【摘要】 目的:探讨脂多糖(lipopolysaccharide,LPS)联合真菌葡聚糖能否活化支气管上皮细胞16HBE内NOD样受体热蛋白结构域相关蛋白3(Nod-like receptor pyrin domain-containing protein 3,NLRP3)炎性小体,芍药苷(paeoniflorin,PF)对该NLRP3炎性小体的活化是否有抑制作用及相关机制。方法:LPS联合真菌葡聚糖建立感染模型,RT-PCR检测细胞内NLRP3、caspase-1、白细胞介素(interleukin,IL)-1βm RNA的表达;ELISA检测细胞上清中IL-1β含量;caspase-1活性检测试剂盒检测胞内caspase-1活化程度;流式检测胞内活性氧(reactive oxygen species,ROS)变化;Western blot检测胞内NLRP3、caspase-1、IL-1β蛋白表达变化。结果:LPS联合真菌葡聚糖联合作用于支气管上皮细胞,胞内NLRP3、caspase-1、IL-1β表达转录加强,细胞上清中IL-1β含量增加,caspase-1活性上调,细胞内ROS升高;PF预作用细胞后,胞内ROS随着药物浓度增加而逐渐降低,NLRP3、caspase-1、IL-1β的转录表达也随之受抑制而下调。结论:LPS联合真菌葡聚糖可有效活化支气管上皮细胞内NLRP3炎性小体,而PF能有效抑制胞内ROS产生从而抑制炎性小体活化、抑制真菌葡聚糖所致的炎性反应。更多还原

【Abstract】 Objective:To investigate if lipopolysaccharide(LPS) combined with fungal glucan(curdlan) activates Nod-like receptor pyrin domain-containing protein(NLRP3) inflammasome,and the activation could be influenced by paeoniflorin(PF) in human bronchial epithelial cells(16HBE),as well as the underlying mechanism. Methods:16HBE cells were stimulated by LPS combined with curdlan to establish inflammation models. The expressions of NLRP3,caspase-1,and interleukin(IL)-1β m RNA were analyzed by RT-PCR. The level of IL-1β in supernatants was evaluated by ELISA. The activity of caspase-1 was measured using a caspase-1activity kit. Reactive oxygen species(ROS) were detected by flow cytometry. The expressions of NLRP3,caspase-1,and IL-1β protein in cells were determined by Western blot. Results:The expressions of NLRP3,caspase-1,IL-1β m RNA and protein were up-regulated in 16 HBE cells activated by LPS combined with curdlan. The activity of caspase-1 and the levels of IL-1β in supernatants,as well as the concentrations of intracellular ROS were significantly increased; these effects were inhibited effectively by PF,especially in high concentration,through reducing the production of intracellular ROS. Conclusion:NLRP3 inflammasome plays an important role in combined LPS and curdlan infection in 16 HBE cells,and PF may possess a certain anti-fungal infection effect via inhibiting the activation of NLRP3 inflammasome.更多还原