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Pseudohypoxic HIF pathway activation dysregulates collagen structure-function in human lung fibrosis

Pseudohypoxic HIF pathway activation dysregulates collagen structure-function in human lung fibrosis
Pseudohypoxic HIF pathway activation dysregulates collagen structure-function in human lung fibrosis
Extracellular matrix (ECM) stiffening with downstream activation of mechanosensitive pathways is strongly implicated in fibrosis. We previously reported that altered collagen nanoarchitecture is a key determinant of pathogenetic ECM structure-function in human fibrosis (Jones et al., 2018). Here, through human tissue, bioinformatic and ex vivo studies we provide evidence that hypoxia-inducible factor (HIF) pathway activation is a critical pathway for this process regardless of the oxygen status (pseudohypoxia). Whilst TGFβ increased rate of fibrillar collagen synthesis, HIF pathway activation was required to dysregulate post-translational modification of fibrillar collagen, promoting pyridinoline cross-linking, altering collagen nanostructure, and increasing tissue stiffness. In vitro, knockdown of Factor Inhibiting HIF (FIH), which modulates HIF activity, or oxidative stress caused pseudohypoxic HIF activation in normal fibroblasts. By contrast, endogenous FIH activity was reduced in fibroblasts from patients with lung fibrosis in association with significantly increased normoxic HIF pathway activation. In human lung fibrosis tissue, HIF mediated signalling was increased at sites of active fibrogenesis whilst subpopulations of human lung fibrosis mesenchymal cells had increases in both HIF and oxidative stress scores. Our data demonstrate that oxidative stress can drive pseudohypoxic HIF pathway activation which is a critical regulator of pathogenetic collagen structure-function in fibrosis.
Collagen, Lung, cell biology, fibrosis, human
2050-084X
Brereton, Christopher, J
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Yao, Liudi
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Davies, Elizabeth R.
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Zhou, Yilu
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Vukmirovic, Milica
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Bell, Joseph
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Wang, Siyuan
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Ridley, Robert
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Dean, Lareb
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Andriotis, Orestis
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Conforti, Franco
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Brewitz, Lennart
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Mohammed, Soran
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Wallis, Timothy
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Tavassoli, Ali
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Ewing, Robert
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Alzetani, Aiman
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Marshall, Benjamin
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Fletcher, Sophie V.
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Thurner, P.J.
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Fabre, Aurelie
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Kaminski, Naftali
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Richeldi, Luca
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Bhaskar, Atul
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Schofield, Christopher J.
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Loxham, Matthew
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Davies, Donna
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Wang, Yihua
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Jones, Mark
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Brereton, Christopher, J
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Yao, Liudi
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Davies, Elizabeth R.
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Zhou, Yilu
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Vukmirovic, Milica
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Bell, Joseph
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Wang, Siyuan
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Ridley, Robert
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Dean, Lareb
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Andriotis, Orestis
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Conforti, Franco
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Brewitz, Lennart
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Mohammed, Soran
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Wallis, Timothy
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Tavassoli, Ali
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Ewing, Robert
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Alzetani, Aiman
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Marshall, Benjamin
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Fletcher, Sophie V.
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Thurner, P.J.
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Fabre, Aurelie
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Kaminski, Naftali
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Richeldi, Luca
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Bhaskar, Atul
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Schofield, Christopher J.
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Loxham, Matthew
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Davies, Donna
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Wang, Yihua
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Jones, Mark
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Brereton, Christopher, J, Yao, Liudi, Davies, Elizabeth R., Zhou, Yilu, Vukmirovic, Milica, Bell, Joseph, Wang, Siyuan, Ridley, Robert, Dean, Lareb, Andriotis, Orestis, Conforti, Franco, Brewitz, Lennart, Mohammed, Soran, Wallis, Timothy, Tavassoli, Ali, Ewing, Robert, Alzetani, Aiman, Marshall, Benjamin, Fletcher, Sophie V., Thurner, P.J., Fabre, Aurelie, Kaminski, Naftali, Richeldi, Luca, Bhaskar, Atul, Schofield, Christopher J., Loxham, Matthew, Davies, Donna, Wang, Yihua and Jones, Mark (2022) Pseudohypoxic HIF pathway activation dysregulates collagen structure-function in human lung fibrosis. eLife, 11, [e69348]. (doi:10.7554/eLife.69348).

