Christian Beyer1, Christoph Zenzmaier2, Katrin Palumbo-Zerr1, Rossella Mancuso1, Alfiya Distler1, Clara Dees1, Pawel Zerr1, Jingang Huang1, Christiane Maier1, Milena L Pachowsky3, Andreas Friebe4, Peter Sandner5, Oliver Distler6, Georg Schett1, Peter Berger7, Jörg H W Distler1. 1. Department of Internal Medicine 3 and Institute for Clinical Immunology, University of Erlangen-Nuremberg, Erlangen, Germany. 2. Institute for Biomedical Aging Research, University of Innsbruck, Innsbruck, Austria Department of Internal Medicine, Innsbruck Medical University, Innsbruck, Austria. 3. Department of Trauma and Orthopaedic Surgery, University Erlangen-Nuremberg, Erlangen, Germany. 4. Institute for Physiology, Julius-Maximilians-University Würzburg, Würzburg, Germany. 5. Bayer Health Care, Global Drug Discovery-Common Mechanism Research, Wuppertal, Germany Hannover Medical School, Institute of Pharmacology, Hannover, Germany. 6. Department of Rheumatology, University Hospital Zurich, Zürich, Switzerland. 7. Institute for Biomedical Aging Research, University of Innsbruck, Innsbruck, Austria.
Abstract
OBJECTIVES: We have previously described the antifibrotic role of the soluble guanylate cyclase (sGC). The mode of action, however, remained elusive. In the present study, we describe a novel link between sGC signalling and transforming growth factor β (TGFβ) signalling that mediates the antifibrotic effects of the sGC. METHODS: Human fibroblasts and murine sGC knockout fibroblasts were treated with the sGC stimulator BAY 41-2272 or the stable cyclic guanosine monophosphate (cGMP) analogue 8-Bromo-cGMP and stimulated with TGFβ. sGC knockout fibroblasts were isolated from sGCI(fl/fl) mice, and recombination was induced by Cre-adenovirus. In vivo, we studied the antifibrotic effects of BAY 41-2272 in mice overexpressing a constitutively active TGF-β1 receptor. RESULTS: sGC stimulation inhibited TGFβ-dependent fibroblast activation and collagen release. sGC knockout fibroblasts confirmed that the sGC is essential for the antifibrotic effects of BAY 41-2272. Furthermore, 8-Bromo-cGMP reduced TGFβ-dependent collagen release. While nuclear p-SMAD2 and 3 levels, SMAD reporter activity and transcription of classical TGFβ target genes remained unchanged, sGC stimulation blocked the phosphorylation of ERK. In vivo, sGC stimulation inhibited TGFβ-driven dermal fibrosis but did not change p-SMAD2 and 3 levels and TGFβ target gene expression, confirming that non-canonical TGFβ pathways mediate the antifibrotic sGC activity. CONCLUSIONS: We elucidated the antifibrotic mode of action of the sGC that increases cGMP levels, blocks non-canonical TGFβ signalling and inhibits experimental fibrosis. Since sGC stimulators have shown excellent efficacy and tolerability in phase 3 clinical trials for pulmonary arterial hypertension, they may be further developed for the simultaneous treatment of fibrosis and vascular disease in systemic sclerosis. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.
OBJECTIVES: We have previously described the antifibrotic role of the soluble guanylate cyclase (sGC). The mode of action, however, remained elusive. In the present study, we describe a novel link between sGC signalling and transforming growth factor β (TGFβ) signalling that mediates the antifibrotic effects of the sGC. METHODS:Human fibroblasts and murine sGC knockout fibroblasts were treated with the sGC stimulator BAY 41-2272 or the stable cyclic guanosine monophosphate (cGMP) analogue 8-Bromo-cGMP and stimulated with TGFβ. sGC knockout fibroblasts were isolated from sGCI(fl/fl) mice, and recombination was induced by Cre-adenovirus. In vivo, we studied the antifibrotic effects of BAY 41-2272 in mice overexpressing a constitutively active TGF-β1 receptor. RESULTS: sGC stimulation inhibited TGFβ-dependent fibroblast activation and collagen release. sGC knockout fibroblasts confirmed that the sGC is essential for the antifibrotic effects of BAY 41-2272. Furthermore, 8-Bromo-cGMP reduced TGFβ-dependent collagen release. While nuclear p-SMAD2 and 3 levels, SMAD reporter activity and transcription of classical TGFβ target genes remained unchanged, sGC stimulation blocked the phosphorylation of ERK. In vivo, sGC stimulation inhibited TGFβ-driven dermal fibrosis but did not change p-SMAD2 and 3 levels and TGFβ target gene expression, confirming that non-canonical TGFβ pathways mediate the antifibrotic sGC activity. CONCLUSIONS: We elucidated the antifibrotic mode of action of the sGC that increases cGMP levels, blocks non-canonical TGFβ signalling and inhibits experimental fibrosis. Since sGC stimulators have shown excellent efficacy and tolerability in phase 3 clinical trials for pulmonary arterial hypertension, they may be further developed for the simultaneous treatment of fibrosis and vascular disease in systemic sclerosis. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.
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