Gasoline engine exhaust (GEE) has been reported to contribute to the pathogenesis of pulmonary diseases. Autophagy, proinflammatory cytokines, and the NF-κB pathway core protein may play roles in the development of lung diseases caused by GEE. However, little is known about the possible toxic effects. Herein, we aimed to examine the crosstalk between GEE and the expression levels of autophagy-associated proteins (microtubule-associated proteins 1A/1B light chain 3A (LC3I/II)), proinflammatory cytokine genes (including interleukin-1β (IL-1β), IL-6 and IL-8), and the NF-kB pathway core protein p65 by conducting an air–liquid interface exposure study in BEAS-2B cells. A CCK-8 assay was conducted to explore the viability of BEAS-2B cells exposed to GEE and 3-methyladenine (3-MA). The protein expression levels of LC3I/II and p65 were detected using Western blotting. The gene expression levels of LC3B, IL-1β, IL-6, and IL-8 were measured using real-time PCR. We found that GEE decreased the viability of BEAS-2B cells in a dose-dependent manner, whereas 10%GEE exposure and 2.5 mM 3-MA had no significant effect. As the dose of GEE increased, LC3I/II protein and gene expression levels, proinflammatory cytokine gene expression levels, and p65 protein expression levels showed varying degrees of changes. Additionally, after treatment with 3-MA, these indicators tended to decrease, but only the gene expression levels of proinflammatory cytokines were statistically significant. These results suggest that GEE could interfere with autophagy and induce an inflammatory response in human bronchial epithelial cells, and that modest changes in autophagy could significantly alleviate this response, thereby providing new insights for the understanding of lung injury caused by GEE.

OBJECTIVE:This study aimed to investigate the role of the nuclear factor erythroid 2-related factor 2/heme oxygenase-1 (Nrf2/HO-1) signaling cascade in the inflammatory responses induced by whole gasoline engine exhaust (GEE) in lung epithelial cells via air-liquid interface (ALI) exposure. MATERIALS AND METHODS:Using an ALI exposure system, human bronchial epithelial cells (BEAS-2B) and type II alveolar epithelial cells (A549) were exposed to whole GEE collected from a two-wheeled motorcycle at various dilution ratios. After a 1 h exposure at 10 mL/min, cell relative viability, intracellular reactive oxygen species (ROS), glutathione (GSH), oxidized glutathione (GSSG) and the GSH/GSSG ratio were measured. Inflammatory cytokines (IL-1β, IL-6, and IL-8) were quantified. The Nrf2 inhibitor brusatol (BR, 300 nM) and the antioxidant N-acetyl-L-cysteine (NAC, 5 mM) were used to modulate the Nrf2/HO-1 pathway and oxidative stress, respectively. Protein and gene expression levels were analyzed by Western Blotting and real-time PCR. RESULTS:Exposure to 10%GEE induced oxidative stress and optimally activated Nrf2/HO-1 expression without cytotoxicity, while higher concentrations suppressed this signaling pathway. Significant correlations were observed between Nrf2/HO-1 levels and inflammatory cytokines. Inhibition of Nrf2/HO-1 with BR reduced inflammatory responses which induced by the 10%GEE in both BEAS-2B and A549 cell lines. Furthermore, attenuating oxidative stress with NAC inhibited both Nrf2/HO-1 expression and the GEE-induced inflammatory response. CONCLUSION:Inhibiting the Nrf2/HO-1 signaling cascade attenuates the pro-inflammatory response induced by GEE in lung epithelial cells following ALI exposure. The Nrf2/HO-1 pathway appears to be a critical regulator of GEE-induced pulmonary inflammation, highlighting its potential as a therapeutic target.

