Wilms tumor 1 impairs apoptotic clearance of fibroblasts in distal fibrotic lung lesions
Harshavardhana H. Ediga, Chanukya P. Vemulapalli, Vishwaraj Sontake, Pradeep K. Patel, Hikaru Miyazaki, Dimitry Popov, Martin B. Jensen, Anil G. Jegga, Steven K. Huang, Christoph Englert, Andreas Schedl, Nishant Gupta, Francis X. McCormack, Satish K. Madala
Harshavardhana H. Ediga, Chanukya P. Vemulapalli, Vishwaraj Sontake, Pradeep K. Patel, Hikaru Miyazaki, Dimitry Popov, Martin B. Jensen, Anil G. Jegga, Steven K. Huang, Christoph Englert, Andreas Schedl, Nishant Gupta, Francis X. McCormack, Satish K. Madala
View: Text | PDF
Research Article Inflammation Pulmonology

Wilms tumor 1 impairs apoptotic clearance of fibroblasts in distal fibrotic lung lesions

Abstract

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disease characterized by impaired fibroblast clearance and excessive extracellular matrix (ECM) protein production. Wilms tumor 1 (WT1), a transcription factor, is selectively upregulated in IPF fibroblasts. However, the mechanisms by which WT1 contributes to fibroblast accumulation and ECM production remain unknown. Here, we investigated the heterogeneity of WT1-expressing mesenchymal cells using single-nucleus RNA-Seq of distal lung tissues from patients with IPF and control donors. WT1 was selectively upregulated in a subset of IPF fibroblasts that coexpressed several prosurvival and ECM genes. The results of both loss-of-function and gain-of-function studies were consistent with a role for WT1 as a positive regulator of prosurvival genes to impair apoptotic clearance and promote ECM production. Fibroblast-specific overexpression of WT1 augmented fibroproliferation, myofibroblast accumulation, and ECM production during bleomycin-induced pulmonary fibrosis in young and aged mice. Together, these findings suggest that targeting WT1 is a promising strategy for attenuating fibroblast expansion and ECM production during fibrogenesis.

Authors

Harshavardhana H. Ediga, Chanukya P. Vemulapalli, Vishwaraj Sontake, Pradeep K. Patel, Hikaru Miyazaki, Dimitry Popov, Martin B. Jensen, Anil G. Jegga, Steven K. Huang, Christoph Englert, Andreas Schedl, Nishant Gupta, Francis X. McCormack, Satish K. Madala

×

Figure 8

WT1 augments fibroblast survival and bleomycin-induced pulmonary fibrosis in aged mice.

Options: View larger image (or click on image) Download as PowerPoint
WT1 augments fibroblast survival and bleomycin-induced pulmonary fibrosi...
(A) Schematic presentation of the study using 15-month-old PDGFRαCreERT (control) and PDGFRαCreERT WT1OE (cWT1OE) mice. (B) Quantification of cell-survival gene transcripts (Wt1, Bcl2, Bcl-XL, and Bcl3) in total lung transcripts for control and cWT1OE mice (n = 3/group). **P < 0.01 and ****P < 0.0001, by multiple unpaired, 2-tailed Student’s t test. (C) The total lung lysates were immunoblotted with antibodies against Wt1, Col1α1, Bax, Bcl2, Bcl-XL, and Gapdh for control and cWT1OE mice. (D) Wt1, Col1α1, Bcl2, and Bcl-XL protein levels were normalized to Gapdh (n = 3/group). *P < 0.05, **P < 0.01, and ***P < 0.001, by 2-tailed Student’s t test. (E) Schematic representation of the animal experiment involving 15-month-old PDGFRαCreERT (control) and PDGFRαCreERT WT1OE (cWT1OE) mice. Mice were treated with bleomycin and tamoxifen as shown in the schema. (F) Representative confocal images of lung sections from 15-month-old control and cWT1OE mice. Arrowheads highlight lung fibroblasts that costained for WT1 (red), vimentin (green), and DAPI (blue) in lung sections from control and cWT1OE mice treated with bleomycin. Scale bars: 20 μm. (G) Representative images of Masson’s trichrome-stained lung sections from control and cWT1OE mice treated with bleomycin. Scale bar: 200 μm. (H) The percentage of fibrotic area was quantified in control and cWT1OE mice using BZ-X image analysis (n = 4–5/group). *P < 0.05, by 2-tailed Student’s t test. (I) Hydroxyproline levels were measured in the right lungs from control and cWT1OE mice treated with bleomycin (n = 4–5/group). *P < 0.05, by 2-tailed Student’s t test. (J) Representative images of αSMA-stained lung sections from control and cWT1OE mice treated with bleomycin. Scale bars: 100 μm. (K) Quantification of αSMA+ area in whole lung sections (n = 4–5/group). *P < 0.05, by 2-tailed Student’s t test. Tam, tamoxifen.