Transcriptomic analysis reveals distinct effects of cigarette smoke on murine airspace and bone-marrow derived macrophages

Abstract

Background: Chronic obstructive pulmonary disease (COPD) is an inflammatory airway disease characterized by emphysema and chronic bronchitis and a leading cause of mortality worldwide. COPD is commonly associated with several comorbid diseases which contribute to exacerbated patient outcomes. Cigarette smoke (CS) is the most prominent risk factor for COPD development and progression and is known to be detrimental to numerous effector functions of lung resident immune cells, including phagocytosis and cytokine production. However, how CS mediates the various pathologies distant from the lung in COPD, and whether CS has a similar biological effect on systemic immune cells remains unknown.

Methods: C57BL/6 mice were exposed to 8 weeks of CS as an experimental model of COPD. Bone marrow cells were isolated from both CS-exposed and room air (RA) control mice and differentiated to bone marrow-derived macrophages (BMDMs). Airspace macrophages (AMs) were isolated from the same CS-exposed and RA mice and bulk RNA-Seq performed. The functional role of differentially expressed genes was assessed through gene ontology analyses. Ingenuity Pathway Analysis was used to determine the activation states of canonical pathways and upstream regulators enriched in differentially expressed genes in both cell types, and to compare the differences between the two cell types.

Results: CS induced transcriptomic changes in BMDMs, including an upregulation of genes in sirtuin signalling and oxidative phosphorylation pathways and a downregulation of genes involved in histone and lysine methylation. In contrast, CS induced decreased expression of genes involved in pathogen response, phagosome formation, and immune cell trafficking in AMs. Little overlap was observed in differentially expressed protein-coding genes in BMDMs compared to AMs and their associated pathways, highlighting the distinct effects of CS on immune cells in different compartments.

Conclusions: CS exposure can induce transcriptomic remodelling in BMDMs which is distinct to that of AMs. Our study highlights the ability of CS exposure to affect immune cell populations distal to the lung and warrants further investigation into the functional effects of these changes and the ensuing role in driving multimorbid disease.

Keywords: COPD; Cigarette smoke; Macrophage; Transcriptome.

Fig. 1

CS exposure induces transcriptomic remodelling in BMDMs. A Schematic of experiment (created using Biorender). B Volcano plot of BMDM differentially expressed genes (DEGs) in CS-exposed mice (n = 3) compared to room air (RA) controls (n = 3). Biological processes associated with BMDM CS-induced upregulated (C) and downregulated (D) DEGs. E Canonical pathways produced from Ingenuity Pathway Analysis enriched in BMDM CS-induced DEGs and their activation z-scores (blue = inhibited, orange = activated)

Fig. 2

CS exposure induces distinct transcriptomic remodelling in BMDMs compared to AMs. A Volcano plot of AM DEGs in CS-exposed mice (n = 4) compared to RA (n = 4). B Biological processes associated with AM CS-induced downregulated DEGs. C Canonical pathways produced from Ingenuity Pathway Analysis enriched in AM CS-induced DEGs and their activation z-scores. D Comparison of Log2 fold change values in BMDMs and AMs from CS exposed mice compared to RA, with significant DEGs common to both cell types highlighted. E Heat map comparing activation z-score of canonical pathways induced by CS in BMDMs and AMs