandⅢsilicosis patients was higher than that in control miners andⅠsilicosis patients (8.8% vs2.4%和3%). Adjusted by JEM category and BI category, silicosis risk in the homozygous major allelecarriers was significantly higher than that in the heterozygous allele carriers and homozygousminor allele carriers(P＜0.05).
The distribution of MMP9-1562C/C carriers and HSP70-2G carriers were marginallydifferent betweenⅡandⅢsilicosis patients andⅠsilicosis and control miners (P=0.087). Inthe subgroup with JEM more than 150, the frequency of HSP70-hom2437C carriers was alsomarginally different in the silicosis group compared with the control miners (P=0.094). Nointeraction was found between cigarette smoking and genotypes. HSP70-1(190)、HSP70-2(1267) and HSP70-hom(2437) were highly linked with each other by LDA software.Haplotype analysis by PHASE analysis soft found that G-A-T and G-G-T haplotypes of thesethree SNPs was more frequent in theⅡandⅡcategory silicosis compared to theⅠsilicosisand controls(P＜0.05).
In conclusion, the expression of HSP70-1, HSP70-2,HSP70-hom, HSP27, HO-1 andMMP9/TIMP1 mRNA of HELF could be induced by AMs supernatants exposed to SiO_2,which displayed a clear dose-effect and time-effect relationships. They had differentcharacteristics of induction, which might be caused by the differences of the expression ofregular sequence in the genes. There were phase differences in the expression of differentresponse protein, which could protect the cells in turn. The phase difference of the expressionof MMP9/TIMP1 mRNA indicated MMP9 expression and activity increased in early phase,and ECM degraded. The appearance of TIMP1 inhibited the activity of MMP9, and theMMP9/TIMP imbalance appeared, which prompted collagen deposition. HSP27,PeroxiredoxinⅡand calreticulin had a closed correlation with oxidative-stress induced bySiO_2.
In order to prevent silicosis, we not only control the dust concentration in the workplace,but also improve personal protection and decrease the dust exposure at the same time. We alsoneed to enhance occupational health education and health promotion in miners, and identifysusceptible population (HSP70-hom2437C C/C genotype, G-A-T or G-G-T haplotype carriersof HSP70-1, HSP70-2, HSP70-hom). Struggle for the WHO goal of eliminating silicosis.
引文
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[27] 王万银，钱令嘉．HSP27研究现状．国外医学：生理病理科学与临床分册，2001：21(6)467-9．
[28] 胡晓荣．金属硫蛋白的生理作用及在医学上的应用．西昌农业高等专科学校学报2000；14(1)：47-50．
[29] 常秀丽．金属硫蛋白及其在镉接触评价中的应用．国外医学：卫生学分册2004；31(1)：32-6．
[30] 张桂春．金属硫蛋白的功能及应用前景．烟台师范学院学报：自然科学版2005；21(2)：142-5．
[31] 周敏，杨芳，贺智敏，等．人金属硫蛋白MT-2a的原核表达、纯化及其抗血清的制备．细胞与分子免疫学杂志2006；22(4)：530-2．
[32] Atkinson JJandSenior RM. Matrix metalloproteinase-9 in lung remodeling. Am J Respir Cell Mol Biol 2003; 28 (1): 12-24.
[33] Perez SE, Cano DA, Dao-Pick T, et al. Matrix metalloproteinases 2 and 9 are dispensable for pancreatic islet formation and function in vivo. Diabetes 2005; 54 (3): 694-701.
[34] 曾爱萍，曾水清，程扬，等．Modulaion of Matrix Metalloproteinase and TIMP-1 Expression by TGF-β1 in Cultured Human RPE Cells．华中科技大学学报：医学英德文版2006；26(3)：363-5．
[35] Nguyen M, Arkell JandJackson CJ. Active and tissue inhibitor of matrix metalloproteinase-free gelatinase B accumulates within human microvascular endothelial vesicles. J Biol Chem 1998; 273 (9): 5400-4.
[36] Qing H, Nie, Ya F, et al. Correlation between TIMP-1 expression and liver fibrosis in two rat liver fibrosis models．世界胃肠病学杂志：英文版2006；12(19)：3044-9．
[37] Feldman LJ, Mazighi M, Scheuble A, et al. Differential expression of matrix metalloproteinases after stent implantation and balloon angioplasty in the hypercholesterolemic rabbit. Circulation 2001; 103 (25): 3117-22.
[38] Douthwaite JA, Johnson TS, Haylor JL, et al. Effects of transforming growth factor-betal on renal extracellular matrix components and their regulating proteins. J Am Soc Nephrol 1999; 10 (10): 2109-19.
[39] Legallicier B, Trugnan G, Murphy G, et al. Expression of the type Ⅳ collagenase system during mouse kidney development and tubule segmentation. J Am Soc Nephrol 2001; 12 (11): 2358-69.
[40] Ma XB, Sun SXandYang F. [Effect of human silicotic alveolar macrophages on the expression of matrix metalloproteinase-1 and tissue inhibitor of metalloproteinases-1 in human lung fibroblasts.]. Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za Zhi 2004; 22 (5): 358-60.
[41] Braundmeier AG, Fazleabas AT, Lessey BA, et al. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) Regulates Metalloproteinases in Human Uterine Endometrium. J Clin Endocrinol Metab 2006.
[42] Scabilloni JF, Wang L, Antonini JM, et al. Matrix metalloproteinase induction in fibrosis and fibrotic nodule formation due to silica inhalation. Am J Physiol Lung Cell Mol Physiol 2005; 288(4): L709-17.
[43] Li H, Cui D, Tong X, et al. [The role of matrix metalloproteinases in extracellular matrix remodelling in chronic obstructive pulmonary disease rat models]. Zhonghua Nei Ke Za Zhi 2002; 41(6): 393-8.
[44] Cheng Z, Winant RCandGambhir SS. A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. J Nucl Med 2005; 46 (5): 878-86.
[45] Westergren-Thorsson G, Malmstrom JandMarko-Varga G. Proteomics — the protein expression technology to study connective tissue biology. J Pharm Biomed Anal 2001; 24(5-6): 815-24.
[46] Zhou Y, Yu C, Miao X, et al. Functional haplotypes in the promoter of matrix metalloproteinase-2 and lung cancer susceptibility. Carcinogenesis June 2005; 26 (6): 1117-21.
[47] Ueki A, Isozaki Y, Tomokuni A, et al. Different distribution of HLA class II alleles in anti-topoisomerase I autoantibody responders between silicosis and systemic sclerosis patients, with a common distinct amino acid sequence in the HLA-DQB1 domain. Immunobiology 2001; 204(4): 458-65.
[48] Bretana A, Goihman-Yahr M, Tapia FJ, et al. Comparative ultrastructure and immunolabeling of MHC-II antigens of alveolar macrophages obtained from patientswith paracoccidioidomycosis and other lung diseases. J Leukoc Biol 1995; 57 (1): 101-9.
[49] Honda K, Kimura A, Dong RP, et al. Immunogenetic analysis of silicosis in Japan. Am J Respir Cell Mol Biol 1993; 8(1): 106-11.
[50] Migliaccio CT, Hamilton RF, Jr.andHolian A. Increase in a distinct pulmonary macrophage subset possessing an antigen-presenting cell phenotype and in vitro APC activity following silica exposure. Toxicol Appl Pharmacol 2005:205 (2): 168-76.
[51] Shanklin DRandSmalley DL. The immunopathology of siliconosis. History, clinical presentation, and relation to silicosis and the chemistry of silicon and silicone. Immunol Res 1998; 18 (3): 125-73.
[52] Zetterberg G, Elmberger G, Johansson A, et al. Rat alveolar and interstitial macrophages in the fibrosing stage following quartz exposure. Hum Exp Toxicol 2000; 19 (7): 402-11.
[53] Nadir R, Jedlicka A, Mintz M, et al. Effect of TNF and LTA polymorphisms on biological markers of response to oxidative stimuli in coal miners: a model of gene-environment interaction. Turnout necrosis factor and lymphotoxin alpha. J Med Genet2003; 40 (2): 96-103.
