The lung extracellular matrix protein landscape in severe early-onset and moderate chronic obstructive pulmonary disease.

Extracellular matrix (ECM) remodeling has been implicated in the irreversible obstruction of airways and destruction of alveolar tissue in chronic obstructive pulmonary disease (COPD). Studies investigating differences in the lung ECM in COPD have mainly focused on some collagens and elastin, leaving an array of ECM components unexplored. We investigated the differences in the ECM landscape comparing severe-early onset (SEO-) COPD and moderate COPD to control lung tissue for collagen type I α chain 1 (COL1A1), COL6A1, COL6A2, COL14A1, fibulin 2 and 5 (FBLN2, FBLN5), latent transforming growth factor-beta binding protein 4 (LTBP4), lumican (LUM), versican (VCAN), decorin (DCN), and elastin (ELN) using image analysis and statistical modelling. Percentage area and/or mean intensity of expression of LUM in the parenchyma, and COL1A1, FBLN2, LTBP4, DCN, and VCAN in the airway walls, was proportionally lower in COPD compared to controls. Lowered levels of most ECM proteins were associated with decreasing FEV1 measurements, indicating a relationship with disease severity. Furthermore, we identified six unique ECM signatures where LUM and COL6A1 in parenchyma and COL1A1, FBLN5, DCN, and VCAN in airway walls appear essential in reflecting the presence and severity of COPD. These signatures emphasize the need to examine groups of proteins to represent an overall difference in the ECM landscape in COPD, that are more likely to be related to functional effects, than individual proteins. Our study revealed differences in the lung ECM landscape between control and COPD and between SEO and moderate COPD signifying distinct pathological processes in the different subgroups.

FAM13A regulates cellular senescence marker p21 and mitochondrial reactive oxygen species production in airway epithelial cells.

Inhalation of noxious gasses induces oxidative stress in airway epithelial cells (AECs), which may lead to cellular senescence and contribute to the development of chronic obstructive pulmonary disease (COPD). FAM13A, a well-known COPD susceptibility gene, is highly expressed in airway epithelium. We studied whether its expression is associated with aging and cellular senescence and affects airway epithelial responses to paraquat, a cellular senescence inducer. The association between age and FAM13A expression was investigated in two datasets of human lung tissue and bronchial brushings from current/ex-smokers with/without COPD. Protein levels of FAM13A and cellular senescence marker p21 were investigated using immunohistochemistry in lung tissue from patients with COPD. In vitro, FAM13A and P21 expression was assessed using qPCR in air-liquid-interface (ALI)-differentiated AECs in absence/presence of paraquat. In addition, FAM13A was overexpressed in human bronchial epithelial 16HBE cells and the effect on P21 expression (qPCR) and mitochondrial reactive oxygen species (ROS) production (MitoSOX staining) was assessed. Lower FAM13A expression was significantly associated with increasing age in lung tissue and bronchial epithelium. In airway epithelium of patients with COPD, we found a negative correlation between FAM13A and p21 protein levels. In ALI-differentiated AECs, the paraquat-induced decrease in FAM13A expression was accompanied by increased P21 expression. In 16HBE cells, the overexpression of FAM13A significantly reduced paraquat-induced P21 expression and mitochondrial ROS production. Our data suggest that FAM13A expression decreases with aging, resulting in higher P21 expression and mitochondrial ROS production in the airway epithelium, thus facilitating cellular senescence and as such potentially contributing to accelerated lung aging in COPD.NEW & NOTEWORTHY To our knowledge, this is the first study investigating the role of the COPD susceptibility gene FAM13A in aging and cellular senescence. We found that FAM13A negatively regulates the expression of the cellular senescence marker P21 and mitochondrial ROS production in the airway epithelium. In this way, the lower expression of FAM13A observed upon aging may facilitate cellular senescence and potentially contribute to accelerated lung aging in COPD.

A proteomics approach to identify COPD-related changes in lung fibroblasts.

Lung fibroblasts are implicated in abnormal tissue repair in chronic obstructive pulmonary disease (COPD). The exact mechanisms are unknown and comprehensive analysis comparing COPD- and control fibroblasts is lacking. The aim of this study is to gain insight into the role of lung fibroblasts in COPD pathology using unbiased proteomic and transcriptomic analysis. Protein and RNA were isolated from cultured parenchymal lung fibroblasts of 17 patients with stage IV COPD and 16 non-COPD controls. Proteins were analyzed using LC-MS/MS and RNA through RNA sequencing. Differential protein and gene expression in COPD was assessed via linear regression, followed by pathway enrichment, correlation analysis, and immunohistological staining in lung tissue. Proteomic and transcriptomic data were compared to investigate the overlap and correlation between both levels of data. We identified 40 differentially expressed (DE) proteins and zero DE genes between COPD and control fibroblasts. The most significant DE proteins were HNRNPA2B1 and FHL1. Thirteen of the 40 proteins were previously associated with COPD, including FHL1 and GSTP1. Six of the 40 proteins were related to telomere maintenance pathways, and were positively correlated with the senescence marker LMNB1. No significant correlation between gene and protein expression was observed for the 40 proteins. We hereby describe 40 DE proteins in COPD fibroblasts including previously described COPD proteins (FHL1, GSTP1) and new COPD research targets like HNRNPA2B1. Lack of overlap and correlation between gene and protein data supports the use of unbiased proteomics analysis and indicates that different types of information are generated with both methods.

A proteome comparison between human fetal and mature renal extracellular matrix identifies EMILIN1 as a regulator of renal epithelial cell adhesion.

Cell-based approaches using tissue engineering and regenerative medicine to replace damaged renal tissue with 3D constructs is a promising emerging therapy for kidney disease. Besides living cells, a template provided by a scaffold based on biomaterials and bioactive factors is needed for successful kidney engineering. Nature's own template for a scaffolding system is the extracellular matrix (ECM). Research has focused on mapping the mature renal ECM; however, the developing fetal ECM matches more the active environment required in 3D renal constructs. Here, we characterized the differences between the human fetal and mature renal ECM using spectrometry-based proteomics of decellularized tissue. We identified 99 different renal ECM proteins of which the majority forms an overlapping core, but also includes proteins enriched in either the fetal or mature ECM. Relative protein quantification showed a significant dominance of EMILIN1 in the fetal ECM. We functionally tested the role of EMILIN1 in the ECM using a novel methodology that permits the reliable anchorage of native cell-secreted ECM to glass coverslips. Depletion of EMILIN1 from the ECM layer using siRNA mediated knock-down technologies does not affect renal epithelial cell growth, but does promote migration. Lack of EMILIN1 in the ECM layer reduces the adhesion strength of renal epithelial cells, shown by a decrease in focal adhesion points and associated stress fibers. We showed in this study the importance of a human renal fetal and mature ECM catalogue for identifying promising ECM components that have high implementation potential in scaffolds for 3D renal constructs.