Whole-component gasoline engine exhaust (GEE) has been classified as possibly carcinogenic to humans, where DNA and chromosome damage may play a key role. This study evaluated DNA and chromosome damage induced by GEE in human bronchial epithelial BEAS-2B cells using an air-liquid interface (ALI) exposure system. Following exposure to GEE at different dilution ratios, the cell relative viability (CRV), the percentage of DNA in the comet tail (TailDNA%), γ-H2AX protein expression, and chromosome damage for BEAS-2B cells were assessed using the Cell Counting Kit-8 (CCK-8) assay, the alkaline comet assay, Western Blotting, and cytokinesis-block micronucleus (CBMN) assay, respectively. Results showed that the relative survival rate of BEAS-2B cells decreased progressively with increasing GEE concentration (decreasing dilution ratios); specifically, a significant reduction was observed from the 1:10 dilution group onwards. TailDNA% increased significantly in all GEE-exposure groups compared to the clean air control, with a significant difference were observed starting from the 1:10 dilution group. γ-H2AX protein expression exhibited a nonsignificant trend of initial increase followed by a decrease. The cell nuclear division index (NDI) decreased significantly from the 1:5 dilution group onwards. The rates of micronuclei (MN), nucleoplasmic bridges (NPBs), and nuclear buds (NBUDs) increased significantly starting from the 1:10 dilution group, nondiluted GEE group, and 1:20 GEE group, respectively. These findings indicate that GEE exposure induces DNA and chromosome damage in BEAS-2B cells, and γ-H2AX may play a crucial role in DNA repair processes, although the specific mechanisms still require further investigation.

Abstract Introduction Exposure to diesel engine exhaust (DEE) is associated with increased lung cancer risk; however, underlying molecular mechanisms remain unclear. We apply an exposome approach to characterize early biological effects of occupational DEE exposure. Methods Plasma samples from 54 diesel engine factory workers and 55 non-exposed control workers were characterized using an integrated exposome platform that combines untargeted gas chromatography (GC-) and liquid chromatography (LC-) with high-resolution mass spectrometry (HRMS). Exposome profiles were evaluated by metabolome-wide association study (MWAS) for molecular features associated with DEE exposure and elemental carbon. Potential molecular mechanisms underlying DEE were further evaluated by integrating exposome profiles with plasma proteomics, urine aminopyrenes and mutagenicity, and buccal gene expression analysis. Results GC- and LC-HRMS untargeted analysis detected 68,285 metabolic features. Comparison of DEE-exposed and non-exposed workers identified 772 molecular features associated with exposure at a FDR <5%, including 102 detected using GC-HRMS and 670 detected using LC-HRMS. Molecular networking and annotation identified compounds consistent with DEE exposure, while metabolic pathway enrichment suggest alterations in oxidative stress and endothelial pathways. We conducted a secondary MWAS to link urinary mutagenicity, reflecting systemic exposure to genotoxic/carcinogenic agents, and associated with tumor development and identified 90 molecular features positively associated with urine mutagenicity at FDR<5%. Discussion Integration of exposome profiles with protein and genome-wide gene expression identified biological alterations consistent with many of the key characteristics of carcinogens. Conclusion Integrated exposome characterization of DEE exposure identified novel DEE biomarkers and biological response profiles in a high exposure setting.

Diesel exhaust (DE) is an important pollution source widely existing in the living and production environment, which is closely related to the health of the public and occupational groups. The International Agency for Research on Cancer has classified DE as a Group 1 carcinogen. Considering the negative health impacts on the respiratory system due to DE exposure in vitro, it is crucial to apply reliable test systems allowing accurate assessment of the biological effects of DE. The exposure technology of respiratory system in vitro is considered as one of the feasible measures to implement the 3R (reduce, refine, and replace) principle in animal experiments. Compared with the traditional submerged culture in vitro models, the air-liquid interface (ALI) exposure technology has the advantages including fewer influencing factors, easier exposure condition control, and shorter exposure cycle. ALI has become an important tool to study molecular events associated with physiology and pathology of respiratory system, and action modes and interactions of different cell types. Also, ALI has been increasingly widely used because it can simulate the actual processes of human respiratory system cells and/or tissues to DE exposure. This review was intended to introduce the development and advantages of ALI exposure technology, and further summarized the application progress of ALI exposure technology in studying the respiratory toxicity induced by DE exposure in vitro, so as to provide new ideas and pathways for the use of ALI exposure technology in the study of biomarkers and mechanisms of respiratory toxicity associated with DE exposure, and provide basic data to screen and promote biomarkers for exposed populations.