[54] Kim KA, Cho YY, Cho JS, et al. Tumor necrosis factor-alpha gene promoter polymorphism in coal workers' pneumoconiosis. Mol Cell Biochem 2002:234-235 (1-2): 205-9.
[55] Vanhee D, Gosset P, Marquette CH, et al. Secretion and mRNA expression of TNF alpha and IL-6 in the lungs of pneumoconiosis patients. Am J Respir Crit Care Med 1995; 152 (1): 298-306.
[56] Nadif R, Mintz M, Marzec J, et al. IL18 and IL18R1 potymorphisms, lung HRCT and fibrosis: a longitudinal study in coal miners. Eur Respir J 2006.
[57] Barbarin V, Xing Z, Delos M, et al. Pulmonary overexpression of IL-10 augments lung fibrosis and Th2 responses induced by silica particles. Am J Physiol Lung Cell Mol Physiol 2005; 288(5): L841-8.
[58] Lesur OJ, Mancini NM, Humbert JC, et al. Interleukin-6, interferon-gamma, and phospholipid levels in the alveolar lining fluid of human lungs. Profiles in coal worker's pneumoconiosis and idiopathic pulmonary fibrosis. Chest 1994:106 (2): 407-13.
[59] Lassalle P, Gosset P, Aerts C, et al. Abnormal secretion of interleukin-1 and tumor necrosis factor alpha by alveolar macrophages in coal worker's pneumoconiosis: comparison between simple pneumoconiosis and progressive massive fibrosis. Exp Lung Res 1990; 16(1): 73-80.
[60] 任鹏，吴秀珍，赵静，等．转化生长因子-β与肺纤维化．中国煤炭工业医学杂志，2006：9(6)：535-7．
[61] 朱剑萍，姚宏伟，陈季强．肺泡上皮细胞在肺纤维化中的作用及机制研究进展．国际呼吸杂志，2006；26(7)：506-9．
[62] Yao W, Wang ZM, Wang MZ, et al. [Oxidative injury and serum cytokines in coal workers with pneumoconiosis]. Sichuan Da Xue Xue Bao Yi Xue Ban 2005; 36 (4): 510-2.
[63] 张国高，贺涵贞．热应激蛋白及其在职业医学中应用研究的展望．中华劳动卫生职业病杂志1998：16(2)：67-9．
[64] Sawatzky DA, Foster R, Seed MP, et al. Heat-shock proteins and their role in chondrocyte protection, an application for autologous transplantation. Inflammopharmacology2005; 12 (5): 569-89.
[65] Barrow REandDasu MR. Oxidative and heat stress gene changes in hypertrophic scar fibroblasts stimulated with interleukin-1beta. J Surg Res 2005; 126 (1): 59-65.
[66] Torronteguy C, Frasson A, Zerwes F, et al. Inducible heat shock protein 70 expression as a potential predictive marker of metastasis in breast tumors. Cell Stress Chaperones 2006; 11 (1): 34-43.
[67] Simko M, Hartwig C, Lantow M, et al. Hsp70 expression and free radical release after exposure to non-thermal radio-frequency electromagnetic fields and ultrafine particles in human Mono Mac 6 cells. Toxicol Lett 2006; 161 (1): 73-82.
[68] Multhoff G. Heat shock proteins in immunity. Handb Exp Pharmacol 2006 (172): 279-304.
[69] Bogatu B Fau-Contag B, Contag B, Ishihara Y Fau-Nishikawa T, et al. Adsorption of lipoproteins onto mineral dust surfaces: a possible factor in the pathogenesis of particle-induced pulmonary fibrosis。Expression of matrix metalloproteinase, tissue inhibitors of metalloproteinase and adhesion molecules in silicotic mice with lung tumor metastasis. (0341-0382 (Print)).
[70] Cerdan S Fau-Rodrigues TB, Rodrigues Tb Fau-Sierra A, Sierra A Fau-Benito M, et al. The redox switch/redox coupling hypothesis[Significance of current bio- and molecular-markers in urological cancers] A review of the cytokine network in multiple myeloma: diagnostic, prognostic, and therapeutic implications. (0197-0186 (Print)).
[71] 何燕，邓国忠，高天．肺纤维化细胞因子机制研究进展．中华创伤杂志，2006；22(6)：477-80．
[72] Porter DW, Millecchia LL, Willard P, et al. Nitric Oxide and Reactive Oxygen Species Production Causes Progressive Damage in Rats after Cessation of Silica Inhalation. Toxicological Sciences March 2006; 90 (1): 188-97.
[73] Delgado L, Parra ERandCapelozzi VL. Apoptosis and extracellular matrix remodelling in human silicosis. Histopathology 2006; 49 (3): 283-9.
[74] Radons JandMulthoff G. Immunostimulatory functions of membrane-bound and exported heat shock protein 70. Exert Immunol Rev 2005; 11: 17-33.
[75] Melchers I. [Immunogenetics--HLA-association, molecular chaperones and "related" diseases]. Z Rheumatol 2005; 64 (6): 402-7.
[76] van Eden W. Immunoregulation of autoimmune diseases. Hum Immunol 2006; 67 (6): 446-53.
[77] Copland IBandPost M. Stretch-activated signaling pathways responsible for early response gene expression in fetal lung epithelial cells. J Cell Physiol 2006; 210 (1): 133-43.
[78] Ito H, Sekimura K, Sasaki N, et al. [Cytoprotective effects of heat shock protein-70 in bronchial epithelium against neutrophil elastase-induced cell injury]. Arerugi 2006; 55 (7): 820-6.
[79] 赵京山，尹桂山．金属硫蛋白的生理功能及其研究进展．生命的化学1998；18(3)：9-11．
[80] 刘志勇，魏国林．金属硫蛋白研究进展．江西科学2004；22(2)：104-9．
[81] 李守明，刘志勇．金属硫蛋白与疾病关系．江西医药2004；39(6)：434-7．
[82] 安建平，王海娟，韩沛平，等．属硫蛋白生理功能的研究进展．天水师范学院学报2005；25 (2)：34-8．
[83] Yang F Fau-Zhou M, Zhou M Fau-He Z, He Z Fau-Liu X, et al. High-yield expression in Escherichia coli of soluble human MT2A with native functions A BamHI RFLP at MT2A on human chromosome 16. (1046-5928 (Print)).
[84] Bates Lj Fau-Mulley JCandMulley JC. A BamHI RFLP at MT2A on human chromosome 16. (0305-1048 (Print)).
[85] Cipriano C Fau-Malavolta M, Malavolta M Fau-Costarelli L, Costarelli L Fau-Giacconi R, et al. Polymorphisms in MTla gene coding region are associated with longevity in Italian Central female population. (1389-5729 (Print)).
[86] 常秀丽，金泰廙，陈亮，等．金属硫蛋白基因亚型表达在镉接触中作为生物标志物的研究．中华劳动卫生职业病杂志2006；24(1)：12-15．
[87] 杨丽娜，李鸿雁，刘雅文，等．MT2A基因多态性与2型糖尿病的关系．吉林大学学报(医学版)2005；31(5)：758-760．
[88] 龚鹏飞，张国高．染矽尘大鼠肺泡巨噬细胞和肺组织热应激蛋白HSP70的表达．中华劳动卫生职业病杂志1995；13(3)：144-6．
[89] 蒋莹，马秀英，曲利明．肝纤维化组织中MMP1、TIMP1的表达及其临床意义．新疆医科大学学报2004；27(4)：372-4．
[90] 杨坤禹，刘莉，张涛，等．基质金属蛋白酶及其组织抑制物在小鼠肺纤维化发生中的作用．中华内科杂志2006；45(7)：589-90．
[91] 刘志峰，郝理华，张理菁．人巨细胞病毒感染对体外培养肺成纤维细胞中明胶酶的影响．中华微生物学和免疫学杂志2005；25(7)：529-32．
[92] Lim S, Roche N, Oliver BG, et al. Balance of matrix metalloprotease-9 and tissue inhibitor of metalloprotease-1 from alveolar macrophages in cigarette smokers. Regulation by interleukin-10. Am J Respir Crit Care Med 2000; 162 (4 Pt 1): 1355-60.
