OBJECTIVES: Hedgehog signalling plays a critical role during the pathogenesis of fibrosis in systemic sclerosis (SSc). Besides canonical hedgehog signalling with smoothened (SMO)-dependent activation of GLI transcription factors, GLI can be activated independently of classical hedgehog ligands and receptors (so-called non-canonical pathways). Here, we aimed to evaluate the role of non-canonical hedgehog signalling in SSc and to test the efficacy of direct GLI inhibitors that target simultaneously canonical and non-canonical hedgehog pathways. METHODS: The GLI inhibitor GANT-61 was used to inhibit canonical as well as non-canonical hedgehog signalling, while the SMO inhibitor vismodegib was used to selectively target canonical hedgehog signalling. Furthermore, GLI2 was selectively depleted in fibroblasts using the Cre-LoxP system. The effects of pharmacological or genetic of GLI2 on transforming growth factor-β (TGF-β) signalling were analysed in cultured fibroblasts, in bleomycin-induced pulmonary fibrosis and in mice with overexpression of a constitutively active TGF-β receptor I. RESULTS: TGF-β upregulated GLI2 in a Smad3-dependent manner and induced nuclear accumulation and DNA binding of GLI2. Fibroblast-specific knockout of GLI2 protected mice from TBR(act)-induced fibrosis. Combined targeting of canonical and non-canonical hedgehog signalling with direct GLI inhibitors exerted more potent antifibrotic effects than selective targeting of canonical hedgehog signalling with SMO inhibitors in experimental dermal and pulmonary fibrosis. CONCLUSIONS: Our data demonstrate that hedgehog pathways and TGF-β signalling both converge to GLI2 and that GLI2 integrates those signalling to promote tissue fibrosis. These findings may have translational implications as non-selective inhibitors of GLI2 are in clinical use and selective molecules are currently in development.

• MeSH
• anilidy farmakologie   MeSH
• dospělí MeSH
• fibroblasty účinky léků   metabolismus   MeSH
• fibróza MeSH
• genový knockout MeSH
• inhibitor aktivátoru plazminogenu 1 genetika   MeSH
• kolagen typu I genetika   MeSH
• kultivované buňky MeSH
• kůže účinky léků   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• messenger RNA metabolismus   MeSH
• mladý dospělý MeSH
• myši knockoutované MeSH
• myši transgenní MeSH
• myši MeSH
• plicní fibróza chemicky indukované   metabolismus   MeSH
• protein Smad3 metabolismus   MeSH
• protein-serin-threoninkinasy antagonisté a inhibitory   genetika   MeSH
• proteiny hedgehog metabolismus   MeSH
• pteridiny farmakologie   MeSH
• pyridiny farmakologie   MeSH
• pyrimidiny farmakologie   MeSH
• receptor Smoothened antagonisté a inhibitory   MeSH
• receptory transformujícího růstového faktoru beta antagonisté a inhibitory   genetika   MeSH
• rekombinantní proteiny farmakologie   MeSH
• růstový faktor pojivové tkáně genetika   MeSH
• senioři MeSH
• signální transdukce účinky léků   MeSH
• systémová sklerodermie genetika   metabolismus   MeSH
• transformující růstový faktor beta metabolismus   farmakologie   MeSH
• transkripční faktory Krüppel-like antagonisté a inhibitory   genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• myši MeSH
• senioři MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

OBJECTIVES: TWIST1 is a member of the class B of basic helix-loop-helix transcription factors that regulates cell lineage determination and differentiation and has been implicated in epithelial-to-mesenchymal transition. Here, we aimed to investigate the role of TWIST1 for the activation of resident fibroblasts in systemic sclerosis (SSc). METHODS: The expression of Twist1 in fibroblasts was modulated by forced overexpression or siRNA-mediated knockdown. Interaction of Twist1, E12 and inhibitor Of differentiation (Id) was analysed by co-immunoprecipitation. The role of Twist1 in vivo was evaluated using inducible, conditional knockout mice with either ubiquitous or fibroblast-specific depletion of Twist1. Mice were either challenged with bleomycin or overexpressing a constitutively active transforming growth factor (TGF)β receptor I. RESULT: The expression of TWIST1 was increased in fibroblasts in fibrotic human and murine skin in a TGFβ/SMAD3-dependent manner. TWIST1 in turn enhanced TGFβ-induced fibroblast activation in a p38-dependent manner. The stimulatory effects of TWIST1 on resident fibroblasts were mediated by TWIST1 homodimers. TGFβ promotes the formation of TWIST1 homodimers by upregulation of TWIST1 and by induction of inhibitor of DNA-binding proteins, which have high affinity for E12/E47 and compete against TWIST1 for E12/E47 binding. Mice with selective depletion of Twist1 in fibroblasts are protected from experimental skin fibrosis in different murine models to a comparable degree as mice with ubiquitous depletion of Twist1. CONCLUSIONS: Our data identify TWIST1 as a central pro-fibrotic factor in SSc, which facilitates fibroblast activation by amplifying TGFβ signalling. Targeting of TWIST1 may thus be a novel approach to normalise aberrant TGFβ signalling in SSc.

• MeSH
• fibroblasty účinky léků   metabolismus   MeSH
• genový knockdown MeSH
• jaderné proteiny biosyntéza   nedostatek   genetika   metabolismus   fyziologie   MeSH
• kůže patologie   MeSH
• lidé MeSH
• malá interferující RNA genetika   MeSH
• messenger RNA genetika   MeSH
• multimerizace proteinu fyziologie   MeSH
• myši knockoutované MeSH
• regulace genové exprese účinky léků   fyziologie   MeSH
• signální transdukce fyziologie   MeSH
• studie případů a kontrol MeSH
• systémová sklerodermie metabolismus   patologie   MeSH
• transformující růstový faktor beta farmakologie   MeSH
• transkripční faktor Twist biosyntéza   nedostatek   genetika   metabolismus   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH