Profiling of the silica-induced molecular events in lung epithelial cells using the RNA-Seq approach Judy Y. W. Chan a * , Joseph C. C. Tsui b , Patrick T. W. Law c , Winnie K. W. So a , Doris Y. P. Leung a , Michael M. K. Sham d , Stephen K. W. Tsui e and Carmen W. H. Chan a * ABSTRACT: Silicosis is a prolonged, irreversible and incurable occupational disease, and there is a significant number of newly diagnosed cases every year in Hong Kong. Due to the long latency of the disease, the diagnosis can be missed until detailed clinical examination at a later stage. For a better control of this deadly disease, detailing the pro-inflammatory and fibrotic events in the macrophage would be instrumental in understanding the pathogenesis of the disease and essential for the significant biomarkers discovery. In this in vitro study, human cell line model A549 lung epithelial cells were used. The immediate molecular events underneath the activation of quartz silica polymorphs were followed in a time course of 0, 0.5, 2, 8, 16 and 24 h. The transcriptome library was prepared and subjected to RNA-Seq analysis. Data analysis was performed by pathway analysis tools and verified by real-time PCR. The results showed that triggered genes were mainly found in the immune response and inflammatory pathways. An interesting finding was the association of the DNA-binding protein inhibitor (ID) family in the silica exposure to lung cells. The linkage of ID1, ID2 and ID3 to cancer may rationalize themselves to be the markers indicating an early response of silicosis. However, further studies are required to consolidate the roles of these genes in silicosis. Copyright © 2017 John Wiley & Sons, Ltd. Keywords: silicosis; crystalline silica; quartz; DNA-binding inhibitor Introduction Silicosis, an irreversible and incurable lung disease highly associated with industries and occupations such as construction and cement manufacture, is caused by chronic exposure to crystalline silica. It is a worldwide occupational disease that affects more than 35 millions workers in the world including China, Europe and the United States (Rafferty, 2012; Leung et al., 2012; Minnesota Department of Labor and Industry Occupational Safety and Health Division (Minnesota OSH), 2012; Mossman and Glenn, 2013;) and raises health concerns such as silicosis, malignancies and autoimmune diseases (Rimal et al., 2005). There are more than 500 000 cases recorded between 1991 and 1995, and 6000 new cases annually being reported in China. The disease causes about 24 000 deaths annually (Leung et al., 2012). Inhalation of crystalline silica induces various pathogenic events, and the known underlying mechanisms include cytotoxicity to lung cells, oxidative stress, stimulation of inflammatory responses and induction of fibrosis. The toxicity of crystalline silica appears to result from the ability of crystalline silica surfaces to interact with aqueous media, to generate oxygen radicals and to injure target pulmonary cells such as alveolar macrophages. Resultant generation of inflammatory cytokines (e.g. interleukin-1 and tumor necrosis factor beta) by target cells lead to cytokine networking between inflammatory cells and resident pulmonary cells, resulting in inflammation and fibrosis (Rimal et al., 2005). The diagnosis of silicosis is not effective. The effect of silica is most likely chronic and the latency period can be up to 10years. Diagnosis of the disease depends mostly on clinical examinations; however, it is not sensitive to detect those cases without the development of typical nodules (Leung et al., 2012). Efforts are being made to develop strategies to relief the burden of the silicosis to patients and to improve the quality of life of patients (PCFB, 2013). There are, however, no available effective prognosis and diagnosis strategies for this incurable lung disease. As a result, an early prognostic strategy or prediction would be helpful to control this irreversible and incurable disease. The aim of the present study was to discover the genes/pathways which are newly found to be involved in the early stage of silicosis using the RNA-Seq strategy. *Correspondence to: Carmen Chan, Room 732, Esther Lee Building, The Chinese University of Hong Kong, Shatin, Hong Kong. E-mail: whchan@cuhk.edu.hk *Correspondence to: Judy Chan, Room 816, Esther Lee Building, The Chinese University of Hong Kong, Shatin, Hong Kong. E-mail: judychanyw@cuhk.edu.hk a Nethersole School of Nursing, The Chinese University of Hong Kong, Shatin, Hong Kong b Department of Orthopaedics and Traumatology, The Chinese University of Hong Kong, Shatin, Hong Kong c School of Life Sciences, The Chinese University of Hong Kong, Shatin, Hong Kong d Grantham Hospital, Aberdeen, Hong Kong e School of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, Hong Kong J. Appl. Toxicol. 2017 Copyright © 2017 John Wiley & Sons, Ltd. Research article Received: 1 February 2017, Revised: 23 February 2017, Accepted: 24 February 2017 Published online in Wiley Online Library (wileyonlinelibrary.com) DOI 10.1002/jat.3471