Diesel engine exhaust (DEE) is an established lung carcinogen, but the biological mechanisms of diesel‐induced lung carcinogenesis are not well understood. MicroRNAs (miRNAs) are small noncoding RNAs that play a potentially important role in regulating gene expression related to lung cancer. We conducted a cross‐sectional molecular epidemiology study to evaluate whether serum levels of miRNAs are altered in healthy workers occupationally exposed to DEE compared to unexposed controls. We conducted a two‐stage study, first measuring 405 miRNAs in a pilot study of six DEE‐exposed workers exposed and six controls. In the second stage, 44 selected miRNAs were measured using the Fireplex circulating miRNA assay that profiles miRNAs directly from biofluids of 45 workers exposed to a range of DEE (Elemental Carbon (EC), median, range: 47.7, 6.1–79.7 μg/m3) and 46 controls. The relationship between exposure to DEE and EC with miRNA levels was analyzed using linear regression adjusted for potential confounders. Serum levels of four miRNAs were significantly lower (miR‐191‐5p, miR‐93‐5p, miR‐423‐3p, miR‐122‐5p) and one miRNA was significantly higher (miR‐92a‐3p) in DEE exposed workers compared to controls. Of these miRNAs, miR‐191‐5p (ptrend = .001, FDR = 0.04) and miR‐93‐5p (ptrend = .009, FDR = 0.18) showed evidence of an inverse exposure–response with increasing EC levels. Our findings suggest that occupational exposure to DEE may affect circulating miRNAs implicated in biological processes related to carcinogenesis, including immune function.

Diesel exhaust has long been of health concern due to established toxicity including carcinogenicity in humans. However, the precise components of diesel engine emissions that drive carcinogenesis are still unclear. Limited work has suggested that nitrated polycyclic aromatic hydrocarbons (NPAHs) such as 1-nitropyrene and 2-nitro-fluorene may be more abundant in diesel exhaust. The present study aimed to examine whether urinary amino metabolites of these NPAHs were associated with high levels of diesel engine emissions and urinary mutagenicity in a group of highly exposed workers including both smokers and nonsmokers. Spot urine samples were collected immediately following a standard work shift from each of the 54 diesel engine testers and 55 non-tester controls for the analysis of five amino metabolites of NPAHs, and cotinine (a biomarker of tobacco smoke exposure) using liquid chromatography-mass spectrometry. An overnight urine sample was collected in a subgroup of non -smoking participants for mutagenicity analysis using strain YG1041 in the Salmonella (Ames) mutagenicity assay. Personal exposure to fine particles (PM2.5) and more-diesel-specific constituents (elemental carbon and soot) was assessed for the engine testers by measuring breathing-zone concentrations repeatedly over several full work shifts. Results showed that it was 12.8 times more likely to detect 1-aminopyrene and 2.9 times more likely to detect 2-aminofluorene in the engine testers than in unexposed controls. Urinary concentrations of 1-amino-pyrene were significantly higher in engine testers (p < 0.001), and strongly correlated with soot and elemental carbon exposure as well as mutagenicity tested in strain YG1041 with metabolic activation (p < 0.001). Smoking did not affect 1-aminopyrene concentrations and 1-aminopyrene relationships with diesel exposure. In contrast, both engine emissions and smoking affected 2-aminofluorene concentrations. The results confirm that urinary 1-aminopyrene may serve as an exposure biomarker for diesel engine emissions and associated mutagenicity.

Background We previously found that occupational exposure to diesel engine exhaust (DEE) was associated with alterations to 19 biomarkers that potentially reflect the mechanisms of carcinogenesis. Whether DEE is associated with biological alterations at concentrations under existing or recommended occupational exposure limits (OELs) is unclear. Methods In a cross-sectional study of 54 factory workers exposed long-term to DEE and 55 unexposed controls, we reanalysed the 19 previously identified biomarkers. Multivariable linear regression was used to compare biomarker levels between DEE-exposed versus unexposed subjects and to assess elemental carbon (EC) exposure-response relationships, adjusted for age and smoking status. We analysed each biomarker at EC concentrations below the US Mine Safety and Health Administration (MSHA) OEL (<106 µg/m 3 ), below the European Union (EU) OEL (<50 µg/m 3 ) and below the American Conference of Governmental Industrial Hygienists (ACGIH) recommendation (<20 µg/m 3 ). Results Below the MSHA OEL, 17 biomarkers were altered between DEE-exposed workers and unexposed controls. Below the EU OEL, DEE-exposed workers had elevated lymphocytes (p=9E-03, false discovery rate (FDR)=0.04), CD4+ count (p=0.02, FDR=0.05), CD8+ count (p=5E-03, FDR=0.03) and miR-92a-3p (p=0.02, FDR=0.05), and nasal turbinate gene expression (first principal component: p=1E-06, FDR=2E-05), as well as decreased C-reactive protein (p=0.02, FDR=0.05), macrophage inflammatory protein-1β (p=0.04, FDR=0.09), miR-423-3p (p=0.04, FDR=0.09) and miR-122-5p (p=2E-03, FDR=0.02). Even at EC concentrations under the ACGIH recommendation, we found some evidence of exposure-response relationships for miR-423-3p (p trend =0.01, FDR=0.19) and gene expression (p trend =0.02, FDR=0.19). Conclusions DEE exposure under existing or recommended OELs may be associated with biomarkers reflective of cancer-related processes, including inflammatory/immune response.

