ABSTRACT
The incidence of disease relating to nanoparticle exposure has been rising rapidly in recent years, for which there is no effective treatment. Macrophage is suggested to play a crucial role in the development of pulmonary disease. To investigate the changes in macrophage after being stimulated by nanometer silica dust and to explore potential biomarkers and signaling pathways, the gene chip GSE13005 was downloaded from Gene Expression Omnibus database, which contained 21 samples: 3 samples per group and 7 groups in total. Macrophages in the control group were cultured in serum-free medium, while the experimental groups were treated with nanometer silica dust in different sizes and concentrations, respectively. To identify the differentially expressed genes and explore their potential functions, we adopted the gene ontology analysis and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis and also constructed protein-protein interaction network. As a result, 1972 differentially expressed genes were identified from 22,690 microarray data in the gene chip, 1069 genes were upregulated and 903 genes were downregulated. Results of the gene ontology analysis indicated that the differentially expressed genes were widely distributed in intracellular and extracellular regions, regulating macrophage apoptosis, inflammatory response, and cell differentiation. The Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the majority of differentially expressed genes were enriched in cytokine-cytokine receptor interaction, cancer or phagosome transcriptional misregulation. The top 10 hub genes, S100a9, Nos3, Psmd14, Psmd4, Lck, Atp6v1h, Jun, Foxh1, Pex14, and Fadd were identified from protein-protein interaction network. In addition, Nos3, Psmd14, Atp6v1h, and Jun were clustered into module M2 (rc = 0.74, p < 0.01), which mainly regulates cell carcinogenesis and antivirus process. In conclusion, differentially expressed genes screened from this study may provide new insights into the exploration of mechanisms, biomarkers, and therapeutic targets for diseases relating to nanoparticle exposure.
Subject(s)
Computational Biology , Gene Regulatory Networks/genetics , Protein Interaction Maps/genetics , Transcriptome/genetics , Apoptosis/drug effects , Databases, Genetic , Gene Expression Regulation, Neoplastic/drug effects , Gene Ontology , Gene Regulatory Networks/drug effects , Humans , Macrophages/drug effects , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Protein Interaction Maps/drug effects , Signal Transduction/drug effects , Silicon Dioxide/administration & dosage , Silicon Dioxide/chemistryABSTRACT
BACKGROUND: Exposure to crystalline silica is considered to increase the risk of lung fibrosis. The primary effector cell, the myofibroblast, plays an important role in the deposition of extracellular matrix (ECM). DNA methylation change is considered to have a potential effect on myofibroblast differentiation. Therefore, the present study was designed to investigate the genome-wide DNA methylation profiles of lung fibroblasts co-cultured with alveolar macrophages exposed to crystalline silica in vitro. METHODS: AM/fibroblast co-culture system was established. CCK8 was used to assess the toxicity of AMs. mRNA and protein expression of collagen I, α-SMA, MAPK9 and TGF-ß1 of fibroblasts after AMs exposed to 100 µg /ml SiO2 for 0-, 24-, or 48 h were determined by means of quantitative real-time PCR, immunoblotting and immunohistochemistry. Genomic DNA of fibroblasts was isolated using MeDIP-Seq to sequence. R software, GO, KEGG and Cytoscape were used to analyze the data. RESULTS: SiO2 exposure increased the expression of collagen I and α-SMA in fibroblasts in co-culture system. Analysis of fibroblast methylome identified extensive methylation changes involved in several signaling pathways, such as the MAPK signaling pathway and metabolic pathways. Several candidates, including Tgfb1 and Mapk9, are hubs who can connect the gene clusters. MAPK9 mRNA expression was significantly higher in fibroblast exposed to SiO2 in co-culture system for 48 h. MAPK9 protein expression was increased at both 24-h and 48-h treatment groups. TGF-ß1 mRNA expression of fibroblast has a time-dependent manner, but we didn't observe the TGF-ß1 protein expression. CONCLUSION: Tgfb1 and Mapk9 are helpful to explore the mechanism of myofibroblast differentiation. The genome-wide DNA methylation profiles of fibroblasts in this experimental silicosis model will be useful for future studies on epigenetic gene regulation during myofibroblast differentiation.
