Vimentin intermediate filament assembly regulates fibroblast invasion in fibrogenic lung injury
Ranu Surolia, … , Victor J. Thannickal, Veena B. Antony
Ranu Surolia, … , Victor J. Thannickal, Veena B. Antony
Published April 4, 2019
Citation Information: JCI Insight. 2019;4(7):e123253. https://doi.org/10.1172/jci.insight.123253.
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Research Article Pulmonology

Vimentin intermediate filament assembly regulates fibroblast invasion in fibrogenic lung injury

Abstract

Idiopathic pulmonary fibrosis (IPF) is a progressive disease, with a median survival of 3–5 years following diagnosis. Lung remodeling by invasive fibroblasts is a hallmark of IPF. In this study, we demonstrate that inhibition of vimentin intermediate filaments (VimIFs) decreases the invasiveness of IPF fibroblasts and confers protection against fibrosis in a murine model of experimental lung injury. Increased expression and organization of VimIFs contribute to the invasive property of IPF fibroblasts in connection with deficient cellular autophagy. Blocking VimIF assembly by pharmacologic and genetic means also increases autophagic clearance of collagen type I. Furthermore, inhibition of expression of collagen type I by siRNA decreased invasiveness of fibroblasts. In a bleomycin injury model, enhancing autophagy in fibroblasts by an inhibitor of VimIF assembly, withaferin A (WFA), protected from fibrotic lung injury. Additionally, in 3D lung organoids, or pulmospheres, from patients with IPF, WFA reduced the invasiveness of lung fibroblasts in the majority of subjects tested. These studies provide insights into the functional role of vimentin, which regulates autophagy and restricts the invasiveness of lung fibroblasts.

Authors

Ranu Surolia, Fu Jun Li, Zheng Wang, Huashi Li, Kevin Dsouza, Vinoy Thomas, Sergey Mirov, Dolores Pérez-Sala, Mohammad Athar, Victor J. Thannickal, Veena B. Antony

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Figure 5

Inhibition of VimIF assembly induces autophagic clearance of collagen type I in IPF fibroblasts.

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Inhibition of VimIF assembly induces autophagic clearance of collagen ty...
(A) Immunoblot analysis for vimentin and collagen type I (Col I) on cell lysates of IPF fibroblasts treated with WFA for different durations. β-Actin served as loading control. Data are representative of 3 experiments. (B) Immunostaining for Col I in IPF fibroblasts treated with vehicle (control) or WFA. Nuclei were costained with DAPI. Scale bars: 20 μm. (C) IPF fibroblasts were treated with vehicle (control) or WFA, and the presence of Col I in autophagosomes was assessed by indirect immunofluorescence and visualization of Col I– and LC3B-positive structures by immunofluorescence microscopy. Scale bars: 20 μm. Inset box shows magnified area of interest. (D) Measurement of invasiveness (percent zone of invasion) in IPF patient pulmospheres after treatment with an autophagy inhibitor (chloroquine [CQ]) or autophagy inducer (rapamycin [Rapa.]). Each dot represents an individual patient. *P < 0.05, **P < 0.01, as compared with control. (E and F) Immunoblot of Col I siRNA– transfected (siCOL1) human fibroblasts and measurement of invasiveness (zone of invasion percent) in IPF pulmospheres after siCOL1 transfection in the presence or absence of WFA. **P < 0.005, ***P < 0.001. (G) Fibroblasts from IPF subjects were treated with WFA to evaluate the expression of Col I. β-Actin served as loading control. (H) Quantification of soluble collagen in the supernatants of IPF fibroblasts treated with vehicle (control) or WFA. *P < 0.05, compared with vehicle-treated fibroblasts.