OBJECTIVES:Diesel exhaust is an established human carcinogen, however the mechanisms by which it leads to cancer development are not fully understood. Mitochondrial dysfunction is an established contributor to carcinogenesis. Recent studies have improved our understanding of the role played by epigenetic modifications in the mitochondrial genome on tumorigenesis. In this study, we aim to evaluate the association between diesel engine exhaust (DEE) exposure with mitochondrial DNA (mtDNA) methylation levels in workers exposed to DEE.METHODS:The study population consisted of 53 male workers employed at a diesel engine manufacturing facility in Northern China who were routinely exposed to diesel exhaust in their occupational setting, as well as 55 unexposed male control workers from other unrelated factories in the same geographic area. Exposure to DEE, elemental carbon, organic carbon, and particulate matter (PM2.5) were assessed. mtDNA methylation for CpG sites (CpGs) from seven mitochondrial genes (D-Loop, MT-RNR1, MT-CO2, MT-CO3, MT-ATP6, MT-ATP8, MT-ND5) was measured in blood samples. Linear regression models were used to estimate the associations between DEE, elemental carbon, organic carbon and PM2.5 exposures with mtDNA methylation levels, adjusting for potential confounders.RESULTS:DEE exposure was associated with decreased MT-ATP6 (difference = -35.6%, P-value = 0.019) and MT-ATP8 methylation (difference = -30%, P-value = 0.029) compared to unexposed controls. Exposures to elemental carbon, organic carbon, and PM2.5 were also significantly and inversely associated with methylation in MT-ATP6 and MT-ATP8 genes (all P-values < 0.05).CONCLUSIONS:Our findings suggest that DEE exposure perturbs mtDNA methylation, which may be of importance for tumorigenesis.

目的 应用气-液界面(air-liquid interface,ALI)染毒技术,探讨摩托车尾气(motorcycle exhaust,ME)对呼吸道不同细胞的细胞器损伤和相对存活率的影响.方法 用20L Tedlar气袋收集洁净空气和ME.实验分为空白对照组、洁净空气组和ME组.空白对照组未做任何处理,洁净空气组为连接0.2 μm滤器后收集的含有21％O2和79％N2的人造空气,ME组为用气袋直接收集的ME.除了空白对照组,其余两组利用ALI体外染毒技术,在染毒流量为25 mL/min、37℃水浴条件下,对生长在ALI插件多孔膜上的BEAS-2B、A549和人肺巨噬细胞持续染毒60 min.利用透射电镜观察细胞内细胞器的损伤.利用CCK-8(cell counting kit-8)检测法评价细胞的相对存活率.结果 细胞电镜显示,对照组和洁净空气组的三种细胞细胞膜均为完整的单层膜结构;线粒体双层膜完整,线粒体嵴清晰可见;核内染色质分布均匀;洁净空气组中偶见少量自噬小体.ME暴露组中,可见BEAS-2B、A549和人肺巨噬细胞的细胞核膜萎缩,线粒体肿胀并伴有线粒体嵴消解和断裂现象,自噬小体的数量明显增多,自噬小体内可见吞噬的细胞器和众多膜泡状结构.BEAS-2B、A549和人肺巨噬细胞相对存活率分别为(61.65±6.23)％、(46.55±6.12)％和(40.89±1.59)％,与洁净空气组相比,3种细胞存活率均显著下降(P＜0.01).方差分析提示,3种细胞存活率差异有统计学意义(F=62.91,P＜0.01),细胞相对存活率呈现BEAS-2B＞A549＞人肺巨噬细胞的趋势.结论 BEAS-2B、A549和人肺巨噬细胞在气-液界面暴露于ME后可引起呼吸道细胞器结构的改变、细胞相对存活率的下降,对呼吸道不同细胞的细胞器和相对存活率存在显著影响.