Record type: Article

Abstract

Extracellular matrix (ECM) stiffening with downstream activation of mechanosensitive pathways is strongly implicated in fibrosis. We previously reported that altered collagen nanoarchitecture is a key determinant of pathogenetic ECM structure-function in human fibrosis (Jones et al., 2018). Here, through human tissue, bioinformatic and ex vivo studies we provide evidence that hypoxia-inducible factor (HIF) pathway activation is a critical pathway for this process regardless of the oxygen status (pseudohypoxia). Whilst TGFβ increased rate of fibrillar collagen synthesis, HIF pathway activation was required to dysregulate post-translational modification of fibrillar collagen, promoting pyridinoline cross-linking, altering collagen nanostructure, and increasing tissue stiffness. In vitro, knockdown of Factor Inhibiting HIF (FIH), which modulates HIF activity, or oxidative stress caused pseudohypoxic HIF activation in normal fibroblasts. By contrast, endogenous FIH activity was reduced in fibroblasts from patients with lung fibrosis in association with significantly increased normoxic HIF pathway activation. In human lung fibrosis tissue, HIF mediated signalling was increased at sites of active fibrogenesis whilst subpopulations of human lung fibrosis mesenchymal cells had increases in both HIF and oxidative stress scores. Our data demonstrate that oxidative stress can drive pseudohypoxic HIF pathway activation which is a critical regulator of pathogenetic collagen structure-function in fibrosis.

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Accepted/In Press date: 4 January 2022
Published date: 21 February 2022
Additional Information: Acknowledgements This project was supported by Medical Research Council (MR/S025480/1), the Wellcome Trust (100638/Z/12/Z), an Academy of Medical Sciences/the Wellcome Trust Springboard Award [SBF002\1,038], and the AAIR Charity. CJB and LSND acknowledge the support of the NIHR Southampton Biomedical Research Centre. LY was supported by China Scholarship Council. YZ was supported by an Institute for Life Sciences PhD Studentship. FC was supported by Medical Research Foundation [MRF-091–0003-RG-CONFO]. ML was supported by a BBSRC Future Leader Fellowship [BB/PO11365/1] and a NIHR Southampton Biomedical Research Centre Senior Research Fellowship. We thank Carine Fixmer, Maria Lane, Benjamin Johnson, and the nurses of the Southampton Biomedical Research Unit for their help in the collection of human samples, supported by the Wessex Clinical Research Network and the National Institute of Health Research, UK. We also thank Dr. Tammie Bishop (University of Oxford) for her technical support in IHC and Prof Sir Peter Ratcliffe (University of Oxford) for the FIH antibody (mouse monoclonal 162 C) and the UAS-luc/GAL4DBD-HIF1αCAD binary reporter system.
Keywords: Collagen, Lung, cell biology, fibrosis, human

Identifiers

Local EPrints ID: 454167
URI: http://eprints.soton.ac.uk/id/eprint/454167
ISSN: 2050-084X
PURE UUID: c2256124-1a11-4a51-b795-f63e20ff5cdb
ORCID for Lareb Dean: ORCID iD orcid.org/0000-0002-8703-9236
ORCID for Soran Mohammed: ORCID iD orcid.org/0000-0002-3882-6129
ORCID for Timothy Wallis: ORCID iD orcid.org/0000-0001-7936-9764
ORCID for Ali Tavassoli: ORCID iD orcid.org/0000-0002-7420-5063
ORCID for Robert Ewing: ORCID iD orcid.org/0000-0001-6510-4001
ORCID for Matthew Loxham: ORCID iD orcid.org/0000-0001-6459-538X
ORCID for Donna Davies: ORCID iD orcid.org/0000-0002-5117-2991
ORCID for Yihua Wang: ORCID iD orcid.org/0000-0001-5561-0648
ORCID for Mark Jones: ORCID iD orcid.org/0000-0001-6308-6014

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Date deposited: 01 Feb 2022 17:48
Last modified: 19 Oct 2022 01:59

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Contributors

Author: Christopher, J Brereton
Author: Liudi Yao
Author: Elizabeth R. Davies
Author: Yilu Zhou
Author: Milica Vukmirovic
Author: Joseph Bell
Author: Siyuan Wang
Author: Robert Ridley
Author: Lareb Dean ORCID iD
Author: Orestis Andriotis
Author: Franco Conforti
Author: Lennart Brewitz
Author: Soran Mohammed ORCID iD
Author: Timothy Wallis ORCID iD
Author: Ali Tavassoli ORCID iD
Author: Robert Ewing ORCID iD
Author: Aiman Alzetani
Author: Sophie V. Fletcher
Author: P.J. Thurner
Author: Aurelie Fabre
Author: Naftali Kaminski
Author: Luca Richeldi
Author: Atul Bhaskar
Author: Christopher J. Schofield
Author: Matthew Loxham ORCID iD
Author: Donna Davies ORCID iD
Author: Yihua Wang ORCID iD
Author: Mark Jones ORCID iD

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