Joint inflammation is a key player in the pathogenesis of osteoarthritis (OA). Imperatorin, a plant-derived small molecule has been reported to have anti-inflammatory properties; however, its effect on chondrocytes is not known. Here, we investigated the effects of Imperatorin on interleukin-1β (IL-1β) induced expression of inducible nitric oxide synthase (iNOS) and nitric oxide production in primary human OA chondrocytes and cartilage explants culture under pathological conditions and explored the associated signaling pathways. We pretreated chondrocytes or explants with Imperatorin (50 μM) followed by IL-1β (1 ng/ml), and the culture supernatant was used to determine the levels of nitrite production by Griess assay and chondrocytes were harvested to prepare cell lysate or RNA for gene expression analysis of iNOS by Western blot or qPCR and in explants by immunohistochemistry (IHC). Pretreatment of primary chondrocytes and cartilage explants with Imperatorin suppressed IL-1β induced expression of iNOS and NO production. Imperatorin blocked the IL-1β-induced phosphorylation of ERK-MAPK/AP1 signaling pathway to suppress iNOS expression. The role of ERK in the regulation of iNOS expression was verified by using ERK inhibitor. Interestingly, we also found that Imperatorin binds to iNOS protein and inhibits its activity in vitro. Our data demonstrated that Imperatorin possess strong anti-inflammatory activity and may be developed as a therapeutic agent for the management of OA.

关节炎症是骨关节炎 (OA) 发病机制的关键因素。据报道，欧前胡素是一种植物来源的小分子，具有抗炎特性；然而，它对软骨细胞的影响尚不清楚。在这里，我们研究了欧前胡素对病理条件下原代人 OA 软骨细胞和软骨外植体培养物中白细胞介素 1β (IL-1β) 诱导的诱导型一氧化氮合酶 (iNOS) 表达和一氧化氮产生的影响，并探索了相关的信号通路。我们用欧前胡素 (50 μM) 和 IL-1β (1 ng/ml) 预处理软骨细胞或外植体，培养上清液用于通过 Griess 测定确定亚硝酸盐产生水平，收获软骨细胞以制备细胞裂解物或 RNA，用于通过蛋白质印迹或 qPCR 和免疫组织化学 (IHC) 在外植体中进行 iNOS 基因表达分析。用欧前胡素预处理原代软骨细胞和软骨外植体可抑制 IL-1β 诱导的 iNOS 表达和 NO 产生。欧前胡素阻断 IL-1β 诱导的 ERK-MAPK/AP1 信号通路磷酸化，从而抑制 iNOS 表达。通过使用ERK抑制剂验证了ERK在调节iNOS表达中的作用。有趣的是，我们还发现欧前胡素与 iNOS 蛋白结合并抑制其活性 用欧前胡素预处理原代软骨细胞和软骨外植体可抑制 IL-1β 诱导的 iNOS 表达和 NO 产生。欧前胡素阻断 IL-1β 诱导的 ERK-MAPK/AP1 信号通路磷酸化，从而抑制 iNOS 表达。通过使用ERK抑制剂验证了ERK在调节iNOS表达中的作用。有趣的是，我们还发现欧前胡素与 iNOS 蛋白结合并抑制其活性 用欧前胡素预处理原代软骨细胞和软骨外植体可抑制 IL-1β 诱导的 iNOS 表达和 NO 产生。欧前胡素阻断 IL-1β 诱导的 ERK-MAPK/AP1 信号通路磷酸化，从而抑制 iNOS 表达。通过使用ERK抑制剂验证了ERK在调节iNOS表达中的作用。有趣的是，我们还发现欧前胡素与 iNOS 蛋白结合并抑制其活性体外。我们的数据表明，欧前胡素具有很强的抗炎活性，可以开发为治疗 OA 的治疗剂。