We investigated whether exposure to carcinogenic diesel engine exhaust (DEE) was associated with altered adduct levels in human serum albumin (HSA) residues. Nano-liquid chromatography-high resolution mass spectrometry (nLC-HRMS) was used to measure adducts of Cys34 and Lys525 residues in plasma samples from 54 diesel engine factory workers and 55 unexposed controls. An untargeted adductomics and bioinformatics pipeline was used to find signatures of Cys34/Lys525 adductome modifications. To identify adducts that were altered between DEE-exposed and unexposed participants, we used an ensemble feature selection approach that ranks and combines findings from linear regression and penalized logistic regression, then aggregates the important findings with those determined by random forest. We detected 40 Cys34 and 9 Lys525 adducts. Among these findings, we found evidence that 6 Cys34 adducts were altered between DEE-exposed and unexposed participants (i.e., 841.75, 851.76, 856.10, 860.77, 870.43, and 913.45). These adducts were biologically related to antioxidant activity.

OBJECTIVE:This study aimed to use an air-liquid interface (ALI) exposure system to simulate the inhalation exposure of motorcycle exhaust particulates (MEPs) and then investigate the benchmark dose (BMD) of MEPs by evaluating cell relative viability (CRV) in lung epithelial BEAS-2B cells.METHODS:The MEPs dose was characterized by measuring the number concentration (NC), surface area concentration (SAC), and mass concentration (MC). BEAS-2B cells were exposed to MEPs at different concentrations via ALI and CRV was determined using Cell Counting Kit (CCK-8) assay. BMD software was applied to calculate BMD and the lower limit of benchmark dose (BMDL) according to Akaike Information Coefficient (AIC), with P-value based on Hill, Linear, Polynomial, and Power model.RESULTS:Our results reveal that BMD of NC and SAC were estimated by the best-fitting Hill model, while MC was estimated by Polynomial model. The BMDL for CRV following ALI exposure to MEPs were as follows: 364.2#/cm 3 for NC; 0.662 × 10 7 nm 2/cm 3 for SAC; and 0.278 μg/m 3 for MC.CONCLUSION:These results indicate that MEPs exposure via ALI system induces a dose-dependent decrease of CRV and provides the potential exposure threshold of MEPs in a lung cell model.

Abstract Background: Diesel engine exhaust (DEE) is classified as a Group 1 human carcinogen by the International Agency for Research on Cancer because of its carcinogenicity to the lung. However, the underlying molecular mechanisms of DEE carcinogenicity are not well understood. Methods: We previously conducted a cross-sectional molecular epidemiology study of diesel engine factory workers exposed to a wide range of DEE and unexposed comparable controls. Here, we carried out a pilot study of a representative subgroup of 19 exposed workers (total n = 54) and 19 unexposed controls (total n = 55). We measured serum level of 1238 targeted proteins using the SOMAScan assay (SOMALogic, Boulder, CA), which measures protein involved in a wide range of biological processes. We used linear regression to identify proteins associated with DEE (permutation p-value <0.01), and examined their exposure-response relationship using a linear trend test across categories of elemental carbon (EC): unexposed (n = 19), lower exposed (median, range: 56.9, 40.2-62.1 µg/m3, n = 9), and higher exposed (median, range: 72.9, 66.9 - 107.7 µg/m3, n = 10), adjusted for age, smoking status, and body mass index. We further assessed correlations of DEE-related proteins with gene expression signature in the nasal epithelium (measured by Affymetrix microarrays) using Pearson's correlation, and examined their pathway enrichment using MetaCore. Results: Occupational exposure to DEE was significantly associated with altered levels of 22 serum proteins. Of these, 13 proteins (CXCL11, HAPLN1, FLT4, CD40LG, PES1, IGHE.IGK..IGL, TNFSF9, PGD, NAGK, CCL25, CCL4L1, PDXK, and PLA2G1B) showed a significant exposure-response relationship with EC (p-trend <0.01), with serum levels of all but PLA2G1B declining with increasing air levels of EC. For instance, CXCL11 showed the most significant association with DEE (β = -0.25; permutation p-value = 0.00004), where the median serum level of CXCL11 was 35.0 relative fluorescent unit (RFU) among the unexposed compared to 29.7 RFU among the lower exposed and 27.8 RFU among the higher exposed group (p-trend = 0.0005). Furthermore, four DEE-related proteins (CXCL11, PPY, CCL25, and SHH) also showed moderate to strong negative correlations (r = -0.52 to -0.72, all permutation p-values <0.01) with the first principal component of a 225 DEE gene expression signature that we previously reported from nasal epithelial cells. Pathway enrichment analysis suggested that these proteins play a role in immunoregulatory and inflammatory processes, including Th17 cell migration; Th1, Th2, and endothelial cell differentiation; and differentiation and clonal expansion of CD8+ T cells (all FDR <0.05). Conclusion: Results from our pilot study suggest that DEE exposure is associated with alteration of multiple proteins in the serum, which play a role in inflammation and immune regulation. Analysis of a larger sample size will be needed to confirm our findings. Citation Format: Mohammad L. Rahman, Yufei Dai, Roel Vermeulen, Wei Hu, Bryan Bassig, Eduard Drizik, Sean Corbett, Dianzhi Ren, Huawei Duan, Yong Niu, Jun Xu, Wei Fu, Kees Meliefste, Baosen Zhou, Xiaohui Zhang, Jufang Yang, Hanqiao Liu, Meng Ye, Gang Liu, Xiaowei Jia, Tao Meng, Ping Bin, Avrum Spira, Marc E. Lenburg, Debra Silverman, Nathaniel Rothman, Yuxin Zheng, Qing Lan. Proteomic analysis of serum in workers exposed to diesel engine exhaust [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 856.