[93] Yasmin, McEniery CM, Wallace S, et al. Matrix metalloproteinase-9 (MMP-9), MMP-2, and serum elastase activity are associated with systolic hypertension and arterial stiffness. Arterioscler Thromb Vasc Biol 2005; 25 (2): 372.
[94] Russell RE, Culpitt SV, DeMatos C, et al. Release and activity of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 by alveolar macrophages from patients with chronic obstructive pulmonary disease. Am J Respir Cell Mol Biol 2002; 26 (5): 602-9.
[95] Salisbury BA, Pungliya M, Choi JY, et al. SNP and haplotype variation in the human genome. Mutat Res 2003; 526 (1-2): 53-61.
[96] Lee Th Fau-Kim S-U, Kim Su Fau-Yu S-L, Yu Sl Fau-Kim SH, et al. Peroxiredoxin Ⅱ is essential for sustaining life span of erythrocytes in mice. (0006-4971 (Print)).
[97] Rhee Sg Fau-Kang SW, Kang Sw Fau-Chang TS, Chang Ts Fau-Jeong W, et al. Peroxiredoxin, a novel family of peroxidases. (1521-6543 (Print)).
[98] Hung Cc Fau-Ichimura T, Ichimura T Fau-Stevens JL, Stevens Jl Fau-Bonventre JV, et al. Protection of renal epithelial cells against oxidative injury by endoplasmic reticulum stress preconditioning ismediated by ERK1/2 activation. (0021-9258 (Print)).
[99] Chang Ts Fau-Jeong W, Jeong W Fau-Choi SY, Choi Sy Fau-Yu S, et al. Regulation of peroxiredoxin Ⅰ activity by Cdc2-mediated phosphorylation. (0021-9258 (Print)).
[100] Dierick Jf Fau-Wenders F, Wenders F Fau-Chainiaux F, Chainiaux F Fau-Remacle J, et al. Retrovirally mediated overexpression of peroxiredoxin Ⅵ increases the survival of WI-38 human diploid fibroblasts exposed to cytotoxic doses of tert-butylhydroperoxide and UVB. (1389-5729 (Print)).
[101] Woo Ha Fau-Chae HZ, Chae Hz Fau-Hwang SC, Hwang Sc Fau-Yang K-S, et al. Reversing the inactivation of peroxiredoxins caused by cysteine sulfinic acid formation. (1095-9203 (Electronic)).
[102] Tien Nguyen-nhu N Fau-Knoops BandKnoops B. Mitochondrial and cytosolic expression of human peroxiredoxin 5 in Saccharomyces cerevisiae protect yeast cells from oxidative stress induced by paraquat. (0014-5793 (Print)).
[103] Seo Ms Fau-Kim JK, Kim Jk Fau-Lim Y, Lim Y Fau-Kang SW, et al. Rapid degradation of PrxⅠ and PrxⅡ induced by silica in Rat2 cells. (0006-291X (Print)).
[104] Michalak M Fau-Burns K, Bums K Fau-Andrin C, Andrin C Fau-Mesaeli N, et al. Endoplasmic reticulum form of calreticulin modulates glucocorticoid-sensitive gene expression. (0021-9258 (Print)).
[105] 徐菲菲，刘秀华．钙网蛋白的生理及病理生理学作用．生理科学进展 2006；37(03)：216-220．
[106] Gao B Fau-Adhikari R, Adhikari R Fau-Howarth M, Howarth M Fau-Nakamura K, et al. Assembly and antigen-presenting function of MHC class Ⅰ molecules in cells lacking the ER chaperone calreticulin. (1074-7613 (Print)).
[107] Li J Fau-Puceat M, Puceat M Fau-Perez-Terzic C, Perez-Terzic C Fau-Mery A, et al. Calreticulin reveals a critical Ca(2+) checkpoint-in-cardiac myofibritlogenesis. (0021-9525 (Print)).
[108] Mancino L Fau-Rizvi SM, Rizvi Sm Fau-Lapinski PE, Lapinski Pe Fau-Raghavan M, et al. Calreticulin recognizes misfolded HLA-A2 heavy chains. (0027-8424 (Print)).
[109] Papp S, Fadel MP, Kim H, et al. Calreticulin affects fibronectin-based cell-substratum adhesion via the regulation of C-SRC activity. (0021-9258 (Print)).
[110] Mesaeli N Fau-Phillipson CandPhillipson C. Impaired p53 expression, function, and nuclear localization in calreticulin-deficient cells. (1059-1524 (Print)).
[111] Casado P, Zuazua-Villar P, Prado MA, et al. Characterization of HSP27 phosphorylation induced by microtubule interfering agents: Implication of p38 signalling pathway Hsp27 overexpression in the R6/2 mouse model of Huntington's Disease: Chronic Neurodegeneration does not Induce Hsp27 Activation. (0003-9861 (Print)).
[112] Hastie ATandPeters SE Interactions of allergens and irritants in susceptible populations in producing lung dysfunction: implications for future research. Environ Health Perspect 2001 ; 109 Suppl 4: 605-7.
[113] Zee RY, Bates DandRidker PM. A prospective evaluation of the heat shock protein 70 gene polymorphisms and the risk of stroke. Thromb Haemost 2002; 87 (4): 622-5.
[114] Williams KJ, Cruikshank MKandPlopper CG. Pulmonary heat shock protein expression after exposure to a metabolically activated Clara cell toxicant: relationship to protein adduct formation. Toxicol Appl Pharmacol 2003; 192 (2): 107-18.
[115] Wang L Fau-Luhm R, Luhm R Fau-Lei MandLei M. SNP and mutation analysis. (0065-2598 (Print)).
[116] Hnizdo E Fau-Sluis-Cremer GKandSluis-Cremer GK. Risk of silicosis in a cohort of white South African gold miners. (0271-3586 (Print)).
[117] Muir Dc Fau-Julian JA, Julian Ja Fau-Shannon HS, Shannon Hs Fau-Verma DK, et al. Silica exposure and silicosis among Ontario hardrock miners: Ⅲ. Analysis and risk estimates. (0271-3586 (Print)).
[118] Chen W Fau-Zhuang Z, Zhuang Z Fau-Attfield MD, Attfield Md Fau-Chen BT, et al. Exposure to silica and silicosis among tin miners in China: exposure-response analyses and risk assessment.
[119] Brinkman GLandCoates EO, Jr. The effect of bronchitis, smoking, and occupation on ventilation. Am Rev Respir Dis 1963; 87: 684-93.
[120] 何权瀛，林江涛，主编．现代呼吸系统疾病诊断学．北京：中国协和医科大学出版社，2002．
[121] Pae CU, Kim TS, Kwon O J, et al. Polymorphisms of heat shock protein 70 gene (HSPA1A, HSPA1B and HSPA1L) and schizophrenia. Neurosci Res 2005; 53 (1): 8-13.
[122] Ding K, Zhou K, He F, et al. LDA—a java-based linkage disequilibrium analyzer. Bioinformatics 2003; 19 (16): 2147-8.
[123] 张华生，杨剑，覃英标，等．广西锡矿矿尘致矽肺的危险度评价．中华劳动卫生职业病杂志 2002；20(6)：430-432．
[124] Liu P, Sun MandSader S. Matrix metalloproteinases in cardiovascular disease. Can J Cardiol 2006; 22 Suppl B: 25B-30B.
[125] Pardo A, Perez-Ramos J, Segura-Valdez L, et al. Expression and localization of TIMP-1, TIMP-2, MMP-13, MMP-2, and MMP-9 in early and advanced experimental lung silicosis. Ann N YAcad Sci 1999; 878: 587-9.
[126] Lin S, Cutler DJ, Zwick ME, et al. Haplotype inference in random population samples. Am J Hum Genet 2002; 71 (5): 1129-37.
[127] Stephens M, Smith NJandDonnelly P. A new statistical method for haplotype reconstruction from population data. Am J Hum Genet 2001 ; 68 (4): 978-89.
[1] Muir DCF. Toxicity of silica dust [J]. Am J Ind Med, 1994, 25: 769-770.
[2] Parkes R. Occupational Lung Disorders [M]. 3ed. New York: Butterworth-Heinemann, 1994, 285-339.
[3] Wallace WE, Harrison J, Gryson RL, et al. Aluminosilicate surface contamination of respirable quartz particles from coal mine dusts and from clay works dusts [J]. Ann Occup Hyg, 1994, 38(suppl 1): 439-445.
