title: Selective non-steroidal glucocorticoid receptor agonists attenuate inflammation but do not impair intestinal epithelial cell restitution in vitro.
abstract: BACKGROUND
  Despite the excellent anti-inflammatory and immunosuppressive action of glucocorticoids (GCs), their use for the treatment of inflammatory bowel disease (IBD) still carries significant risks in terms of frequently occurring severe side effects, such as the impairment of intestinal tissue repair. The recently-introduced selective glucocorticoid receptor (GR) agonists (SEGRAs) offer anti-inflammatory action comparable to that of common GCs, but with a reduced side effect profile.
  
  
  METHODS
  The in vitro effects of the non-steroidal SEGRAs Compound A (CpdA) and ZK216348, were investigated in intestinal epithelial cells and compared to those of Dexamethasone (Dex). GR translocation was shown by immunfluorescence and Western blot analysis. Trans-repressive effects were studied by means of NF-κB/p65 activity and IL-8 levels, trans-activation potency by reporter gene assay. Flow cytometry was used to assess apoptosis of cells exposed to SEGRAs. The effects on IEC-6 and HaCaT cell restitution were determined using an in vitro wound healing model, cell proliferation by BrdU assay. In addition, influences on the TGF-β- or EGF/ERK1/2/MAPK-pathway were evaluated by reporter gene assay, Western blot and qPCR analysis.
  
  
  RESULTS
  Dex, CpdA and ZK216348 were found to be functional GR agonists. In terms of trans-repression, CpdA and ZK216348 effectively inhibited NF-κB activity and IL-8 secretion, but showed less trans-activation potency. Furthermore, unlike SEGRAs, Dex caused a dose-dependent inhibition of cell restitution with no effect on cell proliferation. These differences in epithelial restitution were TGF-β-independent but Dex inhibited the EGF/ERK1/2/MAPK-pathway important for intestinal epithelial wound healing by induction of MKP-1 and Annexin-1 which was not affected by CpdA or ZK216348.
  
  
  CONCLUSIONS
  Collectively, our results indicate that, while their anti-inflammatory activity is comparable to Dex, SEGRAs show fewer side effects with respect to wound healing. The fact that SEGRAs did not have a similar effect on cell restitution might be due to a different modulation of EGF/ERK1/2 MAPK signalling.
authors.affiliation: Institute of Pharmaceutical Chemistry, Goethe University Frankfurt/Main, Campus Riedberg, Frankfurt/Main, Germany.
authors.forename: Kerstin C
authors.initials: KC
authors.lastname: Reuter
authors.forename: Stefan M
authors.initials: SM
authors.lastname: Loitsch
authors.forename: Axel U
authors.initials: AU
authors.lastname: Dignass
authors.forename: Dieter
authors.initials: D
authors.lastname: Steinhilber
authors.forename: Jürgen
authors.initials: J
authors.lastname: Stein
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000009:adverse effects* / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects; D001572:Benzofurans / Q000009:adverse effects / Q000494:pharmacology / Q000627:therapeutic use; D048588:Benzoxazines / Q000009:adverse effects / Q000494:pharmacology / Q000627:therapeutic use; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000378:metabolism; D020868:Gene Silencing / Q000187:drug effects; D006801:Humans; D007249:Inflammation / Q000188:drug therapy; D007413:Intestinal Mucosa / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000819:agonists* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000378:metabolism; D014945:Wound Healing / Q000187:drug effects
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D001572:Benzofurans; D048588:Benzoxazines; D011965:Receptors, Glucocorticoid; D016212:Transforming Growth Factor beta; C481822:ZK 216348; D004815:Epidermal Growth Factor; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3