Department of Respiratory Disease, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China.
Respirology. 2013 Nov;18(8):1249-55. doi: 10.1111/resp.12145.
Triptolide, a type of diterpenoid, is the active compound of Tripterygium wilfordii; it plays roles in anti-inflammatory and immune response regulation. Our objective was to investigate the mechanism of the inhibitory effect of triptolide on interleukin-13 (IL-13) gene expression in activated T lymphocytes. Understanding the molecular mechanism by which triptolide exerts a therapeutic function may be useful in developing a pharmaceutical treatment for asthma.
Peripheral blood mononuclear cells (PBMC) and Hut-78 cells were stimulated with anti-CD3/CD28 with or without co-incubation with triptolide. The alteration of IL-13 messenger RNA (mRNA), expression and protein level were analysed using real-time reverse transcription polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay, respectively. The intracellular distribution profile of transcription factor GATA3 and nuclear factor of activated T cells (NFAT1) were analysed by Western blotting. The binding rates of GATA3 and NFAT1 to the promoter sequence of IL-13 were analysed by chromatin immunoprecipitation (ChIP) PCR.
In PBMC, the release of IL-13 was dependent on anti-CD3/CD28 stimulation. Its release could be inhibited by triptolide at the concentration of 500 nmol. In Hut-78 cells, IL-13 mRNA and protein expression were increased with anti-CD3/CD28 stimulation and significantly inhibited by incubation with 28 nmol triptolide. This concentration of triptolide also significantly inhibited the nuclear translocation of GATA3 and NFAT1 reducing the binding rate to the IL-13 gene promoter.
Triptolide inhibits IL-13 gene transcription and protein expression by inhibiting GATA3 and NFAT1 nuclear translocation and their binding rates to the IL-13 gene promoter region.
雷公藤红素是一种二萜类化合物，是雷公藤的活性成分，具有抗炎和免疫反应调节作用。本研究旨在探讨雷公藤红素抑制活化 T 淋巴细胞白细胞介素-13（IL-13）基因表达的作用机制。了解雷公藤红素发挥治疗作用的分子机制可能有助于开发治疗哮喘的药物。
外周血单个核细胞（PBMC）和 Hut-78 细胞用抗-CD3/CD28 刺激，或与雷公藤红素共同孵育。采用实时逆转录聚合酶链反应（PCR）和酶联免疫吸附试验（ELISA）分别分析 IL-13 信使 RNA（mRNA）、表达和蛋白水平的变化。采用 Western blot 分析转录因子 GATA3 和活化 T 细胞核因子（NFAT1）的细胞内分布。采用染色质免疫沉淀（ChIP）PCR 分析 GATA3 和 NFAT1 与 IL-13 启动子序列的结合率。
在 PBMC 中，IL-13 的释放依赖于抗-CD3/CD28 刺激，其释放可被 500 nmol 雷公藤红素抑制。在 Hut-78 细胞中，抗-CD3/CD28 刺激可增加 IL-13 mRNA 和蛋白表达，28 nmol 雷公藤红素孵育可显著抑制其表达。该浓度的雷公藤红素还可显著抑制 GATA3 和 NFAT1 的核转位，降低其与 IL-13 基因启动子的结合率。
雷公藤红素通过抑制 GATA3 和 NFAT1 的核转位及其与 IL-13 基因启动子区的结合率，抑制 IL-13 基因转录和蛋白表达。
