The circadian clock protein REVERBα inhibits pulmonary fibrosis development

Peter Cunningham ,Martin K Rutter ,Neil C Henderson ,Hannah J Durrington ,Lee A Borthwick ,Gareth Kitchen ,Boris Hinz ,Andrew J Fisher ,Julie Gibbs ,Paul F Mercer ,Rachel C Chambers ,James Bagnall ,David Ray ,Simon Anderson ,P J J De Meijer ,Rajamiyer Venkateswaran ,Nicola Begley ,Rajesh Shah ,Alicja Nazgiewicz ,Karen Piper Hanley ,Monika Lodyga ,A S I Loudon ,David A Bechtold ,John Blaikley

doi:10.1073/pnas.1912109117

Abstract

Pulmonary inflammatory responses lie under circadian control; however, the importance of circadian mechanisms in the underlying fibrotic phenotype is not understood. Here, we identify a striking change to these mechanisms resulting in a gain of amplitude and lack of synchrony within pulmonary fibrotic tissue. These changes result from an infiltration of mesenchymal cells, an important cell type in the pathogenesis of pulmonary fibrosis. Mutation of the core clock protein REVERBα in these cells exacerbated the development of bleomycin-induced fibrosis, whereas mutation of REVERBα in club or myeloid cells had no effect on the bleomycin phenotype. Knockdown of REVERBα revealed regulation of the little-understood transcription factor TBPL1. Both REVERBα and TBPL1 altered integrinβ1 focal-adhesion formation, resulting in increased myofibroblast activation. The translational importance of our findings was established through analysis of 2 human cohorts. In the UK Biobank, circadian strain markers (sleep length, chronotype, and shift work) are associated with pulmonary fibrosis, making them risk factors. In a separate cohort, REVERBα expression was increased in human idiopathic pulmonary fibrosis (IPF) lung tissue. Pharmacological targeting of REVERBα inhibited myofibroblast activation in IPF fibroblasts and collagen secretion in organotypic cultures from IPF patients, thus suggesting that targeting of REVERBα could be a viable therapeutic approach.

Highlights

Pulmonary inflammatory responses lie under circadian control; the importance of circadian mechanisms in the underlying fibrotic phenotype is not understood
We have previously identified the lung as a highly circadian organ and that responses to environmental insults are regulated and shaped by the circadian clock
We analyzed mouse lung fibrosis, finding newly-emergent, and strong circadian oscillations driven by fibroblasts

Summary

Introduction

Pulmonary inflammatory responses lie under circadian control; the importance of circadian mechanisms in the underlying fibrotic phenotype is not understood. Knockdown of REVERBα revealed regulation of the little-understood transcription factor TBPL1 Both REVERBα and TBPL1 altered integrinβ focal-adhesion formation, resulting in increased myofibroblast activation. Pulmonary fibrosis involves other cell types, e.g., club cells [9] and macrophages [16], regulating the accumulation of fibroblasts and the deposition of the extracellular matrix As these cell types maintain autonomous circadian oscillations [2, 5], examination of circadian factors and mechanisms in the pulmonary fibrotic response is warranted. Disruption of the clock in these cells, which are not widely implicated in circadian pathophysiology, results in a profibrotic phenotype Translation of these findings in humans revealed previously unrecognized important circadian risk factors for pulmonary fibrosis (sleep length, chronotype, and shift work). The resulting 24-h oscillations in protein expression can be disrupted through environmental disruption (e.g., shift-work schedules) or genetic deletion of core clock components, producing inflammatory and metabolic phenotypes [5, 21, 22]

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