Department of Molecular and Medical Pharmacology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan.
Department of Anesthesiology and Pain Relief Center, The University of Tokyo Hospital, Tokyo, Japan.
Int Immunopharmacol. 2018 Sep;62:120-131. doi: 10.1016/j.intimp.2018.06.021. Epub 2018 Jul 10.
Cilostazol, a selective inhibitor of phosphodiesterase type III with anti-platelet, anti-mitogenic, and vasodilating properties, is widely used to treat ischemic symptoms of peripheral vascular disease. Ample evidence has suggested that cilostazol also exhibits an anti-inflammatory effect, but its anti-inflammatory mechanism is not fully understood. Here, we showed that cilostazol specifically inhibited expression of cytokines, which are induced by nuclear factor-κB (NF-κB) activation, in RAW264.7 macrophage cells stimulated with different Toll-like receptor (TLR) ligands. Cilostazol was found to significantly reduce TLR-4 and TLR-3 ligands-stimulated NF-κB transcriptional activity, which was quantified by luciferase reporter assays. However, cilostazol was without effect on IκBα degradation and NF-κB p65 phosphorylation and nuclear translocation after challenge with the TLR-4 ligand lipopolysaccharide (LPS). Cilostazol did not also prevent the LPS-induced increase in phosphorylated levels of the mitogen-activated protein kinase (MAPK) family. On the other hand, using chromatin immunoprecipitation assays, we demonstrated that cilostazol reduced the LPS-induced transcriptional activities of interleukin-6 and tumor necrosis factor-α by preventing the recruitment of NF-κB p65 to these gene promoters. When cilostazol was given to mice by oral gavage daily for 7 days, LPS-induced aberrant pro-inflammatory cytokine production and end-organ tissue injury were significantly reduced. The results of this study suggest that cilostazol is capable of directly interrupting DNA binding activity of NF-κB proteins from the TLR signaling pathways. The therapy to specifically intervene in this pathway may be potentially beneficial for the prevention of different inflammatory disorders.
西洛他唑是一种磷酸二酯酶 III 型选择性抑制剂，具有抗血小板、抗有丝分裂和血管扩张作用，广泛用于治疗外周血管疾病的缺血症状。大量证据表明，西洛他唑还具有抗炎作用，但抗炎机制尚不完全清楚。在这里，我们表明西洛他唑特异性抑制了 RAW264.7 巨噬细胞中由核因子-κB（NF-κB）激活诱导的细胞因子表达，这些细胞因子受到不同 Toll 样受体（TLR）配体的刺激。西洛他唑被发现可显著降低 TLR-4 和 TLR-3 配体刺激的 NF-κB 转录活性，这通过荧光素酶报告基因测定进行定量。然而，西洛他唑在用 TLR-4 配体脂多糖（LPS）刺激后对 IκBα 降解和 NF-κB p65 磷酸化和核易位没有影响。西洛他唑也没有阻止 LPS 诱导的丝裂原激活蛋白激酶（MAPK）家族磷酸化水平的增加。另一方面，我们通过染色质免疫沉淀测定表明，西洛他唑通过阻止 NF-κB p65 募集到这些基因启动子，降低了 LPS 诱导的白细胞介素-6 和肿瘤坏死因子-α的转录活性。当西洛他唑通过口服灌胃每天给予小鼠 7 天时，LPS 诱导的异常促炎细胞因子产生和终末器官组织损伤明显减少。这项研究的结果表明，西洛他唑能够直接阻断 TLR 信号通路中 NF-κB 蛋白的 DNA 结合活性。针对该途径的治疗可能对预防不同的炎症性疾病具有潜在益处。
