基于BRD4/NF-κB/NLRP3通路介导的巨噬细胞焦亡探讨三石汤抑制痛风性关节炎的分子机制研究

Mechanism of Sanshi decoction inhibits macrophage pyroptosis by inhibiting BRD4/NF-κB/NLRP3 pathway in the treatment of gouty arthritis

目的:观察三石汤对BRD4/NF-κB/NLRP3通路介导的巨噬细胞焦亡的影响,从而阐明三石汤抑制痛风性关节炎的分子机制。方法:以佛波酯诱导THP-1人单核细胞株分化巨噬细胞,并分为空白组、模型组、三石汤低剂量、中剂量、高剂量和BRD4抑制剂组。除空白组外,余下各组以尿酸单钠结晶诱导构建痛风性关节炎细胞模型。分别采用CCK8检测巨噬细胞的活性,流式细胞术检测巨噬细胞焦亡水平,酶联免疫吸附法检测LDH的活性和IL-1β及IL-18的含量,Western blot检测BRD4/NF-κB/NLRP3通路中相关蛋白的表达。结果:与空白组比较,模型组巨噬细胞活性下降,焦亡水平、LDH活性、IL-1β与IL-18含量、BRD4、p-NF-κB p65、NLRP3、Caspase-1 p20和IL-1β蛋白的表达均显著的上调,差异均具有统计学意义(P<0.05)。与模型组相比,三石汤组巨噬细胞活性上调,焦亡水平、LDH活性、IL-1β与IL-18含量、BRD4、p-NF-κB p65、NLRP3、Caspase-1 p20和IL-1β蛋白表达均显著的下降,差异均具有统计学意义(P<0.05)。结论:三石汤通过抑制BRD4/NF-κB/NLRP3通路激活抑制巨噬细胞焦亡,从而改善痛风性关节炎的炎症水平。

Objective:To observe the effect of Sanshi decoction on BRD4/NF-κB/NLRP3 pathway-mediated macrophage pyroptosis,so as to elucidate the molecular mechanism of Sanshi decoction in the treatment of gouty arthritis.Methods:THP-1 was induced into macrophages with foboside and the divided into the control group,model group,low-dose,medium-dose,highdose group of Sanshi decoction,and BRD4 inhibitor group.Except for the control group,the remaining groups were induced with monosodium urate crystals to construct a gouty arthritis cell model.The activity of macrophages was detected by CCK8,the level of macrophage pyroptosis was detected by flow cytometry,the activity of LDH,the content of IL-1βand IL-18 were detected by enzyme-linked immunosorbent assay,and the expression of related proteins in the BRD4/NF-κB/NLRP3 pathway was detected by Western blot.Results:Compared with the control group,macrophage activity was decreased in the model group,and the level of pyroptosis,LDH activity,contents of IL-1βand IL-18,expression levels of BRD4,p-NF-κB p65,NLRP3,Caspase-1 p20,and IL-1βprotein were significantly up-regulated,the differences were statistically significant(P<0.05).Compared with the model group,macrophage activity was up-regulated in the Sanshi Decoction,and the level of pyroptosis,LDH activity,IL-1βand IL-18 contents,expression levels of BRD4,p-NF-κB p65,NLRP3,Caspase-1 p20,and IL-1βprotein were significantly decreased with statistically significant differences(P<0.05).Conclusion:Sanshi decoction inhibits macrophage pyroptosis by inhibiting BRD4/NF-κB/NLRP3 pathway activation,thus improving the inflammation level of gouty arthritis.