Celastrol inhibits pro-inflammatory cytokine secretion in Crohn’s disease biopsies

https://doi.org/10.1016/j.bbrc.2004.07.186Get rights and content

Abstract

Crohn’s disease is a chronic intestinal inflammatory process. In modern therapy, TNF-α inhibition is the main goal. The aim here is to characterize the effects of Celastrol, a pentacyclic-triterpene, on the secretion of inflammatory cytokines by LPS-activated human cells. Celastrol dose-dependently inhibited the secretion of all tested pro-inflammatory cytokines with IC50 in the nanomolar range. Effect not related to glucocorticoid receptor activity is shown by competition experiments with the steroid antagonist RU486. Celastrol inhibited the pro-inflammatory cytokine secretion from mucosal inflammatory biopsies from Crohn’s disease patients. Cytometry emphasized that for all tested pro-inflammatory cytokines, CD33+ cells are the most sensitive. Quantitative-PCR and confocal analysis on a human monocytic cell line indicated that Celastrol acts at the transcriptional level by inhibiting LPS-induced NF-κB translocation. Celastrol might be a putative anti-inflammatory drug in the treatment of inflammatory diseases, given its inhibition of cytokine production by intestinal biopsies from Crohn’s disease patients.

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Materials and methods

Reagents and compounds. β-Mercaptoethanol, dexamethasone, glutamine, Hanks’ balanced salt solution (HBSS), heparin, Histopaque-1077, lipopolysaccharide (LPS) from Salmonella abortus equi, mifepristone (RU486), MTT, RPMI 1640, and SigmaFast OPD substrate were purchased from Sigma–Aldrich (Saint-Quentin Fallavier, France). Celastrol was a generous gift of Dr. A.C. Allison (SurroMed, Mountain View, California, USA). Twenty-four-well tissue culture treated plates and 96-well polyvinyl chloride

Celastrol inhibits pro-inflammatory cytokine production by LPS-activated human peripheral blood mononuclear cells and human whole blood

Since various antioxidants have been described to potently inhibit production of several pro-inflammatory cytokines or chemokines, we measured Celastrol’s potential to inhibit IL-1β, -6, -8, and TNF-α production in vitro by human LPS-activated PBMCs. Celastrol inhibits in a dose-dependant way the production of these four cytokines, with IC50’s ranging from 3 to 30 nM (Fig. 1 and Table 1A). The similar activity profile is found for human whole blood, with, as expected, IC50 values approximately

Discussion

Celastrol has been previously described as a potent inhibitor of various inflammatory mediators (NO, IL-1β) in different in vitro models [20]. Here, we focused initially on four cytokines (IL-1β, IL-6, IL-8, and TNF-α) believed to play a key role in chronic immune diseases, particularly Crohn’s disease [8] and overproduced in various immune-mediated pathologies. We then enlarged our study to a panel of cytokines involved in the Th1/Th2 balance and to inflammatory intestinal mucosal biopsies

Acknowledgments

The authors thank Dr. Kenneth TAKEDA (UMR 7034 CNRS) for critical reviewing of the manuscript. Financial support: The Région Alsace, and FORENAP, FRANCE equally supported G.F. Pinna. Y. Arondel thanks the laboratoires FERRING for their financial assistance.

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