[4] 甄铁梅，逯越，宋王华．Ⅰ期矽肺及0+肺通气功能的测定分析[J]．中国工业医学杂志，2001，14(3)：176，185．
[5] 魏剑琴，赫国志，孙桂珍，杨俊玲．国内、国外两种肺功能预测值结果分析[J]．白求恩医科大学学报，1994，20(6)：606-607．
[6] 郑永克，郑纪阳，戴新建，叶绍茗．0+期矽肺患者肺功能检查及其临床意义[J]．浙江中西医结合杂志，2004，14(9)：566-567．
[7] Gosset P, Lassalle P, Vanhee D, et al. Production of tumor necrosis factor-alpha and interleukin-6 by human alveolar macrophages exposed in vitro to coal mine dust [J]. AmJ Respir Cell Mol Biol, 1991, 5(5): 431-436.
[8] Piguet PF, Vesin C. Treatment by human recombinant soluble TNF receptor of pulmonary fibrosis induced by bleomycin or silica in mice [J]. Eur Respir J, 1994, 7(3): 515-518.
[9] Piguet PF, Vesin C, Grau GE, et al. Interleukin 1 receptor antagonist (IL-1ra) prevents or cures pulmonary fibrosis elicited in mice by bleomycin or silica [J]. Cytokine, 1993, 5(1): 57-61.
[10] 何平，李静，潘作东等．特发性肺纤维化患者外周血细胞产生IL-8能力的研究[J]．中国实用内科杂志，2002，22(6)：336—337．
[11] Hata J, Aoki K, Mitsuhashi H, et al. Change in location of cytokine- induced neutrophil chemoattractants (CINCs) in pulmonary silicosis [J]. Exp Mol Pathol, 2003, 75(1): 68-73.
[12] DriscollKE,HowardBW, Carter JM, et al. Alpha-quartz-induced chemokine expression by rat lung epithelial cells: effects of in vivo and in vitro particle exposure [J]. AmJ Pathol, 1996, 149(5): 1627-1637.
[13] Jagirdar J, Begin R, Dufresne A, et al. Transforming growth factor- beta(TGF- b) in silicosis [J]. Am J Respir Crit Care Med, 1996, 154(4 Pt 1): 1076-1081.
[14] Bonner JC, Goodell AL, Coin PG, et al. Brody. Chrysotile asbestos upregulates gene expression and production of alpha-receptors for plateletderived growth factor (PDGF-AA) onrat lung fibroblasts [J]. J Clin Invest, 1993, 92 (1): 425-430.
[15] Chen F, Deng HY, Ding GF, et al. Excessive production of insulin-like growth factor-Ⅰ by silicotic rat alveolar macrophages [J]. APMIS, 1994, 102 (8): 581-588.
[16] 吴浩，张月娥，许祖德等．大鼠肺纤维化血小板源性生长因子、血小板源性生长因子受体、转化生长因子-β、转化生长因子-β受体的表达[J]．中国组织化学与细胞化学杂志，2000，9(2)：157-160．
[17] Kumar RK, Velan GM, O' Grady R. Epidermal growth factor- like activity in bronchoalveolar lavage fluid in experimental silicosis [J]. Growth Factors, 1994, 10(3): 163-170.
[18] Shukla A, Timblin CR, Hubbard AK, et al. Silica-induced activation of c-Jun-NH2-terminal amino kinases, protracted expression of the activator protein-1 proto-oncogene, fra-1, and S-phase alterations are mediated via oxidative stress [J]. Cancer Res, 2001, 61(5): 1 791-1 795.
[19] Vallyathan V, Mega JF, Shi X, et al. Enhanced generation of free radicals from phagocytes induced by mineral dusts [J]. AmJ Respir Cell Mol Biol, 1992, 6(4): 404-413.
[20] Mossman BT, Churg A. Mechanisms in the Pathogenesis of Asbestosis and Silicosis [J]. AmJ Respir Crit Care Med, 1998, 157(5 Pt 1): 1 666-1 680.
[21] Halliwell B. Protection against tissue damage in vivo by desferrioxamine: what is its mechanism of action [J]. Free Radic Biol Med, 1989, 7(6): 645-651.
[22] Vallyathan V, Shi XL, Dalal NS, et al. Generation of free radicals from freshly fractured silica dust: potential role in acute silica-induced lung injury [J]. AmRev Respir Dis, 1988, 138(5):1 213-1 219.
[23] Blackford JA, Antonini JM, Castranova V, et al. Intratracheal instillation of silica up-regulates inducible nitric oxide synthase gene expression and increases nitric oxide production in alveolar macrophages and neutrophils [J]. AmJ Respir Cell Mol Biol, 1994, 11(4):426-431.
[24] Srivastava KD, RomWN, Jagirdar J, et al. Crucial role of interleukin-1 beta and nitric oxide synthase in silica-induced inflammation and apoptosis in mice [J]. AmJ Respir Crit Care Med, 2002, 165(4): 527-533.
[25] Cho YJ, SeoMS, KimJK, et al. Silica-induced generation of reactive oxygen species in Rat2 fibroblast: role in activation of mitogen-activated protein kinase [J]. BiochemBiophys Res Commun, 1999, 262(3): 708-712.
[26] Ding M, Shi X, Dong D, et al. Freshly fractured crystalline silica induce activator protein-1 activation through ERKs and p38 MAPK [J]. J Biol Chem, 1999, 274(43): 30611-30616.
[27] Shi X, Dalai NS, Vallyathan V. ESR evidence for the hydroxyl formation in aqueous suspension of quartz particles and its possible significance to lipid peroxidation in silicosis [J]. J Toxicol Environ Health, 1988, 25(2): 237-245.
[28] BorgesVM, LopesMF, FalcaoH, et al. Apoptosis underlies immunopathogenic mechanisms in acute silicosis [J]. AmJRespir CellMol Biol, 2002, 27(1): 78-84.
[29] Erin MC, Douglas PL, Berran Y, et al. Significance of genetic information in risk assessment and individual classification using silicosis as a case model [J]. Ann Occup Hyg, 2002, 46(4): 375-381.
[30] Koskinen H, Tiilikainen A, Nordman H. Increased prevalence of HLA-Aw19 and of the phenogroup Aw19, B18 in advanced silicosis [J]. Chest, 1983, 83(6):848-852.
[31] Rihs HP, Lipps P, May-Taube K, et al. Immunogenetic sudies on HLA-DR in German coal miners with and without coal worker's pneumoconiosis [J]. Lung, 1994, 172(6): 347-354.
[32] Yucesoy B, Vallyathan V, Landsittel DP, et al. Association of Tumor Necrosis Factor-α and Interleukin-1 Gene Polymorphisms with Silicosis [J]. Toxicol Appl Pharmacol, 2001, 172(1): 75-82.
[33] Corbett EL, Mozzato-Chamay N, Butterworth AE, et al. Polymorphisms in the tumor necrosis factor-alpha gene promoter may predispose to severe silicosis in black South African miners [J]. Am J Respir Crit Care Med, 2002, 165(5): 690-693.
[34] Mcmanus R, Wilson AG, Mansfield J, et al. TNF2, a polymorphism of the tumour necrosis-alpha gene promoter, is a component of the celiac disease major histocompatibility complex haplotype[J]. Eur J Immunol, 1996, 26(9): 2 113-2 118.
[35] Yucesoy B, Vallyathan V, Landsittel DP, et al. Polymorphisms of the IL-1 gene complex in coal miners with silicosis [J]. Am J Ind Med, 2001, 39(3): 286-291.
