Abstract

Idiopathic pulmonary fibrosis is characterized by excessive deposition of collagen in the lung, leading to chronically impaired gas exchange and death1,2,3. Oxidative stress is believed to be critical in this disease pathogenesis4,5,6, although the exact mechanisms remain enigmatic. Protein S-glutathionylation (PSSG) is a post-translational modification of proteins that can be reversed by glutaredoxin-1 (GLRX)7. It remains unknown whether GLRX and PSSG play a role in lung fibrosis. Here, we explored the impact of GLRX and PSSG status on the pathogenesis of pulmonary fibrosis, using lung tissues from subjects with idiopathic pulmonary fibrosis, transgenic mouse models and direct administration of recombinant Glrx to airways of mice with existing fibrosis. We demonstrate that GLRX enzymatic activity was strongly decreased in fibrotic lungs, in accordance with increases in PSSG. Mice lacking Glrx were far more susceptible to bleomycin- or adenovirus encoding active transforming growth factor beta-1 (AdTGFB1)-induced pulmonary fibrosis, whereas transgenic overexpression of Glrx in the lung epithelium attenuated fibrosis. We furthermore show that endogenous GLRX was inactivated through an oxidative mechanism and that direct administration of the Glrx protein into airways augmented Glrx activity and reversed increases in collagen in mice with TGFB1- or bleomycin-induced fibrosis, even when administered to fibrotic, aged animals. Collectively, these findings suggest the therapeutic potential of exogenous GLRX in treating lung fibrosis.

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Acknowledgements

This work was funded by NIH R35HL135828, R01HL079331, R01HL060014 and R03HL095404 (Y.M.W.J.-H.), an ATS unrestricted grant and NIH R01HL122383 (V.A.), Parker B. Francis fellowships (V.A. and J.L.J.v.d.V.), NIH R01HL05646 (A.v.d.V.), T32HL076122 (D.H.M. and J.T.J.), NCRR 1S10RR019246 (D.J.T.), American Heart Association 16GRNT27660006, NIH R01DK103750 and 1UL1TR001430 (M.M.B.), NIH R01HL133013 and R03 AG051857 (R.M.), P30GM103532 (C.G.I.), FAMRI Young Clinical Scientist Award 113393 and 1K01HL125474-01 (J.L.G.) and U01HL108642 and RC2 HL101715 (N.K.). Analysis of biological specimens and data provided by the Lung Tissue Research Consortium (LTRC) was supported by R03HL095404 (Y.M.W.J.-H.). We would like to thank J. Whitsett (University of Cincinnati) for generously providing the CCSP-rtTA, TetOP-Cre mice, J. Gauldie for providing AdTGFB1, N. Bishop for help with imaging of GLRX and collagen, J. M. Siddesha for technical support and P. Vacek for statistical support.

Author information

Affiliations

  1. Department of Pathology and Laboratory Medicine, University of Vermont, Burlington, VT, USA

    • Vikas Anathy
    • , Karolyn G. Lahue
    • , Shi B. Chia
    • , Dylan T. Casey
    • , Reem Aboushousha
    • , Jos L. J. van der Velden
    • , Evan Elko
    • , Sidra M. Hoffman
    • , David H. McMillan
    • , Jane T. Jones
    • , James D. Nolin
    • , Sarah Abdalla
    • , Robert Schneider
    • , David J. Seward
    • , Kelly J. Butnor
    • , Douglas. J. Taatjes
    • , Albert van der Vliet
    •  & Yvonne M. W. Janssen-Heininger
  2. Department of Medicine, University of Vermont, Burlington, VT, USA

    • David G. Chapman
    • , Ralph C. Budd
    •  & Charles G. Irvin
  3. Honors College, University of Vermont, Burlington, VT, USA

    • Elle C. Roberson
  4. Department of Chemistry, University of Vermont, Burlington, VT, USA

    • Matthew D. Liptak
    •  & Morgan E. Cousins
  5. Institute of Environmental Health Sciences, Wayne State University, Detroit, MI, USA

    • Ye-Shih Ho
  6. Department of Medical Biophysics and Immunology, University of Toronto, and the Ontario Cancer Institute/University Health Network, Toronto, Ontario, Canada

    • Razq Hakem
  7. Department of Medicine, Pulmonary, Critical Care and Sleep Section, National Jewish Health and the University of Colorado, Denver, CO, USA

    • Kevin K. Brown
  8. Department of Medicine, Boston University, Boston, MA, USA

    • Reiko Matsui
    •  & Markus M. Bachschmid
  9. Department of Medicine, Yale School of Medicine, New Haven, CT, USA

    • Jose L. Gomez
    •  & Naftali Kaminski

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Contributions

V.A., K.G.L., S.M.H., D.H.M., J.T.J., E.E., J.D.N., D.T.C., J.L.J.v.d.V. and R.S. performed the animal experiments. S.A., S.B.C. and K.G.L. prepared and tested recombinant GLRX; V.A., S.M.H., E.C.R., R.A. and K.G.L. performed S-glutathionylation and glutaredoxin assays; K.G.L. and D.T.C. performed fibrosis analyses; K.J.B. reviewed histopathology; D.J.T. provided critical suggestions regarding imaging and image analysis. Y.-S.H. generated Glrx−/− mice and helped with data interpretation; R.A. and S.B.C. performed collagenolytic activity assays; M.D.L. and M.E.C. performed and analyzed CD studies; R.H. generated Casp8loxP/loxP mice and helped with data interpretation. R.M. and M.M.B. conducted the ageing study and participated in data analyses. A.v.d.V. and R.C.B. supported the studies with key experimental suggestions and helped interpret data. D.J.S., D.G.C., C.G.I., K.J.B., J.L.G., N.K. and K.K.B. evaluated and analyzed clinical data. Y.M.W.J.-H. and V.A. designed the research, interpreted the data and wrote the manuscript.

Competing interests

Y.M.W.J.-H. and V.A. hold patents entitled: ‘Treatments involving glutaredoxins and similar agents’, ‘Treatments of oxidative stress conditions’ (US patent no. 8,679,811 and 9,907,828; Y.M.W.J.-H. and V.A.) and ‘Detection of glutathionylated proteins’ (US patent no. 8,877,447; Y.M.W.J.-H.).

Corresponding author

Correspondence to Yvonne M. W. Janssen-Heininger.

Supplementary information

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  2. Reporting Summary

  3. Supplementary Dataset

    F, t and df values

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https://doi.org/10.1038/s41591-018-0090-y