Integrating murine gene expression studies to understand obstructive lung disease due to chronic inhaled endotoxin

Abstract

Rationale: Endotoxin is a near ubiquitous environmental exposure that that has been associated with both asthma and chronic obstructive pulmonary disease (COPD). These obstructive lung diseases have a complex pathophysiology, making them difficult to study comprehensively in the context of endotoxin. Genome-wide gene expression studies have been used to identify a molecular snapshot of the response to environmental exposures. Identification of differentially expressed genes shared across all published murine models of chronic inhaled endotoxin will provide insight into the biology underlying endotoxin-associated lung disease.

Methods: We identified three published murine models with gene expression profiling after repeated low-dose inhaled endotoxin. All array data from these experiments were re-analyzed, annotated consistently, and tested for shared genes found to be differentially expressed. Additional functional comparison was conducted by testing for significant enrichment of differentially expressed genes in known pathways. The importance of this gene signature in smoking-related lung disease was assessed using hierarchical clustering in an independent experiment where mice were exposed to endotoxin, smoke, and endotoxin plus smoke.

Results: A 101-gene signature was detected in three murine models, more than expected by chance. The three model systems exhibit additional similarity beyond shared genes when compared at the pathway level, with increasing enrichment of inflammatory pathways associated with longer duration of endotoxin exposure. Genes and pathways important in both asthma and COPD were shared across all endotoxin models. Mice exposed to endotoxin, smoke, and smoke plus endotoxin were accurately classified with the endotoxin gene signature.

Conclusions: Despite the differences in laboratory, duration of exposure, and strain of mouse used in three experimental models of chronic inhaled endotoxin, surprising similarities in gene expression were observed. The endotoxin component of tobacco smoke may play an important role in disease development.

Figure 1. Comparison of 3 identified studies of murine inhaled endotoxin exposure used to generate gene signature.

1a. Common intersect of differentially regulated genes (endotoxin vs. control exposure) identifies a gene signature for endotoxin exposure. 1b. Common intersect of arrays used in each studies represents a background distribution to identify the statistical significance of the gene signature. 1c. 2-way hypergeometric tests to identify statistical significance of gene signature.

Figure 2. Comparison of all 4 identified studies at the pathway level.

Pathway enrichment calculated using hypergeometric tests with all known pathways present in Netpath, Wikipathways, Kegg, and Reactome, with Cytoscape for visualization. 2a. Lai et al, 5 day exposure. 2b. Lai et al, 8 week exposure. 2c. Meng et al, 3 week exposure. 2d. Brass et al, 1 week exposure.

Figure 3. Consensus clustering of air, endotoxin, smoke, and endotoxin plus smoke exposed mice using endotoxin gene signature and random gene signature.

The endotoxin gene signature accurately clusters the different exposure groups 99.97% of the time as compared to a randomly chosen gene signature which accurately clusters the different exposure groups 78.15% of the time.