[36] 常筑平，姜乐，等．矽肺宁治疗尘肺38例临床疗效观察．职业卫生与病伤，1996，11(3)：189．
[37] 薛秀云．抗氧化剂对矽肺患者尿羟脯氨酸的影响．职业卫生与病伤，1997，12(1)：35．
[38] 何令媛，张群卫，等．绞股蓝对矽肺纤维化影响的研究．职业卫生与病伤， 1997，12(1)：41．
[39] 黄曙海，樊建康，梁德新，等．Lazaroid U-75412E试验性治疗大鼠矽肺研究。中国职业医学，1999，26(6)：11-13．
[40] 任国平，曾昭玉，张琪风，等．支气管肺泡灌洗术治疗大鼠矽肺的组织形态学观察．中国职业医学，2000，27(6)：18-20．
[41] 付颖．饮食调整对COPD缓解期营养不良患者的影响．国外医学呼吸系统分册，1994，(4)：205．
[42] 李华．矽肺患者住院情况分析．职业医学，1997，24(2)：30．
[43] 龚建新，孙晓武，贡小清．辅助性诊断尘肺方法的研究．中华劳动卫生职业病杂志，1995，13(4)：202．
[44] 杨增明，范雪云，汤化昌．葡萄糖酸锌治疗矽肺的疗效观察．中华劳动卫生职业病杂志，1997，15(1)：37-38．
[45] 徐永芳，吴开国，杨莉．脂质过氧化与矽肺及螺旋藻的抗氧化作用．中国职业医学，2000，27(3)：45-46．
[46] 马萱绒，甘振威，冯彪，等．维生素和蛋氨酸对矽肺鼠细胞免疫功能的影响．营养学报，1997，19(2)：224-226．
[27] 王万银，钱令嘉．HSP27研究现状．国外医学：生理病理科学与临床分册，2001：21(6)467-9．
[28] 胡晓荣．金属硫蛋白的生理作用及在医学上的应用．西昌农业高等专科学校学报2000；14(1)：47-50．
[29] 常秀丽．金属硫蛋白及其在镉接触评价中的应用．国外医学：卫生学分册2004；31(1)：32-6．
[30] 张桂春．金属硫蛋白的功能及应用前景．烟台师范学院学报：自然科学版2005；21(2)：142-5．
[31] 周敏，杨芳，贺智敏，等．人金属硫蛋白MT-2a的原核表达、纯化及其抗血清的制备．细胞与分子免疫学杂志2006；22(4)：530-2．
[32] Atkinson JJandSenior RM. Matrix metalloproteinase-9 in lung remodeling. Am J Respir Cell Mol Biol 2003; 28 (1): 12-24.
[33] Perez SE, Cano DA, Dao-Pick T, et al. Matrix metalloproteinases 2 and 9 are dispensable for pancreatic islet formation and function in vivo. Diabetes 2005; 54 (3): 694-701.
[34] 曾爱萍，曾水清，程扬，等．Modulaion of Matrix Metalloproteinase and TIMP-1 Expression by TGF-β1 in Cultured Human RPE Cells．华中科技大学学报：医学英德文版2006；26(3)：363-5．
[35] Nguyen M, Arkell JandJackson CJ. Active and tissue inhibitor of matrix metalloproteinase-free gelatinase B accumulates within human microvascular endothelial vesicles. J Biol Chem 1998; 273 (9): 5400-4.
[36] Qing H, Nie, Ya F, et al. Correlation between TIMP-1 expression and liver fibrosis in two rat liver fibrosis models．世界胃肠病学杂志：英文版2006；12(19)：3044-9．
[37] Feldman LJ, Mazighi M, Scheuble A, et al. Differential expression of matrix metalloproteinases after stent implantation and balloon angioplasty in the hypercholesterolemic rabbit. Circulation 2001; 103 (25): 3117-22.
[38] Douthwaite JA, Johnson TS, Haylor JL, et al. Effects of transforming growth factor-betal on renal extracellular matrix components and their regulating proteins. J Am Soc Nephrol 1999; 10 (10): 2109-19.
[39] Legallicier B, Trugnan G, Murphy G, et al. Expression of the type Ⅳ collagenase system during mouse kidney development and tubule segmentation. J Am Soc Nephrol 2001; 12 (11): 2358-69.
[40] Ma XB, Sun SXandYang F. [Effect of human silicotic alveolar macrophages on the expression of matrix metalloproteinase-1 and tissue inhibitor of metalloproteinases-1 in human lung fibroblasts.]. Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za Zhi 2004; 22 (5): 358-60.
[41] Braundmeier AG, Fazleabas AT, Lessey BA, et al. Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) Regulates Metalloproteinases in Human Uterine Endometrium. J Clin Endocrinol Metab 2006.
[42] Scabilloni JF, Wang L, Antonini JM, et al. Matrix metalloproteinase induction in fibrosis and fibrotic nodule formation due to silica inhalation. Am J Physiol Lung Cell Mol Physiol 2005; 288(4): L709-17.
[43] Li H, Cui D, Tong X, et al. [The role of matrix metalloproteinases in extracellular matrix remodelling in chronic obstructive pulmonary disease rat models]. Zhonghua Nei Ke Za Zhi 2002; 41(6): 393-8.
[44] Cheng Z, Winant RCandGambhir SS. A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. J Nucl Med 2005; 46 (5): 878-86.
[45] Westergren-Thorsson G, Malmstrom JandMarko-Varga G. Proteomics — the protein expression technology to study connective tissue biology. J Pharm Biomed Anal 2001; 24(5-6): 815-24.
[46] Zhou Y, Yu C, Miao X, et al. Functional haplotypes in the promoter of matrix metalloproteinase-2 and lung cancer susceptibility. Carcinogenesis June 2005; 26 (6): 1117-21.
[47] Ueki A, Isozaki Y, Tomokuni A, et al. Different distribution of HLA class II alleles in anti-topoisomerase I autoantibody responders between silicosis and systemic sclerosis patients, with a common distinct amino acid sequence in the HLA-DQB1 domain. Immunobiology 2001; 204(4): 458-65.
[48] Bretana A, Goihman-Yahr M, Tapia FJ, et al. Comparative ultrastructure and immunolabeling of MHC-II antigens of alveolar macrophages obtained from patientswith paracoccidioidomycosis and other lung diseases. J Leukoc Biol 1995; 57 (1): 101-9.
[49] Honda K, Kimura A, Dong RP, et al. Immunogenetic analysis of silicosis in Japan. Am J Respir Cell Mol Biol 1993; 8(1): 106-11.
[50] Migliaccio CT, Hamilton RF, Jr.andHolian A. Increase in a distinct pulmonary macrophage subset possessing an antigen-presenting cell phenotype and in vitro APC activity following silica exposure. Toxicol Appl Pharmacol 2005:205 (2): 168-76.
[51] Shanklin DRandSmalley DL. The immunopathology of siliconosis. History, clinical presentation, and relation to silicosis and the chemistry of silicon and silicone. Immunol Res 1998; 18 (3): 125-73.
[52] Zetterberg G, Elmberger G, Johansson A, et al. Rat alveolar and interstitial macrophages in the fibrosing stage following quartz exposure. Hum Exp Toxicol 2000; 19 (7): 402-11.
[53] Nadir R, Jedlicka A, Mintz M, et al. Effect of TNF and LTA polymorphisms on biological markers of response to oxidative stimuli in coal miners: a model of gene-environment interaction. Turnout necrosis factor and lymphotoxin alpha. J Med Genet2003; 40 (2): 96-103.
[54] Kim KA, Cho YY, Cho JS, et al. Tumor necrosis factor-alpha gene promoter polymorphism in coal workers' pneumoconiosis. Mol Cell Biochem 2002:234-235 (1-2): 205-9.
[55] Vanhee D, Gosset P, Marquette CH, et al. Secretion and mRNA expression of TNF alpha and IL-6 in the lungs of pneumoconiosis patients. Am J Respir Crit Care Med 1995; 152 (1): 298-306.
[56] Nadif R, Mintz M, Marzec J, et al. IL18 and IL18R1 potymorphisms, lung HRCT and fibrosis: a longitudinal study in coal miners. Eur Respir J 2006.
[57] Barbarin V, Xing Z, Delos M, et al. Pulmonary overexpression of IL-10 augments lung fibrosis and Th2 responses induced by silica particles. Am J Physiol Lung Cell Mol Physiol 2005; 288(5): L841-8.
[58] Lesur OJ, Mancini NM, Humbert JC, et al. Interleukin-6, interferon-gamma, and phospholipid levels in the alveolar lining fluid of human lungs. Profiles in coal worker's pneumoconiosis and idiopathic pulmonary fibrosis. Chest 1994:106 (2): 407-13.
