Pulmonary toxicity and global gene expression changes in response to sub-chronic inhalation exposure to crystalline silica in rats

Occupational exposure to crystalline silica results in serious health effects, most notably, silicosis and cancer. A proper understanding of the mechanism(s) underlying the initiation and progression of silica-induced pulmonary toxicity is critical for the intervention and/or prevention of the adverse health effects associated with crystalline silica exposure. Rats were exposed to crystalline silica by inhalation at a concentration of 15 mg/m3, 6 hours/day, 5 days/week for 3, 6 or 12 weeks. At the end of each exposure time point, toxicity and global gene expression changes were determined in the lungs. In general, silica exposure resulted in pulmonary toxicity that was dependent on the duration of silica exposure. A significant and silica exposure time-dependent increase in lactate dehydrogenase activity and accumulation of alveolar macrophages and infiltrating neutrophils in the bronchoalveolar lavage fluid suggested crystalline silica-induced pulmonary toxicity in the rats. Histological changes indicative of pulmonary toxicity were detectable only in the lungs of rats that were exposed to silica for 6- or 12-weeks. Minimal, sub-acute pulmonary inflammation consisting mainly of macrophage accumulation and infiltration of neutrophils was seen in 2 out of 8 rats in the 6-week silica exposure group. Chronic active inflammation, type II pneumocyte hyperplasia, and fibrosis were detected following 12-weeks of silica exposure in all rat lungs. In addition, crystalline silica was visible in the lungs of the rats belonging to the 12-week exposure group. A significant increase in the number of neutrophils seen in the blood indicated silica-induced systemic inflammation in the rats. Microarray analysis of the global gene expression profiles of the rat lungs detected significant differential expression (FDR p <0.05 and fold change >1.5) of 38, 77 and 99 genes in the rats exposed to silica for 3-, 6- and 12-weeks, respectively, compared to the time-matched controls. Bioinformatics analysis of the differentially expressed genes identified significant enrichment of functions, networks and pathways related to inflammation, cancer, oxidative stress, fibrosis and tissue remodeling in the lungs of the silica exposed rats. Collectively, the results of our study provided insights into the molecular mechanisms underlying pulmonary toxicity following sub-chronic exposure to silica in rats.职业暴露于结晶二氧化硅（crystalline silica）会引发严重健康损害，其中最为显著的是矽肺（silicosis）与癌症。深入理解二氧化硅诱导的肺毒性发生及进展的分子机制，对于干预乃至预防结晶二氧化硅暴露相关的不良健康效应至关重要。本研究通过吸入方式让大鼠暴露于浓度为15 mg/m³的结晶二氧化硅环境中，暴露方案为每日6小时、每周5天，分别持续3、6或12周。在每个暴露周期结束时，检测大鼠肺部的毒性反应及全基因表达变化。总体而言，二氧化硅暴露引发的肺毒性与暴露时长呈显著依赖关系。支气管肺泡灌洗液（bronchoalveolar lavage fluid）中乳酸脱氢酶（lactate dehydrogenase）活性显著升高，同时肺泡巨噬细胞与浸润性中性粒细胞（neutrophils）的积聚量随暴露时间延长而增加，上述结果提示结晶二氧化硅可诱导大鼠产生肺毒性。仅在暴露6周或12周的大鼠肺部中，可检测到提示肺毒性的组织学改变。在6周暴露组的8只大鼠中，有2只出现了以巨噬细胞积聚、中性粒细胞浸润为主要特征的轻度亚急性肺部炎症。在12周暴露组的所有大鼠肺部中，均检测到慢性活动性炎症、II型肺泡上皮细胞（type II pneumocyte）增生以及纤维化（fibrosis）。此外，12周暴露组大鼠的肺部中可见结晶二氧化硅沉积。血液中中性粒细胞数量显著升高，提示二氧化硅诱导大鼠产生了全身性炎症。对大鼠肺部的全基因表达谱进行微阵列分析（microarray analysis）结果显示，与同时间点的对照组相比，暴露3、6、12周的大鼠分别有38、77和99个基因出现显著差异表达（错误发现率（False Discovery Rate, FDR）p值<0.05，折叠变化>1.5）。对差异表达基因进行的生物信息学分析（bioinformatics analysis）表明，二氧化硅暴露大鼠的肺部中，与炎症、癌症、氧化应激、纤维化及组织重塑相关的功能、调控网络及通路均出现显著富集。综上，本研究结果为理解大鼠亚慢性暴露于二氧化硅后肺毒性的分子机制提供了新的见解。