芍药苷通过抑制NLRP3炎症小体活化减轻病毒性心肌炎小鼠心肌损伤

Paeoniflorin alleviated myocardial injury in mice with viral myocarditis by inhibiting NLRP3 inflammasome activation

目的:探讨芍药苷(PF)对柯萨奇B3病毒(CVB3)诱导的病毒性心肌炎(VMC)小鼠心肌损伤的影响及其可能的作用机制。方法:将60只雄性BALB/c小鼠随机分为空白对照组(blank)、VMC模型组(VMC)、PF低剂量组(PF-L,10 mg/kg)、PF高剂量组(PF-H,40 mg/kg)和高剂量PF联合NLRP3激动剂组(PF-H+MSU,40 mg/kg PF+5 mg/kg MSU),每组12只,采用腹腔注射CVB3病毒溶液建立VMC小鼠模型,次日腹腔注射给药,1次/d,连续7 d。HE染色观察各组小鼠心肌损伤病理变化;TUNEL染色观察心肌组织细胞凋亡;ELISA检测血清中心肌损伤标志物心肌肌钙蛋白Ⅰ(cTnⅠ)、肌酸激酶同工酶(CK-MB)、肌红蛋白(Mb)以及心肌组织中IL-1β、IL-18和TNF-α含量;Western blot检测心肌组织中Cleaved caspase-3、NLRP3、ASC和caspase-1蛋白表达水平。结果:与blank组比较,VMC组小鼠心肌损伤严重,心肌组织细胞凋亡显著增加(P<0.05),血清中cTnⅠ、CK-MB、Mb含量和心肌组织中IL-1β、IL-18、TNF-α含量均显著上升(P<0.05),同时心肌组织中Cleaved caspase-3、NLRP3、ASC和caspase-1蛋白表达显著增加(P<0.05);与VMC组比较,PF-H组小鼠心肌损伤有所改善,心肌组织细胞凋亡水平降低(P<0.05),血清中cTnⅠ、CK-MB、Mb含量和心肌组织中IL-1β、IL-18、TNF-α含量均显著下降(P<0.05),同时心肌组织中Cleaved caspase-3、NLRP3、ASC和caspase-1蛋白表达水平显著降低(P<0.05);而PF-L组与VMC组比较,相关指标差异均无统计学意义(P>0.05);然而,联合NLRP3激动剂MSU干预可显著抑制高剂量PF对VMC模型小鼠的改善作用。结论:PF能有效改善VMC模型小鼠的心肌损伤,其作用机制可能与NLRP3炎症小体通路的活性抑制有关。

Objective:To investigate the effect of paeoniflorin(PF)on myocardial injury in mice with coxsackievirus B3(CVB3)-induced viral myocarditis(VMC)and its possible mechanism.Methods:Sixty male BALB/c mice were randomly divided into blank control group(blank),VMC model group(VMC),PF low-dose group(PF-L,10 mg/kg),PF high-dose group(PF-H,40 mg/kg),and high-dose PF combined with NLRP3 agonist group(PF-H+MSU,40 mg/kg PF+5 mg/kg MSU),12 mice in each group.The mouse model of VMC was established by intraperitoneal injection of CVB3 virus solution,and the drug was intraperitoneally injected the next day,once a day for 7 d.HE staining was used to observe the pathological changes of myocardial injury in each group,and TUNEL staining was used to observe the apoptosis of myocardial tissue cells.Serum myocardial injury markers cardiac troponinⅠ(cTnⅠ),creatine kinase-MB(CK-MB),myoglobin(Mb),and the contents of IL-1β,IL-18,TNF-αin myocardial tissue were detected by ELISA,and the protein expression levels of Cleaved caspase-3,NLRP3,ASC,and caspase-1 in myocardial tissue were detected by Western blot.Results:Compared with the blank group,the myocardial injury was destroyed in VMC mice,the apoptosis of myocardial tissue cells were significantly increased(P<0.05),the contents of cTnⅠ,CK-MB,Mb in serum,as the well as the contents of IL-1β,IL-18,TNF-αin myocardial tissue were significantly increased(P<0.05),and the protein expression of Cleaved caspase-3,NLRP3,ASC and caspase-1 in the myocardial tissue were significantly increased(P<0.05).Compared with the VMC group,the myocardial injury in PF-H group was apparently improved,the contents of cTnⅠ,CK-MB,Mb in serum,as the well as the contents of IL-1β,IL-18,TNF-αin myocardial tissue were significantly decreased(P<0.05),and the protein expression levels of Cleaved caspase-3,NLRP3,ASC and caspase-1 in the myocardial tissue were significantly decreased(P<0.05).There was no significant difference between PF-L group and VMC group(P>0.05).However,combined NLRP3 agonist MSU intervention significantly inhibited the improvement of high dose PF in VMC model mice.Conclusion:PF can effectively improve the myocardial injury in mice with VMC,and the mechanism of action may be related to the inhibition of NLRP3 inflammasome pathway.