[59] Lassalle P, Gosset P, Aerts C, et al. Abnormal secretion of interleukin-1 and tumor necrosis factor alpha by alveolar macrophages in coal worker's pneumoconiosis: comparison between simple pneumoconiosis and progressive massive fibrosis. Exp Lung Res 1990; 16(1): 73-80.
[60] 任鹏，吴秀珍，赵静，等．转化生长因子-β与肺纤维化．中国煤炭工业医学杂志，2006：9(6)：535-7．
[61] 朱剑萍，姚宏伟，陈季强．肺泡上皮细胞在肺纤维化中的作用及机制研究进展．国际呼吸杂志，2006；26(7)：506-9．
[62] Yao W, Wang ZM, Wang MZ, et al. [Oxidative injury and serum cytokines in coal workers with pneumoconiosis]. Sichuan Da Xue Xue Bao Yi Xue Ban 2005; 36 (4): 510-2.
[63] 张国高，贺涵贞．热应激蛋白及其在职业医学中应用研究的展望．中华劳动卫生职业病杂志1998：16(2)：67-9．
[64] Sawatzky DA, Foster R, Seed MP, et al. Heat-shock proteins and their role in chondrocyte protection, an application for autologous transplantation. Inflammopharmacology2005; 12 (5): 569-89.
[65] Barrow REandDasu MR. Oxidative and heat stress gene changes in hypertrophic scar fibroblasts stimulated with interleukin-1beta. J Surg Res 2005; 126 (1): 59-65.
[66] Torronteguy C, Frasson A, Zerwes F, et al. Inducible heat shock protein 70 expression as a potential predictive marker of metastasis in breast tumors. Cell Stress Chaperones 2006; 11 (1): 34-43.
[67] Simko M, Hartwig C, Lantow M, et al. Hsp70 expression and free radical release after exposure to non-thermal radio-frequency electromagnetic fields and ultrafine particles in human Mono Mac 6 cells. Toxicol Lett 2006; 161 (1): 73-82.
[68] Multhoff G. Heat shock proteins in immunity. Handb Exp Pharmacol 2006 (172): 279-304.
[69] Bogatu B Fau-Contag B, Contag B, Ishihara Y Fau-Nishikawa T, et al. Adsorption of lipoproteins onto mineral dust surfaces: a possible factor in the pathogenesis of particle-induced pulmonary fibrosis。Expression of matrix metalloproteinase, tissue inhibitors of metalloproteinase and adhesion molecules in silicotic mice with lung tumor metastasis. (0341-0382 (Print)).
[70] Cerdan S Fau-Rodrigues TB, Rodrigues Tb Fau-Sierra A, Sierra A Fau-Benito M, et al. The redox switch/redox coupling hypothesis[Significance of current bio- and molecular-markers in urological cancers] A review of the cytokine network in multiple myeloma: diagnostic, prognostic, and therapeutic implications. (0197-0186 (Print)).
[71] 何燕，邓国忠，高天．肺纤维化细胞因子机制研究进展．中华创伤杂志，2006；22(6)：477-80．
[72] Porter DW, Millecchia LL, Willard P, et al. Nitric Oxide and Reactive Oxygen Species Production Causes Progressive Damage in Rats after Cessation of Silica Inhalation. Toxicological Sciences March 2006; 90 (1): 188-97.
[73] Delgado L, Parra ERandCapelozzi VL. Apoptosis and extracellular matrix remodelling in human silicosis. Histopathology 2006; 49 (3): 283-9.
[74] Radons JandMulthoff G. Immunostimulatory functions of membrane-bound and exported heat shock protein 70. Exert Immunol Rev 2005; 11: 17-33.
[75] Melchers I. [Immunogenetics--HLA-association, molecular chaperones and "related" diseases]. Z Rheumatol 2005; 64 (6): 402-7.
[76] van Eden W. Immunoregulation of autoimmune diseases. Hum Immunol 2006; 67 (6): 446-53.
[77] Copland IBandPost M. Stretch-activated signaling pathways responsible for early response gene expression in fetal lung epithelial cells. J Cell Physiol 2006; 210 (1): 133-43.
[78] Ito H, Sekimura K, Sasaki N, et al. [Cytoprotective effects of heat shock protein-70 in bronchial epithelium against neutrophil elastase-induced cell injury]. Arerugi 2006; 55 (7): 820-6.
[79] 赵京山，尹桂山．金属硫蛋白的生理功能及其研究进展．生命的化学1998；18(3)：9-11．
[80] 刘志勇，魏国林．金属硫蛋白研究进展．江西科学2004；22(2)：104-9．
[81] 李守明，刘志勇．金属硫蛋白与疾病关系．江西医药2004；39(6)：434-7．
[82] 安建平，王海娟，韩沛平，等．属硫蛋白生理功能的研究进展．天水师范学院学报2005；25 (2)：34-8．
[83] Yang F Fau-Zhou M, Zhou M Fau-He Z, He Z Fau-Liu X, et al. High-yield expression in Escherichia coli of soluble human MT2A with native functions A BamHI RFLP at MT2A on human chromosome 16. (1046-5928 (Print)).
[84] Bates Lj Fau-Mulley JCandMulley JC. A BamHI RFLP at MT2A on human chromosome 16. (0305-1048 (Print)).
[85] Cipriano C Fau-Malavolta M, Malavolta M Fau-Costarelli L, Costarelli L Fau-Giacconi R, et al. Polymorphisms in MTla gene coding region are associated with longevity in Italian Central female population. (1389-5729 (Print)).
[86] 常秀丽，金泰廙，陈亮，等．金属硫蛋白基因亚型表达在镉接触中作为生物标志物的研究．中华劳动卫生职业病杂志2006；24(1)：12-15．
[87] 杨丽娜，李鸿雁，刘雅文，等．MT2A基因多态性与2型糖尿病的关系．吉林大学学报(医学版)2005；31(5)：758-760．
[88] 龚鹏飞，张国高．染矽尘大鼠肺泡巨噬细胞和肺组织热应激蛋白HSP70的表达．中华劳动卫生职业病杂志1995；13(3)：144-6．
[89] 蒋莹，马秀英，曲利明．肝纤维化组织中MMP1、TIMP1的表达及其临床意义．新疆医科大学学报2004；27(4)：372-4．
[90] 杨坤禹，刘莉，张涛，等．基质金属蛋白酶及其组织抑制物在小鼠肺纤维化发生中的作用．中华内科杂志2006；45(7)：589-90．
[91] 刘志峰，郝理华，张理菁．人巨细胞病毒感染对体外培养肺成纤维细胞中明胶酶的影响．中华微生物学和免疫学杂志2005；25(7)：529-32．
[92] Lim S, Roche N, Oliver BG, et al. Balance of matrix metalloprotease-9 and tissue inhibitor of metalloprotease-1 from alveolar macrophages in cigarette smokers. Regulation by interleukin-10. Am J Respir Crit Care Med 2000; 162 (4 Pt 1): 1355-60.
[93] Yasmin, McEniery CM, Wallace S, et al. Matrix metalloproteinase-9 (MMP-9), MMP-2, and serum elastase activity are associated with systolic hypertension and arterial stiffness. Arterioscler Thromb Vasc Biol 2005; 25 (2): 372.
[94] Russell RE, Culpitt SV, DeMatos C, et al. Release and activity of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 by alveolar macrophages from patients with chronic obstructive pulmonary disease. Am J Respir Cell Mol Biol 2002; 26 (5): 602-9.
[95] Salisbury BA, Pungliya M, Choi JY, et al. SNP and haplotype variation in the human genome. Mutat Res 2003; 526 (1-2): 53-61.
[96] Lee Th Fau-Kim S-U, Kim Su Fau-Yu S-L, Yu Sl Fau-Kim SH, et al. Peroxiredoxin Ⅱ is essential for sustaining life span of erythrocytes in mice. (0006-4971 (Print)).
[97] Rhee Sg Fau-Kang SW, Kang Sw Fau-Chang TS, Chang Ts Fau-Jeong W, et al. Peroxiredoxin, a novel family of peroxidases. (1521-6543 (Print)).
