Neural stem cells are fundamental to development of the central nervous system (CNS)-as well as its plasticity and regeneration-and represent a potential tool for neuro transplantation therapy and research. This study is focused on examination of the proliferation dynamic and fate of embryonic neural stem cells (eNSCs) under differentiating conditions. In this work, we analyzed eNSCs differentiating alone and in the presence of sonic hedgehog (SHH) or triiodothyronine (T3) which play an important role in the development of the CNS. We found that inhibition of the SHH pathway and activation of the T3 pathway increased cellular health and survival of differentiating eNSCs. In addition, T3 was able to increase the expression of the gene for the receptor smoothened (Smo), which is part of the SHH signaling cascade, while SHH increased the expression of the T3 receptor beta gene (Thrb). This might be the reason why the combination of SHH and T3 increased the expression of the thyroxine 5-deiodinase type III gene (Dio3), which inhibits T3 activity, which in turn affects cellular health and proliferation activity of eNSCs.
- Klíčová slova
- cell differentiation, embryonic neural stem cells, sonic hedgehog, triiodothyronine,
- MeSH
- jodidperoxidasa genetika metabolismus MeSH
- kultivované buňky MeSH
- myší embryonální kmenové buňky cytologie metabolismus MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- nervové kmenové buňky cytologie metabolismus MeSH
- neurogeneze * MeSH
- proteiny hedgehog genetika metabolismus MeSH
- receptor Smoothened genetika metabolismus MeSH
- trijodthyronin metabolismus MeSH
- tyreoidální hormony, receptory beta genetika metabolismus MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- iodothyronine deiodinase type III MeSH Prohlížeč
- jodidperoxidasa MeSH
- proteiny hedgehog MeSH
- receptor Smoothened MeSH
- Shh protein, mouse MeSH Prohlížeč
- Smo protein, mouse MeSH Prohlížeč
- trijodthyronin MeSH
- tyreoidální hormony, receptory beta MeSH
The Hedgehog (Hh) signaling pathway is crucial for vertebrate embryonic development, tissue homeostasis and regeneration. Hh signaling is upregulated in basal cell carcinoma and medulloblastoma and Hh pathway inhibitors targeting the Smoothened (SMO) protein are in clinical use. However, the signaling cascade is incompletely understood and novel druggable proteins in the pathway are in high demand. We describe the discovery of the Hh-pathway modulator Pipinib by means of cell-based screening. Target identification and validation revealed that Pipinib selectively inhibits phosphatidylinositol 4-kinase IIIβ (PI4KB) and suppresses GLI-mediated transcription and Hh target gene expression by impairing SMO translocation to the cilium. Therefore, inhibition of PI4KB and, consequently, reduction in phosphatidyl-4-phosphate levels may be considered an alternative approach to inhibit SMO function and thus, Hedgehog signaling.
- Klíčová slova
- Hedgehog signaling, PI4KB, biological activity, inhibitors,
- MeSH
- buněčné linie MeSH
- cilie metabolismus MeSH
- exprese genu účinky léků MeSH
- fosfotransferasy s alkoholovou skupinou jako akceptorem antagonisté a inhibitory genetika metabolismus MeSH
- lidé MeSH
- malá interferující RNA metabolismus MeSH
- morfoliny farmakologie MeSH
- myši MeSH
- osteogeneze účinky léků MeSH
- proteiny hedgehog antagonisté a inhibitory genetika metabolismus MeSH
- protinádorové látky chemie farmakologie MeSH
- puriny farmakologie MeSH
- receptor Smoothened genetika metabolismus MeSH
- RNA interference MeSH
- signální transdukce účinky léků MeSH
- thiofeny chemie farmakologie MeSH
- vedlejší histokompatibilní antigeny genetika metabolismus MeSH
- viabilita buněk účinky léků MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Research Support, N.I.H., Extramural MeSH
- Názvy látek
- fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
- malá interferující RNA MeSH
- morfoliny MeSH
- phosphatidylinositol phosphate 4-kinase MeSH Prohlížeč
- proteiny hedgehog MeSH
- protinádorové látky MeSH
- puriny MeSH
- purmorphamine MeSH Prohlížeč
- receptor Smoothened MeSH
- thiofeny MeSH
- vedlejší histokompatibilní antigeny MeSH
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 TBRact-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.
- Klíčová slova
- Fibroblasts, Systemic Sclerosis, Treatment,
- 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 Gli2 s motivem zinkových prstů 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
- TGF-beta receptor I. typu 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
- Názvy látek
- anilidy MeSH
- GANT 61 MeSH Prohlížeč
- Gli2 protein, mouse MeSH Prohlížeč
- HhAntag691 MeSH Prohlížeč
- inhibitor aktivátoru plazminogenu 1 MeSH
- kolagen typu I MeSH
- messenger RNA MeSH
- protein Gli2 s motivem zinkových prstů MeSH
- protein Smad3 MeSH
- protein-serin-threoninkinasy MeSH
- proteiny hedgehog MeSH
- pteridiny MeSH
- pyridiny MeSH
- pyrimidiny MeSH
- receptor Smoothened MeSH
- receptory transformujícího růstového faktoru beta MeSH
- rekombinantní proteiny MeSH
- růstový faktor pojivové tkáně MeSH
- SD-208 MeSH Prohlížeč
- TGF-beta receptor I. typu MeSH
- transformující růstový faktor beta MeSH
- transkripční faktory Krüppel-like MeSH
