基于P2X7R/TLR4/NF-κB/NLRP3轴探讨清解化攻方对重症急性胰腺炎炎症反应的作用机制

Exploration on the mechanism of action of Qingjie Huagong Decoction on the inflammatory response of severe acute pancreatitis based on the P2X7R/TLR4/NF-κB/NLRP3 axis

目的:研究P2X7R/TLR4/NF-κB/NLRP3轴在重症急性胰腺炎(SAP)炎症反应的作用机制及清解化攻方的干预作用。方法:采用雨娃素联合脂多糖建立SAP大鼠模型,分别设置空白组、模型组、不同剂量清解化攻方给药组和阳性对照组,通过HE染色观察胰腺组织病理,ELISA检测炎症指标,结合Western Blot和qRT-PCR检测胰腺组织中P2X7R、TLR4、NF-κB、NLRP3蛋白和mRNA的表达情况。结果:与空白组比较,模型组胰腺组织水肿、坏死出血,大鼠血清中IL-8、IL-12、IL-17、IL-18的含量明显升高(P<0.05),胰腺组织中P2X7R、TLR4、NF-κB、NLRP3蛋白和mRNA的表达量显著升高(P<0.05);与模型组比较,清解化攻方各剂量组和阳性对照组能够改善SAP胰腺组织水肿,减少坏死出血,可降低SAP模型大鼠血清中IL-8、IL-12、IL-17、IL-18的含量(P<0.05),中、高剂量组和阳性对照组胰腺组织中P2X7R、TLR4、NF-κB、NLRP3蛋白和mRNA表达量显著降低(P<0.05)。结论:清解化攻方可能通过下调P2X7R/TLR4/NF-κB/NLRP3信号通路的表达,从而控制SAP炎症反应,起到治疗SAP的作用。

Objective:To study the mechanism of P2X7R/TLR4/NF-κB/NLRP3 axis in the inflammatory response of severe acute pancreatitis(SAP)and the interventional effect of Qingjie Huagong Decoction(QJHGD).Methods:A SAP rat model was established by using cerulein combined with lipopolysaccharide.A blank group,a model group,a group administered with different dosages of QJHGD,and a positive control group were set up,and the histopathology of the pancreas was observed by HE staining,the inflammation indexes were detected by ELISA,and the expressions of P2X7R,TLR4,NF-κB and NLRP3 proteins and mRNAs were detected by combining with Western Blot and qRT-PCR in pancreatic tissues.Results:Compared with the blank group,pancreatic tissue edema,necrotic hemorrhage,serum levels of IL-8,IL-12,IL-17 and IL-18 in the model group were significantly higher(P<0.05),and the expression of P2X7R,TLR4,NF-κB,NLRP3 proteins and mRNAs in pancreatic tissues were significantly higher(P<0.05).Compared with the model group,the QJHGD dose groups and positive control group could improve the edema of SAP pancreatic tissues,reduce necrotic hemorrhage,and could reduce the serum levels of IL-8,IL-12,IL-17 and IL-18 in SAP model rats(P<0.05),and the expressions of P2X7R,TLR4,NF-κB,NLRP3 protein and mRNA in pancreatic tissues of medium and high dose groups and positive control group were decreased(P<0.05).Conclusion:The QJHGD may play a role in the treatment of SAP by down-regulating the expression of the P2X7R/TLR4/NF-κB/NLRP3 signaling pathway,thereby controlling the inflammatory response to SAP.