[98] Hung Cc Fau-Ichimura T, Ichimura T Fau-Stevens JL, Stevens Jl Fau-Bonventre JV, et al. Protection of renal epithelial cells against oxidative injury by endoplasmic reticulum stress preconditioning ismediated by ERK1/2 activation. (0021-9258 (Print)).
[99] Chang Ts Fau-Jeong W, Jeong W Fau-Choi SY, Choi Sy Fau-Yu S, et al. Regulation of peroxiredoxin Ⅰ activity by Cdc2-mediated phosphorylation. (0021-9258 (Print)).
[100] Dierick Jf Fau-Wenders F, Wenders F Fau-Chainiaux F, Chainiaux F Fau-Remacle J, et al. Retrovirally mediated overexpression of peroxiredoxin Ⅵ increases the survival of WI-38 human diploid fibroblasts exposed to cytotoxic doses of tert-butylhydroperoxide and UVB. (1389-5729 (Print)).
[101] Woo Ha Fau-Chae HZ, Chae Hz Fau-Hwang SC, Hwang Sc Fau-Yang K-S, et al. Reversing the inactivation of peroxiredoxins caused by cysteine sulfinic acid formation. (1095-9203 (Electronic)).
[102] Tien Nguyen-nhu N Fau-Knoops BandKnoops B. Mitochondrial and cytosolic expression of human peroxiredoxin 5 in Saccharomyces cerevisiae protect yeast cells from oxidative stress induced by paraquat. (0014-5793 (Print)).
[103] Seo Ms Fau-Kim JK, Kim Jk Fau-Lim Y, Lim Y Fau-Kang SW, et al. Rapid degradation of PrxⅠ and PrxⅡ induced by silica in Rat2 cells. (0006-291X (Print)).
[104] Michalak M Fau-Burns K, Bums K Fau-Andrin C, Andrin C Fau-Mesaeli N, et al. Endoplasmic reticulum form of calreticulin modulates glucocorticoid-sensitive gene expression. (0021-9258 (Print)).
[105] 徐菲菲，刘秀华．钙网蛋白的生理及病理生理学作用．生理科学进展 2006；37(03)：216-220．
[106] Gao B Fau-Adhikari R, Adhikari R Fau-Howarth M, Howarth M Fau-Nakamura K, et al. Assembly and antigen-presenting function of MHC class Ⅰ molecules in cells lacking the ER chaperone calreticulin. (1074-7613 (Print)).
[107] Li J Fau-Puceat M, Puceat M Fau-Perez-Terzic C, Perez-Terzic C Fau-Mery A, et al. Calreticulin reveals a critical Ca(2+) checkpoint-in-cardiac myofibritlogenesis. (0021-9525 (Print)).
[108] Mancino L Fau-Rizvi SM, Rizvi Sm Fau-Lapinski PE, Lapinski Pe Fau-Raghavan M, et al. Calreticulin recognizes misfolded HLA-A2 heavy chains. (0027-8424 (Print)).
[109] Papp S, Fadel MP, Kim H, et al. Calreticulin affects fibronectin-based cell-substratum adhesion via the regulation of C-SRC activity. (0021-9258 (Print)).
[110] Mesaeli N Fau-Phillipson CandPhillipson C. Impaired p53 expression, function, and nuclear localization in calreticulin-deficient cells. (1059-1524 (Print)).
[111] Casado P, Zuazua-Villar P, Prado MA, et al. Characterization of HSP27 phosphorylation induced by microtubule interfering agents: Implication of p38 signalling pathway Hsp27 overexpression in the R6/2 mouse model of Huntington's Disease: Chronic Neurodegeneration does not Induce Hsp27 Activation. (0003-9861 (Print)).
[112] Hastie ATandPeters SE Interactions of allergens and irritants in susceptible populations in producing lung dysfunction: implications for future research. Environ Health Perspect 2001 ; 109 Suppl 4: 605-7.
[113] Zee RY, Bates DandRidker PM. A prospective evaluation of the heat shock protein 70 gene polymorphisms and the risk of stroke. Thromb Haemost 2002; 87 (4): 622-5.
[114] Williams KJ, Cruikshank MKandPlopper CG. Pulmonary heat shock protein expression after exposure to a metabolically activated Clara cell toxicant: relationship to protein adduct formation. Toxicol Appl Pharmacol 2003; 192 (2): 107-18.
[115] Wang L Fau-Luhm R, Luhm R Fau-Lei MandLei M. SNP and mutation analysis. (0065-2598 (Print)).
[116] Hnizdo E Fau-Sluis-Cremer GKandSluis-Cremer GK. Risk of silicosis in a cohort of white South African gold miners. (0271-3586 (Print)).
[117] Muir Dc Fau-Julian JA, Julian Ja Fau-Shannon HS, Shannon Hs Fau-Verma DK, et al. Silica exposure and silicosis among Ontario hardrock miners: Ⅲ. Analysis and risk estimates. (0271-3586 (Print)).
[118] Chen W Fau-Zhuang Z, Zhuang Z Fau-Attfield MD, Attfield Md Fau-Chen BT, et al. Exposure to silica and silicosis among tin miners in China: exposure-response analyses and risk assessment.
[119] Brinkman GLandCoates EO, Jr. The effect of bronchitis, smoking, and occupation on ventilation. Am Rev Respir Dis 1963; 87: 684-93.
[120] 何权瀛，林江涛，主编．现代呼吸系统疾病诊断学．北京：中国协和医科大学出版社，2002．
[121] Pae CU, Kim TS, Kwon O J, et al. Polymorphisms of heat shock protein 70 gene (HSPA1A, HSPA1B and HSPA1L) and schizophrenia. Neurosci Res 2005; 53 (1): 8-13.
[122] Ding K, Zhou K, He F, et al. LDA—a java-based linkage disequilibrium analyzer. Bioinformatics 2003; 19 (16): 2147-8.
[123] 张华生，杨剑，覃英标，等．广西锡矿矿尘致矽肺的危险度评价．中华劳动卫生职业病杂志 2002；20(6)：430-432．
[124] Liu P, Sun MandSader S. Matrix metalloproteinases in cardiovascular disease. Can J Cardiol 2006; 22 Suppl B: 25B-30B.
[125] Pardo A, Perez-Ramos J, Segura-Valdez L, et al. Expression and localization of TIMP-1, TIMP-2, MMP-13, MMP-2, and MMP-9 in early and advanced experimental lung silicosis. Ann N YAcad Sci 1999; 878: 587-9.
[126] Lin S, Cutler DJ, Zwick ME, et al. Haplotype inference in random population samples. Am J Hum Genet 2002; 71 (5): 1129-37.
[127] Stephens M, Smith NJandDonnelly P. A new statistical method for haplotype reconstruction from population data. Am J Hum Genet 2001 ; 68 (4): 978-89.
[1] Muir DCF. Toxicity of silica dust [J]. Am J Ind Med, 1994, 25: 769-770.
[2] Parkes R. Occupational Lung Disorders [M]. 3ed. New York: Butterworth-Heinemann, 1994, 285-339.
[3] Wallace WE, Harrison J, Gryson RL, et al. Aluminosilicate surface contamination of respirable quartz particles from coal mine dusts and from clay works dusts [J]. Ann Occup Hyg, 1994, 38(suppl 1): 439-445.
[4] 甄铁梅，逯越，宋王华．Ⅰ期矽肺及0+肺通气功能的测定分析[J]．中国工业医学杂志，2001，14(3)：176，185．
[5] 魏剑琴，赫国志，孙桂珍，杨俊玲．国内、国外两种肺功能预测值结果分析[J]．白求恩医科大学学报，1994，20(6)：606-607．
[6] 郑永克，郑纪阳，戴新建，叶绍茗．0+期矽肺患者肺功能检查及其临床意义[J]．浙江中西医结合杂志，2004，14(9)：566-567．
[7] Gosset P, Lassalle P, Vanhee D, et al. Production of tumor necrosis factor-alpha and interleukin-6 by human alveolar macrophages exposed in vitro to coal mine dust [J]. AmJ Respir Cell Mol Biol, 1991, 5(5): 431-436.
[8] Piguet PF, Vesin C. Treatment by human recombinant soluble TNF receptor of pulmonary fibrosis induced by bleomycin or silica in mice [J]. Eur Respir J, 1994, 7(3): 515-518.
