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Triptolide inhibits HuR‐dependent TNF‐α and IL‐6 mRNA stabilization and protein expression in Raw264.7 cells (657.6)
Author(s) -
Jiang Zhenzhou,
Zhang Luyong
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.657.6
Subject(s) - triptolide , messenger rna , microbiology and biotechnology , chemistry , cytoplasm , tumor necrosis factor alpha , biology , biochemistry , apoptosis , gene , immunology
Triptolide is a diterpenoid isolated from the plant Tripterygium wilfordii Hook. F and has been used for the treatment of a variety of autoimmune and inflammatory diseases including rheumatoid arthritis. In the present study, triptolide was found to inhibit LPS‐induced TNF‐α and IL‐6 expression in Raw 264.7 cells. The aim of this study was to determine if the RNA‐binding protein HuR plays a key role in triptolide regulating on TNF‐α and IL‐6 expressions. Methods: The mRNA degradation of TNF‐α and IL‐6 was analyzed by real‐time PCR in Actinomycin D assay. The HuR shuttling from nucleus to cytoplasm was detected by Immunofluorescence staining. Immunoprecipitation of HuR‐mRNA complexes was performed to assess the association of endogenous HuR with TNF‐α and IL‐6 mRNAs. The effect of triptolide on TNF‐α and IL‐6 3’‐ untranslated region (UTR) was detected by GFP reporter gene assay. Results : Triptolide decreased the half‐life of TNF‐α and IL‐6 mRNA dramatically and it inhibited 3’‐UTR fluorescence reporter gene activity. Meanwhile, triptolide inhibited the HuR shuttling from nucleus to cytoplasm. After triptolide treatment, decreased TNF‐α and IL‐6 mRNA in pull‐down experiments with anti‐HuR antibodies was observed, indicating that the decreased cytoplasmic HuR is responsible for the decreased mRNA. Conclusions: HuR might have a key impact on anti‐inflammatory and anti‐cancer effects of triptolide.