Airway MMP-12 and DNA methylation in COPD: an integrative approach

Abstract

Background: In COPD, the balance between matrix metalloproteinases (MMPs) and their natural inhibitors [tissue inhibitors of metalloproteinases (TIMPs)] is shifted towards excessive degradation, reflected in bronchoalveolar lavage (BAL) as increased MMP concentrations. Because of their critical role in lung homeostasis, MMP activity is tightly regulated, but to what extent this regulation occurs through epigenetic mechanisms remains unknown.

Methods: To explore the interplay between MMPs, TIMPs, and DNA methylation (DNAm) we (1) analysed MMP-9, -12, and TIMP-1 concentrations in BAL fluid, and profiled DNAm in BAL cells from 18 COPD and 30 control subjects, (2) estimated protein-COPD relationships using multivariable regression, (3) identified protein quantitative trait methylation loci (pQTMs) with COPD as a potential modifier in a separate interaction model, and (4) integrated significant interactions with a previous COPD GWAS meta-analysis.

Results: COPD was associated with higher levels of BAL MMP-12 (p = 0.016) but not with MMP-9 or TIMP-1. Further examination of MMP-12 identified association with DNAm at 34 loci (pQTMs), with TGFBR2 (p = 2.25 × 10^-10) and THBS4 (p = 1.11 × 10^-9) among the top ten pQTM genes. The interaction model identified 66 sites where the DNAm-MMP-12 association was significantly different in COPD compared to controls. Of these, one was colocalized with SNPs previously associated with COPD.

Conclusions: Our findings indicate that airway MMP-12 may partially be regulated by epigenetic mechanisms and that this regulation is disrupted in COPD. Furthermore, integration with COPD GWAS data suggests that this dysregulation is influenced by a combination of environmental factors, disease processes, and genetics, with the latter potentially playing a lesser role.

Keywords: Bronchoscopy; Chronic obstructive pulmonary disease (COPD); DNA methylation; Extracellular matrix remodelling; Matrix metalloproteinases (MMPs); Multiomics.

Fig. 1

Study overview. DNA methylation in BAL cells and concentrations of proteins MMP-9, MMP-12, and TIMP-1 in BAL fluid were measured. The relationship between COPD and protein concentrations was tested using multivariable regression, and proteins exhibiting significant associations were examined further in a protein-epigenome-wide association study (pEWAS). To evaluate the effect of COPD on protein–DNA methylation-relationships, an interaction pEWAS was implemented. All pQTMs were annotated to genes, and interaction pQTMs were tested for colocalization with genetic variation associated with COPD. BAL: bronchoalveolar lavage; MMP: matrix metalloproteinase; TIMP: tissue inhibitor of metalloproteinases; GREAT: Genomic Regions Enrichment of Annotations Tool; GWAS: genome-wide association study; pQTM: protein quantitative trait methylation loci; UCSC: The University of California, Santa Cruz, Genome Browser Database

Fig. 2

Concentrations of MMP-9, -12, and TIMP-1 in bronchoalveolar lavage. Dotted line indicates median, bars the interquartile range. MMP: Matrix metalloproteinase. TIMP: tissue inhibitor of metalloproteinases. COPD: chronic obstructive pulmonary disease

Fig. 3

Association between airway MMP-12 and DNA methylation. pEWAS Manhattan plot. Three most significant pQTMs labelled with annotated genes. Black line represents the Bonferroni-corrected threshold for significance. pEWAS: protein-epigenome-wide association study; MMP-12: Matrix metalloproteinase 12; pQTM: protein quantitative trait methylation loci; Chr: chromosome; HOXD1: Homeobox D1; LZTS1-AS1: LZTS1 Antisense RNA 1; ERO1A: Endoplasmic Reticulum Oxidoreductase 1 Alpha

Fig. 4

The effect of COPD on the MMP-12–DNA methylation relationship. A Interaction pEWAS Manhattan plot indicating interactions between COPD and DNA methylation associated with MMP-12. Three most significant interaction pQTMs labelled with annotated genes. Black line represents the Bonferroni-corrected threshold for significance. B Selected interaction pQTMs. X-axis: DNA methylation, y-axis: MMP-12 concentrations. pEWAS: protein-epigenome-wide association study; MMP-12: Matrix metalloproteinase 12; pQTM: protein quantitative trait methylation loci; Chr: chromosome; COPD: chronic obstructive pulmonary disease; PRKAG2: Protein Kinase AMP-Activated Non-Catalytic Subunit Gamma 2. ACSF3: Acyl-CoA Synthetase Family Member 3; LINC00378: Long Intergenic Non-Protein Coding RNA 378