DNA Methylation Profiling in a Cigarette Smoke-Exposed Mouse Model of Airway Inflammation

Abstract

Purpose

DNA methylation, a major epigenetic modification, has been documented to play an important role in chronic obstructive pulmonary disease (COPD). In this study, we aimed to profile the DNA methylation patterns in a mouse model of airway inflammation induced by cigarette smoke (CS), a foremost risk factor of COPD.

Material and Methods

To establish a model of airway inflammation, wild-type mice were exposed to mainstream CS or room air for 2 hours twice daily, 6 days per week for consecutive 4 weeks. Lung tissues of the mice were collected for genome-wide DNA methylation analysis by liquid hybridization capture-based bisulfite sequencing, which were used for intersection analysis with gene expression by cDNA microarray to identify candidate methylated genes. Then, functional enrichment analyses with protein–protein interaction (PPI) network regarding these genes were conducted to explore the potential mechanisms.

Results

After 4-week CS exposure, the level of DNA methylation accompanied by a subacute airway inflammation was markedly enhanced, and 2002 differentially methylated genes (DMGs) were annotated, including 565 DMGs contained methylations in gene promoters, which were used for intersection with the differentially expressed genes. Then, 135 candidate methylated genes were further selected by the intersection, among which 58 genes with functional methylated modification were finally identified. Further analyses revealed candidate methylated genes were significantly enriched in a complicated network of signals and processes, including interleukins, toll-like receptors, T-cells differentiation, oxidative stress, mast cells activation, stem cells proliferation, etc., as well as the 58 functional methylated genes were partially located at key positions in PPI network, especially CXCL1, DDX58 and JAK3.

Conclusion

This study suggests CS exposure significantly enhances DNA methylated level, and the potential functional methylated genes are closely related to complicated inflammatory-immune responses, which may provide some new experimental evidence in understanding the epigenetic mechanisms of CS-induced airway inflammation in COPD.

Keywords:

• chronic obstructive pulmonary disease
• airway inflammation
• cigarette smoke
• DNA methylation
• liquid hybridization capture-based bisulfite sequencing

Abbreviations

COPD, chronic obstructive pulmonary disease; CS, cigarette smoke; CXCL1, C-X-C Motif Chemokine Ligand 1; DDX58, DExD/H-Box Helicase 58; DEGs, differentially expressed genes; DMGs, differentially methylated genes; DMRs, differentially methylated regions; GO, Gene Ontology; HFMD, hand, foot and mouth disease; IQR, interquartile range; JAK3, Janus Kinase 3; KEGG, Kyoto Encyclopedia of Genes and Genomes; LHC-BS, Liquid hybridization capture-based bisulfite sequencing; PPI, protein–protein interaction; PRR, pattern recognition receptor; TLR6, Toll Like Receptor 6; TPM, total particulate matter.

Data Sharing Statement

The datasets generated and analyzed during the present study are available from the corresponding authors on reasonable requests.

Author Contributions

All authors made a significant contribution to the work reported, whether that is in the conception, study design, execution, acquisition of data, analysis and interpretation, or in all these areas; took part in drafting, revising or critically reviewing the article; gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agree to be accountable for all aspects of the work.

Disclosure

The authors report no conflicts of interest in this work.

Additional information

Funding

This study was supported in part by grant 81970040 from the National Natural Science Foundation of China and grant 18PJ410 from the Health and Family Planning Commission of Sichuan Province.