Diesel engine exhaust (DEE) is classified as a Group 1 human carcinogen. Using a targeted proteomics approach, we aimed to identify proteins associated with DEE and characterize these markers to understand the mechanisms of DEE-induced carcinogenicity. In this cross-sectional molecular epidemiology study, we measured elemental carbon (EC) using a personal air monitor and quantified 1317 targeted proteins in the serum using the SOMAScan assay (SOMALogic) among 19 diesel exposed factory workers and 19 unexposed controls. We used linear regressions to identify proteins associated with DEE and examined their exposure-response relationship across levels of EC using linear trend tests. We further examined pathway enrichment of DEE-related proteins using MetaCore. Occupational exposure to DEE was associated with altered levels of 22 serum proteins (permutation p < .01). Of these, 13 proteins (CXCL11, HAPLN1, FLT4, CD40LG, PES1, IGHE.IGK..IGL, TNFSF9, PGD, NAGK, CCL25, CCL4L1, PDXK, and PLA2G1B) showed an exposure-response relationship with EC (p trend < .01), with serum levels of all but PLA2G1B declining with increasing air levels of EC. For instance, C-X-C Motif Chemokine Ligand 11 (CXCL11) showed the most significant association with DEE (β = -0.25; permutation p = .00004), where mean serum levels were 4121.1, 2356.7, and 2298.8 relative fluorescent units among the unexposed, lower exposed (median, range : 56.9, 40.2-62.1 μg/m3 EC), and higher exposed (median, range of EC: 72.9, 66.9-107.7 μg/m3 EC) groups, respectively (p trend = .0005). Pathway analysis suggested that these proteins are enriched in pathways related to inflammation and immune regulation. Our study suggests that DEE exposure is associated with altered serum proteins, which play a role in inflammation and immune regulation.

目的 应用气—液界面(Air-liquid interface,ALI)染毒技术模拟肺部细胞暴露摩托车尾气(Motorcycle exhaust,ME)的方式,探讨ME中非甲烷总烃(Non methane total hydrocarbon,NMHC)和氮氧化物(Nitrogen oxides,NOx)水平与BEAS-2B细胞存活率的关系.方法 固定一辆配备三元催化器的摩托车,用洁净空气将ME直接稀释成体积比VME∶V洁净空气的比例分别为1∶20、1∶10、1∶5和1∶2的不同配比稀释的ME;用采气袋收集不同配比稀释后的ME,气相色谱法分析NMHC的水平;用盐酸萘乙二胺分光光度法测定NOx的浓度.采用ALI染毒系统,在染毒流量为25 ml/min,37℃水浴条件下,对BEAS-2B细胞持续染毒60 min.通过CCK-8细胞毒性检测试剂盒测定细胞的相对存活率.结果 随着ME配比稀释的倍数增加,尾气中NMHC和NOx的浓度呈现降低的趋势,NMHC和NOx的浓度与稀释倍数存在有统计学意义的负相关性(rSpearman分别为-0.859,-0.982,P值均为＜0.001);细胞相对存活率随着ME配比稀释的倍数增加而增加,存在有统计学意义的正相关性(rSpearman为0.839,P＜0.001),从1∶20稀释组到未稀释尾气组,细胞相对存活率的均数依次降低了4.71％、18.28％、26.98％、44.92％和42.59％;ME中NMHC和NOx的水平与BEAS-2B细胞的相对存活率呈负相关性,并具有统计学意义(r分别为-0.956、-0.892,P值分别为0.011、0.042).结论 ME中不同水平的NMHC和NOx可明显影响BEAS-2B细胞的相对存活率,并具有一定的剂量—效应关系.