[9] Piguet PF, Vesin C, Grau GE, et al. Interleukin 1 receptor antagonist (IL-1ra) prevents or cures pulmonary fibrosis elicited in mice by bleomycin or silica [J]. Cytokine, 1993, 5(1): 57-61.
[10] 何平，李静，潘作东等．特发性肺纤维化患者外周血细胞产生IL-8能力的研究[J]．中国实用内科杂志，2002，22(6)：336—337．
[11] Hata J, Aoki K, Mitsuhashi H, et al. Change in location of cytokine- induced neutrophil chemoattractants (CINCs) in pulmonary silicosis [J]. Exp Mol Pathol, 2003, 75(1): 68-73.
[12] DriscollKE,HowardBW, Carter JM, et al. Alpha-quartz-induced chemokine expression by rat lung epithelial cells: effects of in vivo and in vitro particle exposure [J]. AmJ Pathol, 1996, 149(5): 1627-1637.
[13] Jagirdar J, Begin R, Dufresne A, et al. Transforming growth factor- beta(TGF- b) in silicosis [J]. Am J Respir Crit Care Med, 1996, 154(4 Pt 1): 1076-1081.
[14] Bonner JC, Goodell AL, Coin PG, et al. Brody. Chrysotile asbestos upregulates gene expression and production of alpha-receptors for plateletderived growth factor (PDGF-AA) onrat lung fibroblasts [J]. J Clin Invest, 1993, 92 (1): 425-430.
[15] Chen F, Deng HY, Ding GF, et al. Excessive production of insulin-like growth factor-Ⅰ by silicotic rat alveolar macrophages [J]. APMIS, 1994, 102 (8): 581-588.
[16] 吴浩，张月娥，许祖德等．大鼠肺纤维化血小板源性生长因子、血小板源性生长因子受体、转化生长因子-β、转化生长因子-β受体的表达[J]．中国组织化学与细胞化学杂志，2000，9(2)：157-160．
[17] Kumar RK, Velan GM, O' Grady R. Epidermal growth factor- like activity in bronchoalveolar lavage fluid in experimental silicosis [J]. Growth Factors, 1994, 10(3): 163-170.
[18] Shukla A, Timblin CR, Hubbard AK, et al. Silica-induced activation of c-Jun-NH2-terminal amino kinases, protracted expression of the activator protein-1 proto-oncogene, fra-1, and S-phase alterations are mediated via oxidative stress [J]. Cancer Res, 2001, 61(5): 1 791-1 795.
[19] Vallyathan V, Mega JF, Shi X, et al. Enhanced generation of free radicals from phagocytes induced by mineral dusts [J]. AmJ Respir Cell Mol Biol, 1992, 6(4): 404-413.
[20] Mossman BT, Churg A. Mechanisms in the Pathogenesis of Asbestosis and Silicosis [J]. AmJ Respir Crit Care Med, 1998, 157(5 Pt 1): 1 666-1 680.
[21] Halliwell B. Protection against tissue damage in vivo by desferrioxamine: what is its mechanism of action [J]. Free Radic Biol Med, 1989, 7(6): 645-651.
[22] Vallyathan V, Shi XL, Dalal NS, et al. Generation of free radicals from freshly fractured silica dust: potential role in acute silica-induced lung injury [J]. AmRev Respir Dis, 1988, 138(5):1 213-1 219.
[23] Blackford JA, Antonini JM, Castranova V, et al. Intratracheal instillation of silica up-regulates inducible nitric oxide synthase gene expression and increases nitric oxide production in alveolar macrophages and neutrophils [J]. AmJ Respir Cell Mol Biol, 1994, 11(4):426-431.
[24] Srivastava KD, RomWN, Jagirdar J, et al. Crucial role of interleukin-1 beta and nitric oxide synthase in silica-induced inflammation and apoptosis in mice [J]. AmJ Respir Crit Care Med, 2002, 165(4): 527-533.
[25] Cho YJ, SeoMS, KimJK, et al. Silica-induced generation of reactive oxygen species in Rat2 fibroblast: role in activation of mitogen-activated protein kinase [J]. BiochemBiophys Res Commun, 1999, 262(3): 708-712.
[26] Ding M, Shi X, Dong D, et al. Freshly fractured crystalline silica induce activator protein-1 activation through ERKs and p38 MAPK [J]. J Biol Chem, 1999, 274(43): 30611-30616.
[27] Shi X, Dalai NS, Vallyathan V. ESR evidence for the hydroxyl formation in aqueous suspension of quartz particles and its possible significance to lipid peroxidation in silicosis [J]. J Toxicol Environ Health, 1988, 25(2): 237-245.
[28] BorgesVM, LopesMF, FalcaoH, et al. Apoptosis underlies immunopathogenic mechanisms in acute silicosis [J]. AmJRespir CellMol Biol, 2002, 27(1): 78-84.
[29] Erin MC, Douglas PL, Berran Y, et al. Significance of genetic information in risk assessment and individual classification using silicosis as a case model [J]. Ann Occup Hyg, 2002, 46(4): 375-381.
[30] Koskinen H, Tiilikainen A, Nordman H. Increased prevalence of HLA-Aw19 and of the phenogroup Aw19, B18 in advanced silicosis [J]. Chest, 1983, 83(6):848-852.
[31] Rihs HP, Lipps P, May-Taube K, et al. Immunogenetic sudies on HLA-DR in German coal miners with and without coal worker's pneumoconiosis [J]. Lung, 1994, 172(6): 347-354.
[32] Yucesoy B, Vallyathan V, Landsittel DP, et al. Association of Tumor Necrosis Factor-α and Interleukin-1 Gene Polymorphisms with Silicosis [J]. Toxicol Appl Pharmacol, 2001, 172(1): 75-82.
[33] Corbett EL, Mozzato-Chamay N, Butterworth AE, et al. Polymorphisms in the tumor necrosis factor-alpha gene promoter may predispose to severe silicosis in black South African miners [J]. Am J Respir Crit Care Med, 2002, 165(5): 690-693.
[34] Mcmanus R, Wilson AG, Mansfield J, et al. TNF2, a polymorphism of the tumour necrosis-alpha gene promoter, is a component of the celiac disease major histocompatibility complex haplotype[J]. Eur J Immunol, 1996, 26(9): 2 113-2 118.
[35] Yucesoy B, Vallyathan V, Landsittel DP, et al. Polymorphisms of the IL-1 gene complex in coal miners with silicosis [J]. Am J Ind Med, 2001, 39(3): 286-291.
[36] 常筑平，姜乐，等．矽肺宁治疗尘肺38例临床疗效观察．职业卫生与病伤，1996，11(3)：189．
[37] 薛秀云．抗氧化剂对矽肺患者尿羟脯氨酸的影响．职业卫生与病伤，1997，12(1)：35．
[38] 何令媛，张群卫，等．绞股蓝对矽肺纤维化影响的研究．职业卫生与病伤， 1997，12(1)：41．
[39] 黄曙海，樊建康，梁德新，等．Lazaroid U-75412E试验性治疗大鼠矽肺研究。中国职业医学，1999，26(6)：11-13．
[40] 任国平，曾昭玉，张琪风，等．支气管肺泡灌洗术治疗大鼠矽肺的组织形态学观察．中国职业医学，2000，27(6)：18-20．
[41] 付颖．饮食调整对COPD缓解期营养不良患者的影响．国外医学呼吸系统分册，1994，(4)：205．
[42] 李华．矽肺患者住院情况分析．职业医学，1997，24(2)：30．
[43] 龚建新，孙晓武，贡小清．辅助性诊断尘肺方法的研究．中华劳动卫生职业病杂志，1995，13(4)：202．
[44] 杨增明，范雪云，汤化昌．葡萄糖酸锌治疗矽肺的疗效观察．中华劳动卫生职业病杂志，1997，15(1)：37-38．
[45] 徐永芳，吴开国，杨莉．脂质过氧化与矽肺及螺旋藻的抗氧化作用．中国职业医学，2000，27(3)：45-46．
[46] 马萱绒，甘振威，冯彪，等．维生素和蛋氨酸对矽肺鼠细胞免疫功能的影响．营养学报，1997，19(2)：224-226．
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