Targeting gut T cell Ca2+ release-activated Ca2+ channels inhibits T cell cytokine production and T-box transcription factor T-bet in inflammatory bowel disease

Antonio Di Sabatino, Laura Rovedatti, Rejbinder Kaur, Jonathan P Spencer, Jon T Brown, Valerie D Morisset, Paolo Biancheri, Nicholas A B Leakey, Jonathan I Wilde, Laurie Scott, Gino R Corazza, Kevin Lee, Neel Sengupta, Charles H Knowles, Martin J Gunthorpe, Peter G McLean, Thomas T MacDonald, Laurens Kruidenier

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84 Citations (Scopus)

Abstract

Prolonged Ca(2+) entry through Ca(2+) release-activated Ca(2+) (CRAC) channels is crucial in activating the Ca(2+)-sensitive transcription factor NFAT, which is responsible for directing T cell proliferation and cytokine gene expression. To establish whether targeting CRAC might counteract intestinal inflammation, we evaluated the in vitro effect of a selective CRAC inhibitor on T cell cytokine production and T-bet expression by lamina propria mononuclear cells (LPMC) and biopsy specimens from inflammatory bowel disease (IBD) patients. The inhibitory activity of the CRAC blocker was investigated through patch-clamp experiments on rat basophilic leukemia cells and fluorometric imaging plate reader intracellular Ca(2+) assays using thapsigargin-stimulated Jurkat T cells and its detailed selectivity profile defined using a range of in vitro radioligand binding and functional assays. Anti-CD3/CD28-stimulated LPMC and biopsy specimens from 51 patients with IBD were cultured with a range of CRAC inhibitor concentrations (0.01-10 microM). IFN-gamma, IL-2, IL-8, and IL-17 were analyzed by ELISA. T-bet was determined by immunoblotting. We found that the CRAC blocker concentration-dependently inhibited CRAC current in rat basophilic leukemia cells and thapsigargin-induced Ca(2+) influx in Jurkat T cells. A concentration-dependent reduction in T-bet expression and production of IFN-gamma, IL-2, IL-17, but not IL-8, was observed in IBD LPMC and biopsy specimens treated with the CRAC inhibitor. In conclusion, we provide evidence that the suppression of CRAC channel function may dampen the increased T cell response in the inflamed gut, thus suggesting a promising role for CRAC inhibitor drugs in the therapeutic management of patients with IBD.

Original languageEnglish
Pages (from-to)3454-3462
Number of pages9
JournalJournal of Immunology
Volume183
Issue number5
DOIs
Publication statusPublished - 1 Sep 2009

Keywords

  • Adult
  • Aged
  • Animals
  • Calcium Channel Blockers/pharmacology
  • Calcium Channels/metabolism
  • Cell Line, Tumor
  • Cytokines/antagonists & inhibitors
  • Humans
  • Inflammatory Bowel Diseases/immunology
  • Intestinal Mucosa/immunology
  • Jurkat Cells
  • Middle Aged
  • Organ Culture Techniques
  • Patch-Clamp Techniques
  • Rats
  • T-Box Domain Proteins/antagonists & inhibitors
  • T-Lymphocyte Subsets/immunology
  • Young Adult

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