基于ROS/TXNIP/Nlrp3探讨芦荟大黄素对P.g-LPS诱导的小鼠牙周膜成纤维细胞的影响

Effect of aloe-emodin on mouse periodontal ligament fibroblasts by P.g-LPS stimulated based on ROS/TXNIP/NLRP3 signaling pathway

目的研究芦荟大黄素(aloe-emodin,AE)对牙龈卟啉单胞菌脂多糖(P.g-LPS)诱导的牙周膜成纤维细胞Nlrp3-inflammasome活化的影响。方法体外培养牙周膜成纤维细胞,采用P.g-LPS作为刺激剂,通过Western blot观察不同浓度芦荟大黄素(aloe-emodin, AE)对Nlrp3、前体caspase-1(pro-caspase-1)、活化caspase-1(cle-caspase-1)、TXNIP蛋白表达量的影响分子活化程度,利用RT-qPCR方法检测其对前体caspase-1、Nlrp3转录水平的影响,并使用荧光探针法评价对ROS生成量的影响。结果与空白对照相比,给予P.g-LPS刺激的小鼠牙周膜成纤维细胞Nlrp3、前体caspase-1、TXNIP的蛋白表达量及ROS的生成量明显增高,活化caspase-1的生成明显增多,AE可以明显降低Nlrp3、前体caspase-1,活化caspase-1,TXNIP在细胞内的蛋白表达量,同时抑制Nlrp3和前体caspase-1的转录水平并且减少ROS在细胞内的生成量。结论 AE能通过抑制Nlrp3相关蛋白的表达及减弱ROS/TXNIP诱导的Nlrp3-inflammasome聚集明显抑制P.g-LPS诱导的小鼠牙周膜成纤维细胞Nlrp3-inflammasome的活化。

To investigate the protective effect of aloe-emodin(AE)on mouse periodontal ligament fibroblasts(mPDLFs)induced by P.g-LPS of Porphyromonas gingivalis and the relevant mechanism.Methods P.g-LPS was used as the stimuli to induce mPDLFs and Western blotting was employed to investigate the effect of AE on the expression of NLRP3,Pro-caspase-1,Cle-caspase-1 and TXNIP in P.g-LPS-treated mPDLFs.The effects of AE on the transcription levels of Pro-caspase-1 and NLRP3 were further detected by RT-qPCR.In addition,chemical fluorescence was used to investigate the impact of AE on production of ROS in mPDLFs treated by P.g-LPS.Results Compared with control group,P.g-LPS increased the expression of NLRP3,Pro-caspase-1,Cle-caspase-1,TXNIP and the transcription levels of NLRP3 and Pro-caspase-1 and the production of ROS in mPDLFs.AE could decrease the expression of NLRP3,Pro-caspase-1 and Cle-caspase-1 in varying degrees and inhibited the transcription of NLRP3 and Pro-caspase-1.Meanwhile,the production of ROS and the expression of TXNIP were also reduced by AE.Conclusions AE may inhibit the activation of NLRP3 inflammasome in P.g-LPS treated mPDLFs by reducing NLRP3 related protein formation and NLRP3 inflammasome gathering through ROS/TXNIP.