ObjectivesCoactivators are a heterogeneous family of transcriptional regulators that are essential for modulation of transcriptional outcomes and fine-tune numerous cellular processes. The aim of the present study was to evaluate the role of the coactivator peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in the pathogenesis of systemic sclerosis (SSc).MethodsExpression of PGC-1α was analysed by real-time PCR, western blot and immunofluorescence. Modulation of autophagy was analysed by reporter studies by expression of autophagy-related genes. The effects of PGC-1α knockdown on collagen production and myofibroblast differentiation were analysed in cultured human fibroblasts and in two mouse models with fibroblast-specific knockout of PGC-1α.ResultsThe expression of PGC-1α was induced in dermal fibroblasts of patients with SSc and experimental murine fibrosis. Transforming growth factor beta (TGFβ), hypoxia and epigenetic mechanisms regulate the expression of PGC-1α in fibroblasts. Knockdown of PGC-1α prevented the activation of autophagy by TGFβ and this translated into reduced fibroblast-to-myofibroblast differentiation and collagen release. Knockout of PGC-1α in fibroblasts prevented skin fibrosis induced by bleomycin and by overexpression of a constitutively active TGFβ receptor type I. Moreover, pharmacological inhibition of PGC-1α by SR18292 induced regression of pre-established, bleomycin-induced skin fibrosis.ConclusionPGC-1α is upregulated in SSc and promotes autophagy to foster TGFβ-induced fibroblast activation. Targeting of PGC-1α prevents aberrant autophagy, inhibits fibroblast activation and tissue fibrosis and may over therapeutic potential.

Chronic graft-versus-host disease (cGvHD) is a major life-threatening complication of allogeneic hematopoietic stem cell transplantation. The molecular mechanisms underlying cGvHD remain poorly understood and targeted therapies are not well established for clinical use. Here, we examined the role of the canonical WNT pathway in sclerodermatous cGvHD (sclGvHD). WNT signaling was activated in human sclGvHD with increased nuclear accumulation of the transcription factor β-catenin and WNT-biased gene expression signature in lesional skin. Treatment with highly selective tankryase inhibitor G007-LK, CK1α agonist pyrvinium or LRP6 inhibitor salinomycin, abrogated the activation of WNT signaling and protected against experimental cGvHD, without significant impact on graft-versus-leukemia effect (GvL). Treatment with G007-LK, pyrvinium or salinomycin almost completely prevented the development of clinical and histological features in the B10.D2 (H-2d)→BALB/c (H-2d) and in the LP/J (H-2b)→C57BL/6 (H-2b) model of sclGvHD. Inhibition of canonical WNT signaling reduced the release of extracellular matrix from fibroblasts and reduced leukocyte influx, suggesting that WNT signaling stimulates fibrotic tissue remodeling by direct effects on fibroblasts and by indirect, inflammation-dependent effects in sclGvHD. Our findings may have direct translational potential, as pyrvinium is in clinical use and tankyrase inhibitors are in clinical trials for other implications.

Background:Peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) is the best studied member of the family of coactivators. PGC-1α was initially identified through its interaction with PPARγ in brown adipose tissue. Recent evidence further indicates that PGC-1α may also modulate the transcription of autophagy-related genes, which has recently been shown to be required for fibroblast-to-myofibroblast differentiation under fibrotic conditions. However, the role of PGC-1α in the pathogenesis of SSc has not been investigated.Objectives:The aim of the present study was to evaluate the role of the coactivator PGC-1α on autophagy and to evaluate its role in the pathologic activation of fibroblasts in SSc.Methods:Expression of PGC-1α was analyzed by RT-PCR, Western blot and immunofluorescence. Modulation of autophagy was analyzed by reporter studies by expression of autophagy related genes. The effects of PGC-1α knockdown on collagen production and myofibroblast differentiation were analyzed in cultured human fibroblasts and in two mouse models with fibroblast-specific knockout of PGC-1α.Results:PGC-1α overexpression was detected by immunohistochemistry in skin sections of SSc patients and in experimental fibrotic murine skin, particularly in fibroblasts. Knockdown of PGC-1α inhibited the stimulatory effects of TGFβ on fibroblast activation with impaired induction of collagen as compared to control fibroblasts. Fibroblasts specific knockout of PGC-1α ameliorates experimental fibrosis in bleomycin-induced and adTBR-induced murine dermal fibrosis with decreased dermal thickness, hydroxyproline and myofibroblast counts compared to wild-type fibrotic mice. Incubation of dermal fibroblasts with TGFβ activated autophagy in control fibroblasts with increased expression of the autophagy-related genes ATG7 and BECLIN-1, enhanced conversion of LC3 I to LC3 II and decreased ratios of ILC3 I EGFP to LC3 II RFP in LC3 reporter assays. The expression levels of ATG7, BECLIN-1 and ILC3 II of TGFβ-stimulated PGC-1α knockout fibroblasts decreased compare to TGFβ stimulated wild-type fibroblasts. The ratio of ILC3 I EGFP to LC3 II RFP of TGFβ-stimulated PGC-1α knockout fibroblasts in reporter assays were comparable to unstimulated fibroblasts.Conclusion:PGC-1α is upregulated in SSc and promotes autophagy to foster TGFβ-induced fibroblast activation. Targeting of PGC-1α prevents aberrant autophagy, inhibits fibroblast activation and tissue fibrosis.References:[1]Finck BN, Kelly DP. PGC-1 coactivators: inducible regulators of energy metabolism in health and disease. The Journal of clinical investigation. 2006 Mar; 116(3):615-622[2]Lindholm D, Eriksson O, Makela J, Belluardo N, Korhonen L. PGC-1alpha: a master gene that is hard to master. Cellular and molecular life sciences: CMLS. 2012 Aug; 69(15):2465-2468.[3]Li SY, Susztak K. The Role of Peroxisome Proliferator-Activated Receptor gamma Coactivator 1alpha (PGC-1alpha) in Kidney Disease. Semin Nephrol. 2018 Mar; 38(2):121-126.[4]Vainshtein A, Tryon LD, Pauly M, Hood DA. Role of PGC-1alpha during acute exercise-induced autophagy and mitophagy in skeletal muscle. American journal of physiology Cell physiology. 2015 May 1; 308(9):C710-719.[5]Zehender A LN, Stefanica A, Chen CW, Soare A, Wohlfahrt T, Rauber S, Bergmann C, Ramming A, Distler O, Schett G, Distler J. TGFβ Promotes Fibrosis By MYST1-Dependent Epigenetic Regulation of Autophagy [abstract]. Arthritis Rheumatol 2017; 69 (suppl 10).Disclosure of Interests:Yun Zhang: None declared, Katja Dreißigacker: None declared, Diana Distler: None declared, Andrea-Hermina Györfi: None declared, Christina Bergmann: None declared, xiang zhou: None declared, Lichong Shen: None declared, Ingo Ludolph: None declared, Raymund Horch: None declared, Andreas Ramming Grant/research support from: Pfizer, Novartis, Consultant of: Boehringer Ingelheim, Novartis, Gilead, Pfizer, Speakers bureau: Boehringer Ingelheim, Roche, Janssen, Georg Schett Speakers bureau: AbbVie, BMS, Celgene, Janssen, Eli Lilly, Novartis, Roche and UCB, Jörg Distler Grant/research support from: Boehringer Ingelheim, Consultant of: Boehringer Ingelheim, Paid instructor for: Boehringer Ingelheim, Speakers bureau: Boehringer Ingelheim