Background Millions of workers worldwide are exposed to diesel engine exhaust (DEE), a known genotoxic carcinogen. Alu retroelements are repetitive DNA sequences that can multiply and compromise genomic stability. There is some evidence linking altered Alu repeats to cancer and elevated mortality risks. However, whether Alu repeats are influenced by environmental pollutants is unexplored. In an occupational setting with high DEE exposure levels, we investigated associations with Alu repeat copy number. Methods A cross-sectional study of 54 male DEE-exposed workers from an engine testing facility and a comparison group of 55 male unexposed controls was conducted in China. Personal air samples were assessed for elemental carbon, a DEE surrogate, using NIOSH Method 5040. Quantitative PCR (qPCR) was used to measure Alu repeat copy number relative to albumin (Alb) single-gene copy number in leucocyte DNA. The unitless Alu/Alb ratio reflects the average quantity of Alu repeats per cell. Linear regression models adjusted for age and smoking status were used to estimate relations between DEE-exposed workers versus unexposed controls, DEE tertiles (6.1-39.0, 39.1-54.5 and 54.6-107.7 mu g/m(3)) and Alu/Alb ratio. Results DEE-exposed workers had a higher average Alu/Alb ratio than the unexposed controls (p=0.03). Further, we found a positive exposure-response relationship (p=0.02). The Alu/Alb ratio was highest among workers exposed to the top tertile of DEE versus the unexposed controls (1.12 +/- 0.08 SD vs 1.06 +/- 0.07 SD, p=0.01). Conclusion Our findings suggest that DEE exposure may contribute to genomic instability. Further investigations of environmental pollutants, Alu copy number and carcinogenesis are warranted.

ObjectivesTrichloroethylene (TCE) -induced hypersensitivity syndrome (TIHS) is a potentially life-threatening disease. Several genetic susceptibility biomarkers have been found to be associated with TIHS, and this systematic prospective study has been conducted to evaluate the utility of these genetic susceptibility biomarkers in preventing the disease.MethodsThe newly hired TCE-exposed workers were recruited from March 2009 to October 2010. HLA-B*13:01 genotyping and 3-month follow-up procedure were conducted. All workers were monitored for adverse reaction by telephone interview every week. The workers with early symptoms of TIHS were asked to go to the hospital immediately for further examination, diagnosis and treatment. The medical expense record data of patients with TIHS were collected for cost-effectiveness analysis in 2018.ResultsAmong 1651 workers, 158 (9.57%) were found to carry the HLA-B*13:01 allele and 16 (0.97%) were diagnosed with TIHS. HLA-B*13:01 allele was significantly associated with an increased TIHS risk (relative risk=28.4, 95% CI 9.2 to 86.8). As a risk predictor of TIHS, HLA-B*13:01 testing had a sensitivity of 75%, a specificity of 91.1% and an area under curve of 0.83 (95% CI 0.705 to 0.955), the positive and negative predictive values were 7.6% and 99.7%, respectively. The incidence of TIHS was significantly decreased in HLA-B*13:01 non-carriers (0.27%) compared with all workers (0.97%, p=0.014). Cost-effectiveness analysis showed that HLA-B*13:01 screening could produce an economic saving of $4604 per TIHS avoided.ConclusionsProspective HLA-B*13:01 screening may significantly reduce the incidence of TIHS and could be a cost effective option for preventing the disease in TCE-exposed workers.

