Department of Pulmonary Medicine, Faculty of Medicine, University of Tsukuba, Ibaraki, Japan.
Respiratory Disease Center, Showa University Northern Yokohama Hospital, Kanagawa, Japan.
Int Arch Allergy Immunol. 2018;176(2):83-90. doi: 10.1159/000488154. Epub 2018 Apr 12.
IL-17F is involved in the pathogenesis of several inflammatory diseases, including asthma and COPD. However, the effects of steroids on the function of IL-17F signaling mechanisms are largely unknown. One of the transcription elongation factors, positive transcription elongation factor b (P-TEFb) composed of cyclin T1 and cyclin-dependent kinase 9 (CDK9), is known as a novel checkpoint regulator of gene expression via bromodomain-containing protein 4 (Brd4).
Human airway smooth muscle cells were stimulated with IL-17F and the expression of IL-8 was evaluated by real-time PCR and ELISA. Next, the phosphorylation of CDK9 was determined by Western blotting. The CDK9 inhibitor and short interfering RNAs (siRNAs) targeting Brd4, cyclin T1, and CDK9 were used to identify the effect on IL-17F-induced IL-8 expression. Finally, the effect of steroids and its signaling were evaluated.
IL-17F markedly induced the transcription of the IL-8 gene and the expression of the protein. Pretreatment of CDK9 inhibitor and transfection of siRNAs targeting CDK9 markedly abrogated IL-17F-induced IL-8 production. Transfection of siRNAs targeting Brd4 and cyclin T1 diminished IL-17F-induced phosphorylation of CDK9 and IL-8 production. Moreover, budesonide decreased CDK9 phosphorylation and markedly inhibited IL-17F-induced IL-8 production.
This is the first report that P-TEFb is involved in IL-17F-induced IL-8 expression and that steroids diminish it via the inhibition of CDK9 phosphorylation. IL-17F and P-TEFb might be novel therapeutic targets for airway inflammatory diseases.
IL-17F 参与多种炎症性疾病的发病机制，包括哮喘和 COPD。然而，类固醇对 IL-17F 信号转导机制功能的影响在很大程度上尚不清楚。转录延伸因子之一，由细胞周期蛋白 T1 和细胞周期蛋白依赖性激酶 9 (CDK9) 组成的正转录延伸因子 b (P-TEFb)，已知是通过溴结构域蛋白 4 (Brd4) 作为基因表达的新型检查点调节剂。
用 IL-17F 刺激人气道平滑肌细胞，通过实时 PCR 和 ELISA 评估 IL-8 的表达。接下来，通过 Western blot 测定 CDK9 的磷酸化。使用 CDK9 抑制剂和靶向 Brd4、细胞周期蛋白 T1 和 CDK9 的短发夹 RNA (siRNA) 来确定对 IL-17F 诱导的 IL-8 表达的影响。最后，评估类固醇及其信号的作用。
IL-17F 显著诱导 IL-8 基因的转录和蛋白的表达。CDK9 抑制剂预处理和靶向 CDK9 的 siRNA 转染显著减弱了 IL-17F 诱导的 IL-8 产生。靶向 Brd4 和细胞周期蛋白 T1 的 siRNA 转染减弱了 IL-17F 诱导的 CDK9 磷酸化和 IL-8 的产生。此外，布地奈德降低了 CDK9 的磷酸化，并显著抑制了 IL-17F 诱导的 IL-8 产生。
这是第一项表明 P-TEFb 参与 IL-17F 诱导的 IL-8 表达，并且类固醇通过抑制 CDK9 磷酸化来减弱其表达的报告。IL-17F 和 P-TEFb 可能是气道炎症性疾病的新的治疗靶点。
