Celastrol blocks binding of lipopolysaccharides to a Toll-like receptor4/myeloid differentiation factor2 complex in a thiol-dependent manner.

Abstract

ETHNOPHARMACOLOGICAL RELEVANCE:

Tripterygium wilfordii (lei gong teng; Thunder of God Vine), which belongs to the Celastraceae family, has long been used in traditional Chinese medicine to treat inflammation and rheumatoid arthritis. Celastrol is a bioactive compound isolated from T. wilfordii.

AIM OF THE STUDY:

We investigated whether celastrol suppressed binding of lipopolysaccharides (LPS) to myeloid differentiation factor 2 (MD2), thereby downregulating Toll-like receptor4 (TLR4) activation in mouse primary macrophages.

MATERIALS AND METHODS:

Cytokine expression was determined by polymerase chain reaction analysis and enzyme-linked immunosorbent assay in bone marrow-derived primary macrophages (BMDMs). The kinase activity of tank-binding kinase 1 (TBK1) was examined by a luciferase reporter assay and an in vitro kinase assay. LPS binding to MD2 was examined by an in vitro binding assay and confocal microscopy analysis.

RESULTS:

Celastrol reduced LPS-induced expression of inflammatory cytokines, such as tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-12, and IL-1β, at both the mRNA and protein levels in BMDMs. Celastrol suppressed LPS binding to MD2, as shown by the in vitro binding assay, whereas it did not inhibit TBK1. In addition, co-localization of LPS with MD2 in BMDMs was blocked by celastrol. The inhibitory effects of celastrol on LPS binding to MD2 were reversed by thiol donors (N-acetyl-L-cysteine and dithiothreitol), suggesting that the thiol reactivity of celastrol contributes to its inhibitory effects on TLR4 activation in macrophages.

CONCLUSION:

Our results demonstrate that celastrol suppresses TLR4 activation through the inhibition of LPS binding to the TLR4/MD2 complex. These results provide a novel mechanism of action by which celastrol contributes to the anti-inflammatory activity of T. wilfordii.

• Authors: Jin Young Lee, Byung Ho Lee, Nam Doo Kim, Joo Young Lee
• Journal: Journal of ethnopharmacology (J Ethnopharmacol) Vol. 172 Pg. 254-60 (Aug 22 2015) ISSN: 1872-7573 [Electronic] Ireland
• PMID: 26116162 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Chemical References

• Anti-Inflammatory Agents
• Cytokines
• Lipopolysaccharides
• Ly96 protein, mouse
• Lymphocyte Antigen 96
• Pentacyclic Triterpenes
• Sulfhydryl Compounds
• Toll-Like Receptor 4
• Triterpenes
• celastrol
• Dithiothreitol
• Acetylcysteine

Topics

• Acetylcysteine (pharmacology)
• Animals
• Anti-Inflammatory Agents (isolation & purification, pharmacology)
• Cytokines (metabolism)
• Dithiothreitol (pharmacology)
• Enzyme-Linked Immunosorbent Assay
• Lipopolysaccharides (metabolism)
• Lymphocyte Antigen 96 (metabolism)
• Macrophages (drug effects, metabolism)
• Medicine, Chinese Traditional
• Mice
• Mice, Inbred C57BL
• Pentacyclic Triterpenes
• Polymerase Chain Reaction
• Sulfhydryl Compounds (chemistry)
• Toll-Like Receptor 4 (metabolism)
• Tripterygium (chemistry)
• Triterpenes (isolation & purification, pharmacology)