Fibrocytes Regulate Wilms Tumor 1–Positive Cell Accumulation in Severe Fibrotic Lung Disease

Abstract

Collagen-producing myofibroblast transdifferentiation is considered a crucial determinant in the formation of scar tissue in the lungs of patients with idiopathic pulmonary fibrosis (IPF). Multiple resident pulmonary cell types and bone marrow-derived fibrocytes have been implicated as contributors to fibrotic lesions due to the transdifferentiation potential of these cells into myofibroblasts. In this study, we assessed the expression of Wilms’ tumor 1 (WT1), a known marker of mesothelial cells, in various cell types in normal and fibrotic lungs. We demonstrate that WT1 is expressed by both mesothelial and mesenchymal cells in IPF lungs, but has limited or no expression in normal human lungs. We also demonstrate that WT1-positive cells accumulate in fibrotic lung lesions, using two different mouse models of pulmonary fibrosis and WT1 promoter-driven fluorescent reporter mice. Reconstitution of bone-marrow cells into a transforming growth factor-α transgenic-mouse model demonstrated that fibrocytes do not transform into WT1-positive mesenchymal cells, but do augment accumulation of WT1-positive cells in severe fibrotic lung disease. Importantly, the number of WT1-positive cells in fibrotic lesions were correlated with severity of lung disease as assessed by changes in lung function, histology, and hydroxyproline levels in mice. Finally, inhibition of WT1 expression was sufficient to attenuate collagen and other extracellular-matrix gene production by mesenchymal cells from both murine and human fibrotic lungs. Thus, the results of this study demonstrate a novel association between fibrocyte-driven WT1-positive cell accumulation and severe fibrotic lung disease.

Heterogeneity of Fibroblasts and Myofibroblasts in Pulmonary Fibrosis

“…Most studies characterizing pulmonary mesothelial cells in lung fibrosis utilize Wilms’ tumor suppressor gene, wt1 , as a marker for these cells. Wt1 is exclusively expressed in embryonic mesothelial cells as early as embryonic day E10.5 and by E18.5 its expression levels significantly declines such that this gene is not expressed in adult lungs[21, 43]. Importantly, several studies have shown that this tumor suppressor is induced after lung injury and/or remodeling[44, 45, 43].…”

“…Wt1 is exclusively expressed in embryonic mesothelial cells as early as embryonic day E10.5 and by E18.5 its expression levels significantly declines such that this gene is not expressed in adult lungs[21, 43]. Importantly, several studies have shown that this tumor suppressor is induced after lung injury and/or remodeling[44, 45, 43]. Two studies utilizing Wt1 tm1EGFP/Cre)wtp/ J to label all cells with an active wt1 promoter with EGFP have showed that lineage-labeled cells invaded into the pulmonary parenchyma and a subset of these cells expressed αSMA at 4 to 24 h after intratracheal delivery of active TGFß[44, 45].…”

“…This was confirmed using a different wt1 lineage tracing mouse strain, Wt1 Tm2(Cre/ERT2)Wtp /J; B6.129S4-Gt(ROSA)-26Sor tm1Sor /J to label wt1 -expressing cells irreversibly with ß-gal at 4 h after intratracheal TGFß administration[44]. Further, in a CCSP/TGFα transgenic mouse model of lung fibrosis, Wt1-immunostained cells appeared to invade into the pulmonary parenchyma and express vimentin by co-immunofluorescent analysis[43]. Interestingly, utilizing a Wt1 CreERT1/+ ROSA mTmG/+ strain to label all cells with tdTomato and wt1 -expressing cells with GFP, intradermal bleomycin administration induced pleural thickening and increased wt1 lineage-labeled cells in the pulmonary pleura but these cells were not observed to invade into the pulmonary parenchyma[43].…”

Role of CXCL12/CXCR4-Mediated Circulating Fibrocytes in Pulmonary Fibrosis

“…12,15,16 Additionally, fibrocytes acted as a regulator that promoted progressive accumulation of Wilms' Tumor 1 (WT1)-positive cells, a sizable subset of lung-resident mesenchymal cells, in fibrotic lesions of both human idiopathic pulmonary fibrosis (IPF) and mouse models of pulmonary fibrosis. 17 The circulating fibrocytes secrete periostin, a matricellular…”

Unique and Redundant Functions of p70 Ribosomal S6 Kinase Isoforms Regulate Mesenchymal Cell Proliferation and Migration in Pulmonary Fibrosis

“…Separately, wild-type and S6K knockout (KO) mice were administered bleomycin (0.1 mg) daily in 0.05 ml saline solution intradermally 5 days per week for 4 weeks, as previously described (20).…”