基于p38 MAPK信号通路分析槲皮素保护骨性关节炎关节软骨的机制

Mechanism of Quercetin in Protecting Articular Cartilage from Osteoarthritis Based on p38 MAPK Pathway

目的:探讨槲皮素通过靶向干预p38丝裂原活化蛋白激酶(p38 MAPK)信号通路保护膝关节骨性关节炎关节软骨的作用机制。方法:笔者通过网络药理学技术,科学的对槲皮素保护骨性关节炎患者关节软骨的靶点因子、信号通路进行预测,并对其进行分析。选择一条与骨性关节炎相关密切的预测通路对其采用体外细胞实验进行验证,通过细胞计数试剂-8(CCK-8)方法筛选出药物最佳干预浓度,采用酶联免疫吸附测定(ELISA)对骨性关节炎及密切相关炎性因子白细胞介素(IL)-1β,肿瘤坏死因子-α(TNF-α)进行检测,采用实时荧光定量聚合酶链式反应(Real-time PCR)和蛋白免疫印迹法(Western blot)检测槲皮素干预后p38 MAPK信号通路下的相关mRNA,蛋白的表达。结果:通过网络药理学预测MAPK信号通路与骨性关节炎患者(OA)存在十分密切的相关性,并选取该通路中最为密切的p38 MAPK信号通路进行研究,通过药物浓度筛查发现在100μmol·L^(-1)的槲皮素干预后相关炎性因子降低最为明显,并且,在该浓度的槲皮素干预下抑制p38 MAPK信号通路中p38,磷酸化(p)-p38,基质金属蛋白酶-13(MMP-13),解聚蛋白样金属蛋白酶-4(ADAMTS-4)等关键相关因子表达,促进Ⅱ型骨胶原蛋白(CollagenⅡ)的表达。结论:槲皮素在骨性关节炎疾病防治中具有降低软骨炎性因子表达的作用,可通过抑制p38MAPK通路相关因子表达而向好发展,所有结果显示,槲皮素具有降低OA炎性因子表达,保护OA患者关节软骨的作用。

Objective: To investigate the protective effect of quercetin(Qu)on articular cartilage of knee osteoarthritis and its mechanism by inhibiting p38 mitogen activated protein kinase(MAPK)signaling pathway. Method: Through the network pharmacology technology,we scientifically predicted and analyzed the target factors and signal pathways of Qu in the protection of articular cartilage in patients with osteoarthritis. We selected a prediction pathway closely related to osteoarthritis and validated it by cell experiment in vitro. The best intervention concentration of the drug was selected by cell counting kit-8(CCK-8)method. The osteoarthritis and its closely related inflammatory factors interleukin(IL)-1β and tumor necrosis factor(TNF)-α were detected by enzyme linked immunosorbent assay(ELISA). The expression of related mRNA and protein in p38 signal pathway after Qu intervention were detected by quantitative real time polymerase chain reaction(Real-time PCR)and Western blot. Result: It was predicted that MAPK signal pathway was closely related to osteoarthritis by network pharmacology,and p38 MAPK pathway,which was most closely related to osteoarthritis,was selected for study. The results showed that 100 μmol·L^(-1) Qu had the most obvious effect in decreasing the expression of related inflammatory factors,inhibited the expression of p38,phosphorylated(p)-p38,matrix metalloproteinase-13(MMP-13),A disintegrin-like and metalloproteinase with thrombospondin type-1 motifs-4(ADAMTS-4)in the pathway,and promoted the expression of Collagen Ⅱ. Conclusion: Qu could decrease the expression of cartilage inflammatory factors in the prevention and treatment of osteoarthritis,and the effect can be well developed by intervening and inhibiting p38 MAPK pathway related factor expression level. All the results show that Qu can decrease osteoarthritis inflammatory factors and protect articular cartilage in patients with osteoarthritis.