Objective: To assess the association between circulating C-reactive protein (CRP), and CRP polymorphisms in the diesel engine exhaust (DEE)-exposed workers. Methods: In 137 DEE-exposed workers and 127 unexposed comparable control workers, six urinary mono-hydroxylated polycyclic aromatic hydrocarbons (OH-PAHs) and serum CRP levels were assayed. Genotyping of four CRP single-nucleotide polymorphisms (SNPs) was measured. Results: Serum CRP levels increased in exposed versus control workers (all p < 0.001). In the DEE-exposed workers, two CRP polymorphisms were associated with serum CRP levels, the subjects of rs1205 TT genotype had lower serum CRP levels (p < 0.05 compared to TC or CC). Conclusions: Our findings suggest that polymorphisms in CRP and circulating CRP involved in the inflammatory process may play significant roles in human sensitivity to lung function injury caused by DEE exposure. This study will help investigate the underlying mechanisms of adverse respiratory effects induced by DEE.

Diesel engine exhaust (DEE) is a known lung carcinogen and may be associated with other tumors, however, the mechanisms of action by which DEE causes cancer is not well understood. MicroRNAs (miRNAs), which are small non-coding RNA molecules that play a role in post-transcriptional regulation of gene expression, are altered in multiple tumors and have been observed to be differentially expressed in a variety of biospecimen types in smokers as well as in relation to short-term air pollution exposure. To evaluate whether serum levels of miRNAs are altered in healthy workers occupationally exposed to DEE, we analyzed samples collected in a cross-sectional molecular epidemiology study of diesel engine truck testing facility workers and comparable unexposed controls in China. A panel of 44 miRNAs were measured in 46 workers exposed to relatively high air levels of DEE and 45 controls using the Fireplex circulating miRNA assay (Abcam, Inc.), which profiles miRNAs directly from biofluids. The exposure-response relationship between categorical EC levels and each miRNA was analyzed by linear regression adjusted for age, body mass index, smoking status, current alcohol use and recent infection. We identified two miRNAs that showed a monotonic inverse exposure-response association with DEE: miR-191-5p and miR-93-5p. Levels of miR-191-5p in arbitrary units (A.U.) of fluorescence were 400.7, 333.0, 322.6, and 260.0 in controls and across increasing tertiles of EC, respectively (p for trend = 0.001, FDR = 0.05). Levels of miR-93-5p were 747.1, 713.7, 720.5, and 625.4 A.U. in controls and increasing tertiles of EC, respectively (p for trend = 0.008, FDR = 0.18). Both miRNAs have been reported to influence several biological processes important in carcinogenesis. Our results suggest that occupational exposure to DEE may affect circulating miRNAs in healthy workers. Citation Format: Wei Hu, Bryan A. Bassig, Yufei Dai, Dianzhi Ren, Huawei Duan, Yong Niu, Jun Xu, Wei Fu, Kees Meliefste, Baosen Zhou, Jufang Yang, Meng Ye, Xiaowei Jia, Tao Meng, Ping Bin, Jason YY Wong, Dean H. Hosgood, Nathaniel Rothman, Roel C. Vermeulen, Debra T. Silverman, Yuxin Zheng, Qing Lan. Occupational exposure to diesel engine exhaust and alternations in serum microRNAs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 597.

Mechanisms responsible for diesel exhaust particle (DEP)-induced toxicity in respiratory disorders are poorly understood, recent experimental and controlled exposure studies suggested that oxidative stress might be involved. To investigate the time-course effects DEP on nuclear factor erythroid 2-related factor 2 (Nrf2), a key regulator in cellular adaptive antioxidant response, mice were intratracheal instilled with 100 μg DEP/mouse and sacrificed after 30 min, 6 h, 12 h, 24 h, 48 h, and 72 h. We measured reactive oxygen species (ROS) as well as Nrf2 and antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) and phase II enzymes including heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase-1 (NQO1), glutamate-cysteine ligase catalytic subunit (GCLC), glutamate-cysteine ligase modifier subunit (GCLM) in the lungs. Additionally, histopathological changes were examined. At 6 h, ROS peaked, most of the enzymes were activated, and the histology showed the lungs were damaged. At 12 h, ROS returned to normal level and CAT activity decreased, while protein expression of Nrf2, HO-1, NQO1, GCLC, and GCLM increased, and the lungs were recovering from damage. After 24 h, ROS started to decrease and Nrf2 showed a decreasing trend at both gene and protein levels, while the lung damage had been entirely restored. These results suggested that a single exposure to DEP induce transient oxidative stress in the lungs, with time-dependent effects on Nrf2 and antioxidant enzymes and phase II enzymes.